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  • Protein Binding  (52)
  • Nature Publishing Group (NPG)  (52)
  • American Institute of Physics (AIP)
  • 2010-2014  (52)
  • 2000-2004
  • 1990-1994
  • 1985-1989
  • 2013  (52)
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  • 2010-2014  (52)
  • 2000-2004
  • 1990-1994
  • 1985-1989
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  • 1
    Publication Date: 2013-03-29
    Description: Chromosomal replication machines contain coupled DNA polymerases that simultaneously replicate the leading and lagging strands. However, coupled replication presents a largely unrecognized topological problem. Because DNA polymerase must travel a helical path during synthesis, the physical connection between leading- and lagging-strand polymerases causes the daughter strands to entwine, or produces extensive build-up of negative supercoils in the newly synthesized DNA. How DNA polymerases maintain their connection during coupled replication despite these topological challenges is unknown. Here we examine the dynamics of the Escherichia coli replisome, using ensemble and single-molecule methods, and show that the replisome may solve the topological problem independent of topoisomerases. We find that the lagging-strand polymerase frequently releases from an Okazaki fragment before completion, leaving single-strand gaps behind. Dissociation of the polymerase does not result in loss from the replisome because of its contact with the leading-strand polymerase. This behaviour, referred to as 'signal release', had been thought to require a protein, possibly primase, to pry polymerase from incompletely extended DNA fragments. However, we observe that signal release is independent of primase and does not seem to require a protein trigger at all. Instead, the lagging-strand polymerase is simply less processive in the context of a replisome. Interestingly, when the lagging-strand polymerase is supplied with primed DNA in trans, uncoupling it from the fork, high processivity is restored. Hence, we propose that coupled polymerases introduce topological changes, possibly by accumulation of superhelical tension in the newly synthesized DNA, that cause lower processivity and transient lagging-strand polymerase dissociation from DNA.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3618558/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3618558/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Kurth, Isabel -- Georgescu, Roxana E -- O'Donnell, Mike E -- GM38839/GM/NIGMS NIH HHS/ -- R01 GM038839/GM/NIGMS NIH HHS/ -- Howard Hughes Medical Institute/ -- England -- Nature. 2013 Apr 4;496(7443):119-22. doi: 10.1038/nature11988. Epub 2013 Mar 27.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉The Rockefeller University, Howard Hughes Medical Institute, 1230 York Avenue, New York, New York 10065, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23535600" target="_blank"〉PubMed〈/a〉
    Keywords: DNA/chemistry/genetics/metabolism ; DNA Primase/metabolism ; *DNA Replication ; DNA, Bacterial/biosynthesis/chemistry/genetics/*metabolism ; DNA, Superhelical/biosynthesis/chemistry/genetics/metabolism ; DNA-Binding Proteins/chemistry/metabolism ; DNA-Directed DNA Polymerase/chemistry/*metabolism ; Escherichia coli/*enzymology/*genetics ; Microscopy, Fluorescence ; Multienzyme Complexes/chemistry/*metabolism ; *Nucleic Acid Conformation ; Protein Binding
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  • 2
    Publication Date: 2013-10-15
    Description: The mechanisms by which genetic variation affects transcription regulation and phenotypes at the nucleotide level are incompletely understood. Here we use natural genetic variation as an in vivo mutagenesis screen to assess the genome-wide effects of sequence variation on lineage-determining and signal-specific transcription factor binding, epigenomics and transcriptional outcomes in primary macrophages from different mouse strains. We find substantial genetic evidence to support the concept that lineage-determining transcription factors define epigenetic and transcriptomic states by selecting enhancer-like regions in the genome in a collaborative fashion and facilitating binding of signal-dependent factors. This hierarchical model of transcription factor function suggests that limited sets of genomic data for lineage-determining transcription factors and informative histone modifications can be used for the prioritization of disease-associated regulatory variants.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3994126/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3994126/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Heinz, S -- Romanoski, C E -- Benner, C -- Allison, K A -- Kaikkonen, M U -- Orozco, L D -- Glass, C K -- 5T32DK007494/DK/NIDDK NIH HHS/ -- CA17390/CA/NCI NIH HHS/ -- DK063491/DK/NIDDK NIH HHS/ -- DK091183/DK/NIDDK NIH HHS/ -- P01 DK074868/DK/NIDDK NIH HHS/ -- P30 CA023100/CA/NCI NIH HHS/ -- P30 DK063491/DK/NIDDK NIH HHS/ -- R01 CA173903/CA/NCI NIH HHS/ -- R01 DK091183/DK/NIDDK NIH HHS/ -- T32 AR059033/AR/NIAMS NIH HHS/ -- England -- Nature. 2013 Nov 28;503(7477):487-92. doi: 10.1038/nature12615. Epub 2013 Oct 13.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉1] Department of Cellular and Molecular Medicine, University of California, San Diego, 9500 Gilman Drive, Mail Code 0651, La Jolla, California 92093, USA [2].〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/24121437" target="_blank"〉PubMed〈/a〉
    Keywords: Amino Acid Motifs/genetics ; Animals ; Base Sequence ; Cell Lineage/genetics ; DNA-Binding Proteins/metabolism ; Enhancer Elements, Genetic/*genetics ; Gene Expression Regulation/*genetics ; Genetic Variation/*genetics ; Histones/chemistry/metabolism ; Macrophages/metabolism ; Male ; Mice ; Mice, Inbred BALB C ; Mice, Inbred C57BL ; Models, Biological ; Mutation/genetics ; NF-kappa B/metabolism ; Protein Binding ; Reproducibility of Results ; Selection, Genetic/*genetics ; Transcription Factor RelA/metabolism ; Transcription Factors/*metabolism
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  • 3
    Publication Date: 2013-03-05
    Description: The contraction and relaxation of muscle cells is controlled by the successive rise and fall of cytosolic Ca(2+), initiated by the release of Ca(2+) from the sarcoplasmic reticulum and terminated by re-sequestration of Ca(2+) into the sarcoplasmic reticulum as the main mechanism of Ca(2+) removal. Re-sequestration requires active transport and is catalysed by the sarcoplasmic reticulum Ca(2+)-ATPase (SERCA), which has a key role in defining the contractile properties of skeletal and heart muscle tissue. The activity of SERCA is regulated by two small, homologous membrane proteins called phospholamban (PLB, also known as PLN) and sarcolipin (SLN). Detailed structural information explaining this regulatory mechanism has been lacking, and the structural features defining the pathway through which cytoplasmic Ca(2+) enters the intramembranous binding sites of SERCA have remained unknown. Here we report the crystal structure of rabbit SERCA1a (also known as ATP2A1) in complex with SLN at 3.1 A resolution. The regulatory SLN traps the Ca(2+)-ATPase in a previously undescribed E1 state, with exposure of the Ca(2+) sites through an open cytoplasmic pathway stabilized by Mg(2+). The structure suggests a mechanism for selective Ca(2+) loading and activation of SERCA, and provides new insight into how SLN and PLB inhibition arises from stabilization of this E1 intermediate state without bound Ca(2+). These findings may prove useful in studying how autoinhibitory domains of other ion pumps modulate transport across biological membranes.〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Winther, Anne-Marie L -- Bublitz, Maike -- Karlsen, Jesper L -- Moller, Jesper V -- Hansen, John B -- Nissen, Poul -- Buch-Pedersen, Morten J -- England -- Nature. 2013 Mar 14;495(7440):265-9. doi: 10.1038/nature11900. Epub 2013 Mar 3.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Pcovery, Thorvaldsensvej 57, DK-1871 Frederiksberg, Denmark.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23455424" target="_blank"〉PubMed〈/a〉
    Keywords: Animals ; Binding Sites ; Calcium/*metabolism ; Calcium-Binding Proteins/chemistry/metabolism ; Crystallography, X-Ray ; Cytoplasm/*metabolism ; Enzyme Activation ; Magnesium/metabolism ; Models, Molecular ; Muscle Proteins/chemistry/*metabolism ; Phosphorylation ; Protein Binding ; Proteolipids/chemistry/*metabolism ; Rabbits ; Sarcoplasmic Reticulum Calcium-Transporting ATPases/*chemistry/*metabolism
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  • 4
    Publication Date: 2013-10-15
    Description: The design of G-protein-coupled receptor (GPCR) allosteric modulators, an active area of modern pharmaceutical research, has proved challenging because neither the binding modes nor the molecular mechanisms of such drugs are known. Here we determine binding sites, bound conformations and specific drug-receptor interactions for several allosteric modulators of the M2 muscarinic acetylcholine receptor (M2 receptor), a prototypical family A GPCR, using atomic-level simulations in which the modulators spontaneously associate with the receptor. Despite substantial structural diversity, all modulators form cation-pi interactions with clusters of aromatic residues in the receptor extracellular vestibule, approximately 15 A from the classical, 'orthosteric' ligand-binding site. We validate the observed modulator binding modes through radioligand binding experiments on receptor mutants designed, on the basis of our simulations, either to increase or to decrease modulator affinity. Simulations also revealed mechanisms that contribute to positive and negative allosteric modulation of classical ligand binding, including coupled conformational changes of the two binding sites and electrostatic interactions between ligands in these sites. These observations enabled the design of chemical modifications that substantially alter a modulator's allosteric effects. Our findings thus provide a structural basis for the rational design of allosteric modulators targeting muscarinic and possibly other GPCRs.〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Dror, Ron O -- Green, Hillary F -- Valant, Celine -- Borhani, David W -- Valcourt, James R -- Pan, Albert C -- Arlow, Daniel H -- Canals, Meritxell -- Lane, J Robert -- Rahmani, Raphael -- Baell, Jonathan B -- Sexton, Patrick M -- Christopoulos, Arthur -- Shaw, David E -- England -- Nature. 2013 Nov 14;503(7475):295-9. doi: 10.1038/nature12595. Epub 2013 Oct 13.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉1] D. E. Shaw Research, 120 West 45th Street, 39th Floor, New York, New York 10036, USA [2].〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/24121438" target="_blank"〉PubMed〈/a〉
    Keywords: Allosteric Regulation/physiology ; Animals ; Binding Sites ; CHO Cells ; Cricetulus ; *Drug Design ; Humans ; Models, Chemical ; Molecular Conformation ; Molecular Dynamics Simulation ; Mutation ; Protein Binding ; Receptors, G-Protein-Coupled/*antagonists & inhibitors/*chemistry/genetics ; Reproducibility of Results
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  • 5
    Publication Date: 2013-07-19
    Description: Structural analysis of class B G-protein-coupled receptors (GPCRs), cell-surface proteins that respond to peptide hormones, has been restricted to the amino-terminal extracellular domain, thus providing little understanding of the membrane-spanning signal transduction domain. The corticotropin-releasing factor receptor type 1 is a class B receptor which mediates the response to stress and has been considered a drug target for depression and anxiety. Here we report the crystal structure of the transmembrane domain of the human corticotropin-releasing factor receptor type 1 in complex with the small-molecule antagonist CP-376395. The structure provides detailed insight into the architecture of class B receptors. Atomic details of the interactions of the receptor with the non-peptide ligand that binds deep within the receptor are described. This structure provides a model for all class B GPCRs and may aid in the design of new small-molecule drugs for diseases of brain and metabolism.〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Hollenstein, Kaspar -- Kean, James -- Bortolato, Andrea -- Cheng, Robert K Y -- Dore, Andrew S -- Jazayeri, Ali -- Cooke, Robert M -- Weir, Malcolm -- Marshall, Fiona H -- England -- Nature. 2013 Jul 25;499(7459):438-43. doi: 10.1038/nature12357. Epub 2013 Jul 17.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Heptares Therapeutics Ltd, BioPark, Broadwater Road, Welwyn Garden City AL7 3AX, UK.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23863939" target="_blank"〉PubMed〈/a〉
    Keywords: Amino Acid Motifs ; Amino Acid Sequence ; Aminopyridines/chemistry/metabolism/pharmacology ; Binding Sites ; Conserved Sequence ; Crystallography, X-Ray ; HEK293 Cells ; Humans ; Ligands ; Models, Molecular ; Molecular Sequence Data ; Protein Binding ; Protein Structure, Tertiary ; Receptors, Corticotropin-Releasing Hormone/antagonists & ; inhibitors/*chemistry/*classification/metabolism ; Receptors, Dopamine D3/antagonists & inhibitors/chemistry/classification
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  • 6
    Publication Date: 2013-10-25
    Description: Protein biosynthesis depends on the availability of ribosomes, which in turn relies on ribosomal RNA production. In eukaryotes, this process is carried out by RNA polymerase I (Pol I), a 14-subunit enzyme, the activity of which is a major determinant of cell growth. Here we present the crystal structure of Pol I from Saccharomyces cerevisiae at 3.0 A resolution. The Pol I structure shows a compact core with a wide DNA-binding cleft and a tightly anchored stalk. An extended loop mimics the DNA backbone in the cleft and may be involved in regulating Pol I transcription. Subunit A12.2 extends from the A190 jaw to the active site and inserts a transcription elongation factor TFIIS-like zinc ribbon into the nucleotide triphosphate entry pore, providing insight into the role of A12.2 in RNA cleavage and Pol I insensitivity to alpha-amanitin. The A49-A34.5 heterodimer embraces subunit A135 through extended arms, thereby contacting and potentially regulating subunit A12.2.〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Fernandez-Tornero, Carlos -- Moreno-Morcillo, Maria -- Rashid, Umar J -- Taylor, Nicholas M I -- Ruiz, Federico M -- Gruene, Tim -- Legrand, Pierre -- Steuerwald, Ulrich -- Muller, Christoph W -- England -- Nature. 2013 Oct 31;502(7473):644-9. doi: 10.1038/nature12636. Epub 2013 Oct 23.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉1] Centro de Investigaciones Biologicas, Consejo Superior de Investigaciones Cientificas, Ramiro de Maeztu 9, 28040 Madrid, Spain [2].〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/24153184" target="_blank"〉PubMed〈/a〉
    Keywords: Catalytic Domain ; Crystallography, X-Ray ; DNA/chemistry/metabolism ; Models, Molecular ; Peptide Chain Elongation, Translational ; Protein Binding ; Protein Conformation ; Protein Multimerization ; Protein Subunits/*chemistry ; RNA Polymerase I/*chemistry ; RNA Polymerase II/chemistry ; RNA Polymerase III/chemistry ; Saccharomyces cerevisiae/*enzymology ; Transcription, Genetic
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  • 7
    Publication Date: 2013-05-24
    Description: Facilitates chromatin transcription (FACT) is a conserved histone chaperone that reorganizes nucleosomes and ensures chromatin integrity during DNA transcription, replication and repair. Key to the broad functions of FACT is its recognition of histones H2A-H2B (ref. 2). However, the structural basis for how histones H2A-H2B are recognized and how this integrates with the other functions of FACT, including the recognition of histones H3-H4 and other nuclear factors, is unknown. Here we reveal the crystal structure of the evolutionarily conserved FACT chaperone domain Spt16M from Chaetomium thermophilum, in complex with the H2A-H2B heterodimer. A novel 'U-turn' motif scaffolded onto a Rtt106-like module embraces the alpha1 helix of H2B. Biochemical and in vivo assays validate the structure and dissect the contribution of histone tails and H3-H4 towards Spt16M binding. Furthermore, we report the structure of the FACT heterodimerization domain that connects FACT to replicative polymerases. Our results show that Spt16M makes several interactions with histones, which we suggest allow the module to invade the nucleosome gradually and block the strongest interaction of H2B with DNA. FACT would thus enhance 'nucleosome breathing' by re-organizing the first 30 base pairs of nucleosomal histone-DNA contacts. Our snapshot of the engagement of the chaperone with H2A-H2B and the structures of all globular FACT domains enable the high-resolution analysis of the vital chaperoning functions of FACT, shedding light on how the complex promotes the activity of enzymes that require nucleosome reorganization.〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Hondele, Maria -- Stuwe, Tobias -- Hassler, Markus -- Halbach, Felix -- Bowman, Andrew -- Zhang, Elisa T -- Nijmeijer, Bianca -- Kotthoff, Christiane -- Rybin, Vladimir -- Amlacher, Stefan -- Hurt, Ed -- Ladurner, Andreas G -- Howard Hughes Medical Institute/ -- England -- Nature. 2013 Jul 4;499(7456):111-4. doi: 10.1038/nature12242. Epub 2013 May 22.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Department of Physiological Chemistry, Butenandt Institute and LMU Biomedical Center, Faculty of Medicine, Ludwig Maximilians University of Munich, Butenandtstrasse 5, 81377 Munich, Germany.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23698368" target="_blank"〉PubMed〈/a〉
    Keywords: Amino Acid Motifs ; Chaetomium/*chemistry ; Conserved Sequence ; Crystallography, X-Ray ; DNA/chemistry/metabolism ; DNA Replication ; Fungal Proteins/*chemistry/*metabolism ; Histones/chemistry/*metabolism ; Hydrophobic and Hydrophilic Interactions ; Models, Molecular ; Molecular Chaperones/*chemistry/*metabolism ; Nucleosomes/chemistry/metabolism ; Protein Binding ; Protein Multimerization ; Protein Structure, Secondary ; Protein Structure, Tertiary ; Substrate Specificity
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  • 8
    Publication Date: 2013-07-31
    Description: In Gram-positive bacteria, T-box riboswitches regulate the expression of aminoacyl-tRNA synthetases and other proteins in response to fluctuating transfer RNA aminoacylation levels under various nutritional states. T-boxes reside in the 5'-untranslated regions of the messenger RNAs they regulate, and consist of two conserved domains. Stem I contains the specifier trinucleotide that base pairs with the anticodon of cognate tRNA. 3' to stem I is the antiterminator domain, which base pairs with the tRNA acceptor end and evaluates its aminoacylation state. Despite high phylogenetic conservation and widespread occurrence in pathogens, the structural basis of tRNA recognition by this riboswitch remains ill defined. Here we demonstrate that the ~100-nucleotide T-box stem I is necessary and sufficient for specific, high-affinity (dissociation constant (Kd) ~150 nM) tRNA binding, and report the structure of Oceanobacillus iheyensis glyQ stem I in complex with its cognate tRNA at 3.2 A resolution. Stem I recognizes the overall architecture of tRNA in addition to its anticodon, something accomplished by large ribonucleoproteins such as the ribosome, or proteins such as aminoacyl-tRNA synthetases, but is unprecedented for a compact mRNA domain. The C-shaped stem I cradles the L-shaped tRNA, forming an extended (1,604 A(2)) intermolecular interface. In addition to the specifier-anticodon interaction, two interdigitated T-loops near the apex of stem I stack on the tRNA elbow in a manner analogous to those of the J11/12-J12/11 motif of RNase P and the L1 stalk of the ribosomal E-site. Because these ribonucleoproteins and T-boxes are unrelated, this strategy to recognize a universal tRNA feature probably evolved convergently. Mutually induced fit of stem I and the tRNA exploiting the intrinsic flexibility of tRNA and its conserved post-transcriptional modifications results in high shape complementarity, which in addition to providing specificity and affinity, globally organizes the T-box to orchestrate tRNA-dependent transcription regulation.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3808885/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3808885/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Zhang, Jinwei -- Ferre-D'Amare, Adrian R -- Z99 HL999999/Intramural NIH HHS/ -- ZIA HL006102-02/Intramural NIH HHS/ -- ZIA HL006150-01/Intramural NIH HHS/ -- England -- Nature. 2013 Aug 15;500(7462):363-6. doi: 10.1038/nature12440. Epub 2013 Jul 28.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉National Heart, Lung and Blood Institute, 50 South Drive, MSC 8012, Bethesda, Maryland 20892-8012, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23892783" target="_blank"〉PubMed〈/a〉
    Keywords: Bacillaceae/*chemistry ; Bacterial Proteins/*chemistry ; *Models, Molecular ; Protein Binding ; Protein Structure, Quaternary ; RNA, Transfer/*chemistry ; *Riboswitch ; T-Box Domain Proteins/*chemistry
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  • 9
    Publication Date: 2013-06-14
    Description: 53BP1 (also called TP53BP1) is a chromatin-associated factor that promotes immunoglobulin class switching and DNA double-strand-break (DSB) repair by non-homologous end joining. To accomplish its function in DNA repair, 53BP1 accumulates at DSB sites downstream of the RNF168 ubiquitin ligase. How ubiquitin recruits 53BP1 to break sites remains unknown as its relocalization involves recognition of histone H4 Lys 20 (H4K20) methylation by its Tudor domain. Here we elucidate how vertebrate 53BP1 is recruited to the chromatin that flanks DSB sites. We show that 53BP1 recognizes mononucleosomes containing dimethylated H4K20 (H4K20me2) and H2A ubiquitinated on Lys 15 (H2AK15ub), the latter being a product of RNF168 action on chromatin. 53BP1 binds to nucleosomes minimally as a dimer using its previously characterized methyl-lysine-binding Tudor domain and a carboxy-terminal extension, termed the ubiquitination-dependent recruitment (UDR) motif, which interacts with the epitope formed by H2AK15ub and its surrounding residues on the H2A tail. 53BP1 is therefore a bivalent histone modification reader that recognizes a histone 'code' produced by DSB signalling.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3955401/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3955401/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Fradet-Turcotte, Amelie -- Canny, Marella D -- Escribano-Diaz, Cristina -- Orthwein, Alexandre -- Leung, Charles C Y -- Huang, Hao -- Landry, Marie-Claude -- Kitevski-LeBlanc, Julianne -- Noordermeer, Sylvie M -- Sicheri, Frank -- Durocher, Daniel -- 84297-1/Canadian Institutes of Health Research/Canada -- 84297-2/Canadian Institutes of Health Research/Canada -- MOP84297/Canadian Institutes of Health Research/Canada -- England -- Nature. 2013 Jul 4;499(7456):50-4. doi: 10.1038/nature12318. Epub 2013 Jun 12.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Samuel Lunenfeld Research Institute, Mount Sinai Hospital, 600 University Avenue, Toronto, Ontario M5G 1X5, Canada.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23760478" target="_blank"〉PubMed〈/a〉
    Keywords: Amino Acid Motifs ; Amino Acid Sequence ; Animals ; Cell Cycle Proteins/chemistry/metabolism ; Cell Line ; Chromosomal Proteins, Non-Histone/chemistry/deficiency/genetics ; DNA Breaks, Double-Stranded ; *DNA Damage ; DNA-Binding Proteins/chemistry/deficiency/genetics ; Female ; Histones/*chemistry/*metabolism ; Humans ; Intracellular Signaling Peptides and ; Proteins/chemistry/deficiency/genetics/*metabolism ; Lysine/*metabolism ; Male ; Mice ; Molecular Sequence Data ; Mutant Proteins/chemistry/metabolism ; Nuclear Proteins/chemistry/metabolism ; Nucleosomes/chemistry/metabolism ; Protein Binding ; Protein Structure, Tertiary ; Schizosaccharomyces ; Schizosaccharomyces pombe Proteins/chemistry/metabolism ; Signal Transduction ; Ubiquitin/*metabolism ; *Ubiquitination
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  • 10
    Publication Date: 2013-07-23
    Description: During translation initiation in eukaryotes, the small ribosomal subunit binds messenger RNA at the 5' end and scans in the 5' to 3' direction to locate the initiation codon, form the 80S initiation complex and start protein synthesis. This simple, yet intricate, process is guided by multiple initiation factors. Here we determine the structures of three complexes of the small ribosomal subunit that represent distinct steps in mammalian translation initiation. These structures reveal the locations of eIF1, eIF1A, mRNA and initiator transfer RNA bound to the small ribosomal subunit and provide insights into the details of translation initiation specific to eukaryotes. Conformational changes associated with the captured functional states reveal the dynamics of the interactions in the P site of the ribosome. These results have functional implications for the mechanism of mRNA scanning.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3748252/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3748252/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Lomakin, Ivan B -- Steitz, Thomas A -- GM022778/GM/NIGMS NIH HHS/ -- P01 GM022778/GM/NIGMS NIH HHS/ -- Howard Hughes Medical Institute/ -- England -- Nature. 2013 Aug 15;500(7462):307-11. doi: 10.1038/nature12355. Epub 2013 Jul 21.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut 06520-8114, USA. ivan.lomakin@yale.edu〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23873042" target="_blank"〉PubMed〈/a〉
    Keywords: Animals ; Crystallography, X-Ray ; Eukaryotic Initiation Factor-1/chemistry/metabolism ; Humans ; *Models, Molecular ; Protein Binding ; *Protein Biosynthesis ; Protein Structure, Quaternary ; RNA, Messenger/*chemistry/*metabolism ; RNA, Transfer, Met/chemistry/metabolism ; Rabbits ; Ribosome Subunits, Small, Eukaryotic/chemistry/metabolism ; Ribosomes/metabolism
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  • 11
    Publication Date: 2013-10-29
    Description: Pentatricopeptide repeat (PPR) proteins represent a large family of sequence-specific RNA-binding proteins that are involved in multiple aspects of RNA metabolism. PPR proteins, which are found in exceptionally large numbers in the mitochondria and chloroplasts of terrestrial plants, recognize single-stranded RNA (ssRNA) in a modular fashion. The maize chloroplast protein PPR10 binds to two similar RNA sequences from the ATPI-ATPH and PSAJ-RPL33 intergenic regions, referred to as ATPH and PSAJ, respectively. By protecting the target RNA elements from 5' or 3' exonucleases, PPR10 defines the corresponding 5' and 3' messenger RNA termini. Despite rigorous functional characterizations, the structural basis of sequence-specific ssRNA recognition by PPR proteins remains to be elucidated. Here we report the crystal structures of PPR10 in RNA-free and RNA-bound states at resolutions of 2.85 and 2.45 A, respectively. In the absence of RNA binding, the nineteen repeats of PPR10 are assembled into a right-handed superhelical spiral. PPR10 forms an antiparallel, intertwined homodimer and exhibits considerable conformational changes upon binding to its target ssRNA, an 18-nucleotide PSAJ element. Six nucleotides of PSAJ are specifically recognized by six corresponding PPR10 repeats following the predicted code. The molecular basis for the specific and modular recognition of RNA bases A, G and U is revealed. The structural elucidation of RNA recognition by PPR proteins provides an important framework for potential biotechnological applications of PPR proteins in RNA-related research areas.〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Yin, Ping -- Li, Quanxiu -- Yan, Chuangye -- Liu, Ying -- Liu, Junjie -- Yu, Feng -- Wang, Zheng -- Long, Jiafu -- He, Jianhua -- Wang, Hong-Wei -- Wang, Jiawei -- Zhu, Jian-Kang -- Shi, Yigong -- Yan, Nieng -- Howard Hughes Medical Institute/ -- England -- Nature. 2013 Dec 5;504(7478):168-71. doi: 10.1038/nature12651. Epub 2013 Oct 27.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉1] State Key Laboratory of Bio-membrane and Membrane Biotechnology, Tsinghua University, Beijing 100084, China [2] Center for Structural Biology, School of Life Sciences and School of Medicine, Tsinghua-Peking Center for Life Sciences, Tsinghua University, Beijing 100084, China [3].〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/24162847" target="_blank"〉PubMed〈/a〉
    Keywords: Binding Sites ; Crystallography, X-Ray ; *Models, Molecular ; Plant Proteins/*chemistry/genetics/*metabolism ; Protein Binding ; Protein Structure, Tertiary ; RNA/chemistry/*metabolism ; Zea mays/*chemistry/genetics/metabolism
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  • 12
    Publication Date: 2013-03-12
    Description: The regulated loading of the Mcm2-7 DNA helicase (comprising six related subunits, Mcm2 to Mcm7) into pre-replicative complexes at multiple replication origins ensures precise once per cell cycle replication in eukaryotic cells. The origin recognition complex (ORC), Cdc6 and Cdt1 load Mcm2-7 into a double hexamer bound around duplex DNA in an ATP-dependent reaction, but the molecular mechanism of this origin 'licensing' is still poorly understood. Here we show that both Mcm2-7 hexamers in Saccharomyces cerevisiae are recruited to origins by an essential, conserved carboxy-terminal domain of Mcm3 that interacts with and stimulates the ATPase activity of ORC-Cdc6. ATP hydrolysis can promote Mcm2-7 loading, but can also promote Mcm2-7 release if components are missing or if ORC has been inactivated by cyclin-dependent kinase phosphorylation. Our work provides new insights into how origins are licensed and reveals a novel ATPase-dependent mechanism contributing to precise once per cell cycle replication.〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Frigola, Jordi -- Remus, Dirk -- Mehanna, Amina -- Diffley, John F X -- Cancer Research UK/United Kingdom -- England -- Nature. 2013 Mar 21;495(7441):339-43. doi: 10.1038/nature11920. Epub 2013 Mar 10.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Cancer Research UK London Research Institute, Clare Hall Laboratories, South Mimms EN6 3LD, UK.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23474987" target="_blank"〉PubMed〈/a〉
    Keywords: Adenosine Triphosphatases/*metabolism ; Adenosine Triphosphate/metabolism ; Amino Acid Sequence ; Cell Cycle Proteins/metabolism ; Chromosomal Proteins, Non-Histone/metabolism ; DNA Replication/*genetics ; DNA-Binding Proteins/metabolism ; Enzyme Activation ; Hydrolysis ; Minichromosome Maintenance Complex Component 3 ; Minichromosome Maintenance Complex Component 7 ; Nuclear Proteins/metabolism ; Protein Binding ; Replication Origin/*genetics ; Saccharomyces cerevisiae/cytology/enzymology/*genetics/*metabolism ; Saccharomyces cerevisiae Proteins/metabolism ; Sequence Alignment
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  • 13
    Publication Date: 2013-07-09
    Description: The newly emergent Middle East respiratory syndrome coronavirus (MERS-CoV) can cause severe pulmonary disease in humans, representing the second example of a highly pathogenic coronavirus, the first being SARS-CoV. CD26 (also known as dipeptidyl peptidase 4, DPP4) was recently identified as the cellular receptor for MERS-CoV. The engagement of the MERS-CoV spike protein with CD26 mediates viral attachment to host cells and virus-cell fusion, thereby initiating infection. Here we delineate the molecular basis of this specific interaction by presenting the first crystal structures of both the free receptor binding domain (RBD) of the MERS-CoV spike protein and its complex with CD26. Furthermore, binding between the RBD and CD26 is measured using real-time surface plasmon resonance with a dissociation constant of 16.7 nM. The viral RBD is composed of a core subdomain homologous to that of the SARS-CoV spike protein, and a unique strand-dominated external receptor binding motif that recognizes blades IV and V of the CD26 beta-propeller. The atomic details at the interface between the two binding entities reveal a surprising protein-protein contact mediated mainly by hydrophilic residues. Sequence alignment indicates, among betacoronaviruses, a possible structural conservation for the region homologous to the MERS-CoV RBD core, but a high variation in the external receptor binding motif region for virus-specific pathogenesis such as receptor recognition.〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Lu, Guangwen -- Hu, Yawei -- Wang, Qihui -- Qi, Jianxun -- Gao, Feng -- Li, Yan -- Zhang, Yanfang -- Zhang, Wei -- Yuan, Yuan -- Bao, Jinku -- Zhang, Buchang -- Shi, Yi -- Yan, Jinghua -- Gao, George F -- England -- Nature. 2013 Aug 8;500(7461):227-31. doi: 10.1038/nature12328. Epub 2013 Jul 7.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉CAS Key Laboratory of Pathogenic Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23831647" target="_blank"〉PubMed〈/a〉
    Keywords: Conserved Sequence/genetics ; Coronavirus/*chemistry/genetics/*metabolism ; Dipeptidyl Peptidase 4/*chemistry/metabolism ; Humans ; Protein Binding ; Protein Interaction Domains and Motifs/genetics ; Protein Structure, Tertiary/genetics ; Receptors, Virus/*chemistry/*metabolism ; *Virus Attachment
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  • 14
    Publication Date: 2013-01-22
    Description: In various cellular membrane systems, vacuolar ATPases (V-ATPases) function as proton pumps, which are involved in many processes such as bone resorption and cancer metastasis, and these membrane proteins represent attractive drug targets for osteoporosis and cancer. The hydrophilic V(1) portion is known as a rotary motor, in which a central axis DF complex rotates inside a hexagonally arranged catalytic A(3)B(3) complex using ATP hydrolysis energy, but the molecular mechanism is not well defined owing to a lack of high-resolution structural information. We previously reported on the in vitro expression, purification and reconstitution of Enterococcus hirae V(1)-ATPase from the A(3)B(3) and DF complexes. Here we report the asymmetric structures of the nucleotide-free (2.8 A) and nucleotide-bound (3.4 A) A(3)B(3) complex that demonstrate conformational changes induced by nucleotide binding, suggesting a binding order in the right-handed rotational orientation in a cooperative manner. The crystal structures of the nucleotide-free (2.2 A) and nucleotide-bound (2.7 A) V(1)-ATPase are also reported. The more tightly packed nucleotide-binding site seems to be induced by DF binding, and ATP hydrolysis seems to be stimulated by the approach of a conserved arginine residue. To our knowledge, these asymmetric structures represent the first high-resolution view of the rotational mechanism of V(1)-ATPase.〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Arai, Satoshi -- Saijo, Shinya -- Suzuki, Kano -- Mizutani, Kenji -- Kakinuma, Yoshimi -- Ishizuka-Katsura, Yoshiko -- Ohsawa, Noboru -- Terada, Takaho -- Shirouzu, Mikako -- Yokoyama, Shigeyuki -- Iwata, So -- Yamato, Ichiro -- Murata, Takeshi -- England -- Nature. 2013 Jan 31;493(7434):703-7. doi: 10.1038/nature11778. Epub 2013 Jan 13.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Department of Chemistry, Graduate School of Science, Chiba University, 1-33 Yayoi-cho, Inage, Chiba 263-8522, Japan.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23334411" target="_blank"〉PubMed〈/a〉
    Keywords: Binding Sites ; Crystallization ; Enterococcus/*enzymology/genetics ; *Models, Molecular ; Mutation ; Nucleotides/metabolism ; Protein Binding ; Protein Structure, Tertiary ; Protein Subunits ; Rotation ; Vacuolar Proton-Translocating ATPases/*chemistry/genetics
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  • 15
    Publication Date: 2013-07-13
    Description: Cell-surface receptors frequently use scaffold proteins to recruit cytoplasmic targets, but the rationale for this is uncertain. Activated receptor tyrosine kinases, for example, engage scaffolds such as Shc1 that contain phosphotyrosine (pTyr)-binding (PTB) domains. Using quantitative mass spectrometry, here we show that mammalian Shc1 responds to epidermal growth factor (EGF) stimulation through multiple waves of distinct phosphorylation events and protein interactions. After stimulation, Shc1 rapidly binds a group of proteins that activate pro-mitogenic or survival pathways dependent on recruitment of the Grb2 adaptor to Shc1 pTyr sites. Akt-mediated feedback phosphorylation of Shc1 Ser 29 then recruits the Ptpn12 tyrosine phosphatase. This is followed by a sub-network of proteins involved in cytoskeletal reorganization, trafficking and signal termination that binds Shc1 with delayed kinetics, largely through the SgK269 pseudokinase/adaptor protein. Ptpn12 acts as a switch to convert Shc1 from pTyr/Grb2-based signalling to SgK269-mediated pathways that regulate cell invasion and morphogenesis. The Shc1 scaffold therefore directs the temporal flow of signalling information after EGF stimulation.〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Zheng, Yong -- Zhang, Cunjie -- Croucher, David R -- Soliman, Mohamed A -- St-Denis, Nicole -- Pasculescu, Adrian -- Taylor, Lorne -- Tate, Stephen A -- Hardy, W Rod -- Colwill, Karen -- Dai, Anna Yue -- Bagshaw, Rick -- Dennis, James W -- Gingras, Anne-Claude -- Daly, Roger J -- Pawson, Tony -- MOP-13466-6849/Canadian Institutes of Health Research/Canada -- England -- Nature. 2013 Jul 11;499(7457):166-71. doi: 10.1038/nature12308.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Samuel Lunenfeld Research Institute, Mount Sinai Hospital, 600 University Avenue, Toronto M5G 1X5, Canada.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23846654" target="_blank"〉PubMed〈/a〉
    Keywords: Animals ; Breast/cytology ; Cell Line ; Epidermal Growth Factor/*metabolism ; Epithelial Cells/cytology ; Extracellular Signal-Regulated MAP Kinases/metabolism ; Feedback, Physiological ; GRB2 Adaptor Protein/deficiency/genetics/metabolism ; Humans ; Mice ; Multiprotein Complexes/chemistry/metabolism ; Phosphorylation ; Protein Binding ; Protein-Tyrosine Kinases ; Proto-Oncogene Proteins c-akt/metabolism ; Rats ; Receptor, Epidermal Growth Factor/agonists/metabolism ; Shc Signaling Adaptor Proteins/deficiency/genetics/*metabolism ; *Signal Transduction ; Time Factors
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  • 16
    Publication Date: 2013-12-18
    Description: Strigolactones (SLs), a newly discovered class of carotenoid-derived phytohormones, are essential for developmental processes that shape plant architecture and interactions with parasitic weeds and symbiotic arbuscular mycorrhizal fungi. Despite the rapid progress in elucidating the SL biosynthetic pathway, the perception and signalling mechanisms of SL remain poorly understood. Here we show that DWARF 53 (D53) acts as a repressor of SL signalling and that SLs induce its degradation. We find that the rice (Oryza sativa) d53 mutant, which produces an exaggerated number of tillers compared to wild-type plants, is caused by a gain-of-function mutation and is insensitive to exogenous SL treatment. The D53 gene product shares predicted features with the class I Clp ATPase proteins and can form a complex with the alpha/beta hydrolase protein DWARF 14 (D14) and the F-box protein DWARF 3 (D3), two previously identified signalling components potentially responsible for SL perception. We demonstrate that, in a D14- and D3-dependent manner, SLs induce D53 degradation by the proteasome and abrogate its activity in promoting axillary bud outgrowth. Our combined genetic and biochemical data reveal that D53 acts as a repressor of the SL signalling pathway, whose hormone-induced degradation represents a key molecular link between SL perception and responses.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4096652/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4096652/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Zhou, Feng -- Lin, Qibing -- Zhu, Lihong -- Ren, Yulong -- Zhou, Kunneng -- Shabek, Nitzan -- Wu, Fuqing -- Mao, Haibin -- Dong, Wei -- Gan, Lu -- Ma, Weiwei -- Gao, He -- Chen, Jun -- Yang, Chao -- Wang, Dan -- Tan, Junjie -- Zhang, Xin -- Guo, Xiuping -- Wang, Jiulin -- Jiang, Ling -- Liu, Xi -- Chen, Weiqi -- Chu, Jinfang -- Yan, Cunyu -- Ueno, Kotomi -- Ito, Shinsaku -- Asami, Tadao -- Cheng, Zhijun -- Wang, Jie -- Lei, Cailin -- Zhai, Huqu -- Wu, Chuanyin -- Wang, Haiyang -- Zheng, Ning -- Wan, Jianmin -- R01 CA107134/CA/NCI NIH HHS/ -- Howard Hughes Medical Institute/ -- England -- Nature. 2013 Dec 19;504(7480):406-10. doi: 10.1038/nature12878. Epub 2013 Dec 11.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉1] National Key Laboratory for Crop Genetics and Germplasm Enhancement, Jiangsu Plant Gene Engineering Research Center, Nanjing Agricultural University, Nanjing 210095, China [2] National Key Facility for Crop Gene Resources and Genetic Improvement, Institute of Crop Science, Chinese Academy of Agricultural Sciences, Beijing 100081, China. ; National Key Facility for Crop Gene Resources and Genetic Improvement, Institute of Crop Science, Chinese Academy of Agricultural Sciences, Beijing 100081, China. ; National Key Laboratory for Crop Genetics and Germplasm Enhancement, Jiangsu Plant Gene Engineering Research Center, Nanjing Agricultural University, Nanjing 210095, China. ; 1] Department of Pharmacology, University of Washington, Seattle, Washington 98195, USA [2] Howard Hughes Medical Institute, Box 357280, University of Washington, Seattle, Washington 98195, USA. ; National Centre for Plant Gene Research (Beijing), Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, 1-2 Beichen West Road, Beijing 100101, China. ; Department of Applied Biological Chemistry, The University of Tokyo, 1-1-1 Yayoi, Bunkyo, Tokyo 113-8657, Japan.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/24336215" target="_blank"〉PubMed〈/a〉
    Keywords: Amino Acid Sequence ; Cloning, Molecular ; Gene Expression Regulation, Plant ; Lactones/*metabolism ; Molecular Sequence Data ; Mutation/genetics ; Oryza/genetics/*metabolism ; Phenotype ; Plant Growth Regulators/*metabolism ; Plant Proteins/genetics/*metabolism ; Proteasome Endopeptidase Complex/metabolism ; Protein Binding ; *Proteolysis ; SKP Cullin F-Box Protein Ligases/*metabolism ; *Signal Transduction
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  • 17
    Publication Date: 2013-12-18
    Description: Strigolactones (SLs) are a group of newly identified plant hormones that control plant shoot branching. SL signalling requires the hormone-dependent interaction of DWARF 14 (D14), a probable candidate SL receptor, with DWARF 3 (D3), an F-box component of the Skp-Cullin-F-box (SCF) E3 ubiquitin ligase complex. Here we report the characterization of a dominant SL-insensitive rice (Oryza sativa) mutant dwarf 53 (d53) and the cloning of D53, which encodes a substrate of the SCF(D3) ubiquitination complex and functions as a repressor of SL signalling. Treatments with GR24, a synthetic SL analogue, cause D53 degradation via the proteasome in a manner that requires D14 and the SCF(D3) ubiquitin ligase, whereas the dominant form of D53 is resistant to SL-mediated degradation. Moreover, D53 can interact with transcriptional co-repressors known as TOPLESS-RELATED PROTEINS. Our results suggest a model of SL signalling that involves SL-dependent degradation of the D53 repressor mediated by the D14-D3 complex.〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Jiang, Liang -- Liu, Xue -- Xiong, Guosheng -- Liu, Huihui -- Chen, Fulu -- Wang, Lei -- Meng, Xiangbing -- Liu, Guifu -- Yu, Hong -- Yuan, Yundong -- Yi, Wei -- Zhao, Lihua -- Ma, Honglei -- He, Yuanzheng -- Wu, Zhongshan -- Melcher, Karsten -- Qian, Qian -- Xu, H Eric -- Wang, Yonghong -- Li, Jiayang -- England -- Nature. 2013 Dec 19;504(7480):401-5. doi: 10.1038/nature12870. Epub 2013 Dec 11.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉1] State Key Laboratory of Plant Genomics and National Center for Plant Gene Research (Beijing), Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing 100101, China [2]. ; State Key Laboratory of Plant Genomics and National Center for Plant Gene Research (Beijing), Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing 100101, China. ; VARI-SIMM Center, Center for Structure and Function of Drug Targets, CAS-Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China. ; Laboratory of Structural Sciences, Van Andel Research Institute, 333 Bostwick Avenue Northeast, Grand Rapids, Michigan 49503, USA. ; State Key Laboratory of Rice Biology, China National Rice Research Institute, Chinese Academy of Agricultural Sciences, Hangzhou 310006, China. ; 1] VARI-SIMM Center, Center for Structure and Function of Drug Targets, CAS-Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China [2] Laboratory of Structural Sciences, Van Andel Research Institute, 333 Bostwick Avenue Northeast, Grand Rapids, Michigan 49503, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/24336200" target="_blank"〉PubMed〈/a〉
    Keywords: Cloning, Molecular ; Gene Expression Regulation, Plant ; Lactones/*antagonists & inhibitors/*metabolism ; Models, Biological ; Multiprotein Complexes/chemistry/metabolism ; Mutation/genetics ; Oryza/genetics/*metabolism ; Plant Growth Regulators/antagonists & inhibitors/*metabolism ; Plant Proteins/chemistry/genetics/*metabolism ; Proteasome Endopeptidase Complex/metabolism ; Protein Binding ; Proteolysis ; *Signal Transduction ; Ubiquitin/metabolism
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  • 18
    Publication Date: 2013-08-09
    Description: The bacterial type VI secretion system (T6SS) is a large multicomponent, dynamic macromolecular machine that has an important role in the ecology of many Gram-negative bacteria. T6SS is responsible for translocation of a wide range of toxic effector molecules, allowing predatory cells to kill both prokaryotic as well as eukaryotic prey cells. The T6SS organelle is functionally analogous to contractile tails of bacteriophages and is thought to attack cells by initially penetrating them with a trimeric protein complex called the VgrG spike. Neither the exact protein composition of the T6SS organelle nor the mechanisms of effector selection and delivery are known. Here we report that proteins from the PAAR (proline-alanine-alanine-arginine) repeat superfamily form a sharp conical extension on the VgrG spike, which is further involved in attaching effector domains to the spike. The crystal structures of two PAAR-repeat proteins bound to VgrG-like partners show that these proteins sharpen the tip of the T6SS spike complex. We demonstrate that PAAR proteins are essential for T6SS-mediated secretion and target cell killing by Vibrio cholerae and Acinetobacter baylyi. Our results indicate a new model of the T6SS organelle in which the VgrG-PAAR spike complex is decorated with multiple effectors that are delivered simultaneously into target cells in a single contraction-driven translocation event.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3792578/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3792578/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Shneider, Mikhail M -- Buth, Sergey A -- Ho, Brian T -- Basler, Marek -- Mekalanos, John J -- Leiman, Petr G -- AI-01845/AI/NIAID NIH HHS/ -- AI-026289/AI/NIAID NIH HHS/ -- R01 AI018045/AI/NIAID NIH HHS/ -- R01 AI026289/AI/NIAID NIH HHS/ -- England -- Nature. 2013 Aug 15;500(7462):350-3. doi: 10.1038/nature12453. Epub 2013 Aug 7.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Ecole Polytechnique Federale de Lausanne (EPFL), BSP-415, 1015 Lausanne, Switzerland.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23925114" target="_blank"〉PubMed〈/a〉
    Keywords: Acinetobacter/genetics/metabolism ; Bacterial Proteins/*chemistry/*secretion ; Bacterial Secretion Systems/*genetics ; Microsatellite Repeats/*physiology ; Protein Binding ; Vibrio cholerae/genetics/metabolism
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  • 19
    Publication Date: 2013-01-08
    Description: The initiation of gene transcription by RNA polymerase II is regulated by a plethora of proteins in human cells. The first general transcription factor to bind gene promoters is transcription factor IID (TFIID). TFIID triggers pre-initiation complex formation, functions as a coactivator by interacting with transcriptional activators and reads epigenetic marks. TFIID is a megadalton-sized multiprotein complex composed of TATA-box-binding protein (TBP) and 13 TBP-associated factors (TAFs). Despite its crucial role, the detailed architecture and assembly mechanism of TFIID remain elusive. Histone fold domains are prevalent in TAFs, and histone-like tetramer and octamer structures have been proposed in TFIID. A functional core-TFIID subcomplex was revealed in Drosophila nuclei, consisting of a subset of TAFs (TAF4, TAF5, TAF6, TAF9 and TAF12). These core subunits are thought to be present in two copies in holo-TFIID, in contrast to TBP and other TAFs that are present in a single copy, conveying a transition from symmetry to asymmetry in the TFIID assembly pathway. Here we present the structure of human core-TFIID determined by cryo-electron microscopy at 11.6 A resolution. Our structure reveals a two-fold symmetric, interlaced architecture, with pronounced protrusions, that accommodates all conserved structural features of the TAFs including the histone folds. We further demonstrate that binding of one TAF8-TAF10 complex breaks the original symmetry of core-TFIID. We propose that the resulting asymmetric structure serves as a functional scaffold to nucleate holo-TFIID assembly, by accreting one copy each of the remaining TAFs and TBP.〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Bieniossek, Christoph -- Papai, Gabor -- Schaffitzel, Christiane -- Garzoni, Frederic -- Chaillet, Maxime -- Scheer, Elisabeth -- Papadopoulos, Petros -- Tora, Laszlo -- Schultz, Patrick -- Berger, Imre -- England -- Nature. 2013 Jan 31;493(7434):699-702. doi: 10.1038/nature11791. Epub 2013 Jan 6.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉European Molecular Biology Laboratory Grenoble Outstation, Unit of Virus Host Cell Interactions UVHCI, UJF-CNRS-EMBL Unite Mixte International UMI 3265, 6 rue Jules Horowitz, 38042 Grenoble Cedex 9, France.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23292512" target="_blank"〉PubMed〈/a〉
    Keywords: Cells, Cultured ; Cryoelectron Microscopy ; HeLa Cells ; Humans ; *Models, Molecular ; Protein Binding ; Protein Structure, Tertiary ; Transcription Factor TFIID/*chemistry/genetics/metabolism
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  • 20
    Publication Date: 2013-10-29
    Description: Integrins have a critical role in thrombosis and haemostasis. Antagonists of the platelet integrin alphaIIbbeta3 are potent anti-thrombotic drugs, but also have the life-threatening adverse effect of causing bleeding. It is therefore desirable to develop new antagonists that do not cause bleeding. Integrins transmit signals bidirectionally. Inside-out signalling activates integrins through a talin-dependent mechanism. Integrin ligation mediates thrombus formation and outside-in signalling, which requires Galpha13 and greatly expands thrombi. Here we show that Galpha13 and talin bind to mutually exclusive but distinct sites within the integrin beta3 cytoplasmic domain in opposing waves. The first talin-binding wave mediates inside-out signalling and also ligand-induced integrin activation, but is not required for outside-in signalling. Integrin ligation induces transient talin dissociation and Galpha13 binding to an EXE motif (in which X denotes any residue), which selectively mediates outside-in signalling and platelet spreading. The second talin-binding wave is associated with clot retraction. An EXE-motif-based inhibitor of Galpha13-integrin interaction selectively abolishes outside-in signalling without affecting integrin ligation, and suppresses occlusive arterial thrombosis without affecting bleeding time. Thus, we have discovered a new mechanism for the directional switch of integrin signalling and, on the basis of this mechanism, designed a potent new anti-thrombotic drug that does not cause bleeding.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3823815/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3823815/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Shen, Bo -- Zhao, Xiaojuan -- O'Brien, Kelly A -- Stojanovic-Terpo, Aleksandra -- Delaney, M Keegan -- Kim, Kyungho -- Cho, Jaehyung -- Lam, Stephen C-T -- Du, Xiaoping -- HL062350/HL/NHLBI NIH HHS/ -- HL080264/HL/NHLBI NIH HHS/ -- HL109439/HL/NHLBI NIH HHS/ -- R01 HL080264/HL/NHLBI NIH HHS/ -- R01 HL109439/HL/NHLBI NIH HHS/ -- T32 HL007829/HL/NHLBI NIH HHS/ -- England -- Nature. 2013 Nov 7;503(7474):131-5. doi: 10.1038/nature12613. Epub 2013 Oct 27.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Department of Pharmacology, University of Illinois at Chicago, 835 South Wolcott Avenue, Chicago, Illinois 60612, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/24162846" target="_blank"〉PubMed〈/a〉
    Keywords: Amino Acid Motifs ; Amino Acid Sequence ; Animals ; Antithrombins/adverse effects/*pharmacology/therapeutic use ; Binding Sites ; Bleeding Time ; *Cell Polarity ; Cytoplasm/metabolism ; GTP-Binding Protein alpha Subunits, G12-G13/metabolism ; Hemorrhage/chemically induced ; Humans ; Integrin beta3/chemistry/genetics/metabolism ; Integrins/chemistry/deficiency/genetics/*metabolism ; Male ; Mice ; Mice, Inbred C57BL ; Molecular Sequence Data ; Platelet Glycoprotein GPIIb-IIIa Complex/metabolism ; Protein Binding ; Protein Structure, Tertiary ; Signal Transduction/*drug effects ; Talin/metabolism ; Thrombosis/*drug therapy/metabolism/pathology
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  • 21
    Publication Date: 2013-11-19
    Description: Eukaryotic ribosomes are assembled by a complex pathway that extends from the nucleolus to the cytoplasm and is powered by many energy-consuming enzymes. Nuclear export is a key, irreversible step in pre-ribosome maturation, but mechanisms underlying the timely acquisition of export competence remain poorly understood. Here we show that a conserved Saccharomyces cerevisiae GTPase Nug2 (also known as Nog2, and as NGP-1, GNL2 or nucleostemin 2 in human) has a key role in the timing of export competence. Nug2 binds the inter-subunit face of maturing, nucleoplasmic pre-60S particles, and the location clashes with the position of Nmd3, a key pre-60S export adaptor. Nug2 and Nmd3 are not present on the same pre-60S particles, with Nug2 binding before Nmd3. Depletion of Nug2 causes premature Nmd3 binding to the pre-60S particles, whereas mutations in the G-domain of Nug2 block Nmd3 recruitment, resulting in severe 60S export defects. Two pre-60S remodelling factors, the Rea1 ATPase and its co-substrate Rsa4, are present on Nug2-associated particles, and both show synthetic lethal interactions with nug2 mutants. Release of Nug2 from pre-60S particles requires both its K(+)-dependent GTPase activity and the remodelling ATPase activity of Rea1. We conclude that Nug2 is a regulatory GTPase that monitors pre-60S maturation, with release from its placeholder site linked to recruitment of the nuclear export machinery.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3880858/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3880858/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Matsuo, Yoshitaka -- Granneman, Sander -- Thoms, Matthias -- Manikas, Rizos-Georgios -- Tollervey, David -- Hurt, Ed -- 077248/Wellcome Trust/United Kingdom -- 092076/Wellcome Trust/United Kingdom -- Wellcome Trust/United Kingdom -- England -- Nature. 2014 Jan 2;505(7481):112-6. doi: 10.1038/nature12731. Epub 2013 Nov 17.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Biochemie-Zentrum der Universitat Heidelberg, Im Neuenheimer Feld 328, Heidelberg D-69120, Germany. ; 1] Wellcome Trust Centre for Cell Biology, The University of Edinburgh, Edinburgh EH9 3JR, UK [2] Centre for Synthetic and Systems Biology, The University of Edinburgh, Edinburgh EH9 3JD, UK [3]. ; 1] Biochemie-Zentrum der Universitat Heidelberg, Im Neuenheimer Feld 328, Heidelberg D-69120, Germany [2]. ; Wellcome Trust Centre for Cell Biology, The University of Edinburgh, Edinburgh EH9 3JR, UK.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/24240281" target="_blank"〉PubMed〈/a〉
    Keywords: Adenosine Triphosphatases/*metabolism ; Cell Nucleus/*metabolism/secretion ; Cytoplasm/metabolism ; GTP Phosphohydrolases/chemistry/genetics/*metabolism ; Genes, Lethal/genetics ; Models, Molecular ; Mutation/genetics ; Potassium/metabolism ; Protein Binding ; Protein Structure, Tertiary/genetics ; RNA-Binding Proteins/chemistry/metabolism ; Ribosomal Proteins/metabolism ; Ribosome Subunits, Large, Eukaryotic/chemistry/metabolism ; Ribosomes/*chemistry/*metabolism/secretion ; Saccharomyces cerevisiae/cytology/enzymology/*metabolism ; Saccharomyces cerevisiae Proteins/chemistry/genetics/*metabolism/secretion
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  • 22
    Publication Date: 2013-11-22
    Description: Despite recent advances in crystallography and the availability of G-protein-coupled receptor (GPCR) structures, little is known about the mechanism of their activation process, as only the beta2 adrenergic receptor (beta2AR) and rhodopsin have been crystallized in fully active conformations. Here we report the structure of an agonist-bound, active state of the human M2 muscarinic acetylcholine receptor stabilized by a G-protein mimetic camelid antibody fragment isolated by conformational selection using yeast surface display. In addition to the expected changes in the intracellular surface, the structure reveals larger conformational changes in the extracellular region and orthosteric binding site than observed in the active states of the beta2AR and rhodopsin. We also report the structure of the M2 receptor simultaneously bound to the orthosteric agonist iperoxo and the positive allosteric modulator LY2119620. This structure reveals that LY2119620 recognizes a largely pre-formed binding site in the extracellular vestibule of the iperoxo-bound receptor, inducing a slight contraction of this outer binding pocket. These structures offer important insights into the activation mechanism and allosteric modulation of muscarinic receptors.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4020789/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4020789/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Kruse, Andrew C -- Ring, Aaron M -- Manglik, Aashish -- Hu, Jianxin -- Hu, Kelly -- Eitel, Katrin -- Hubner, Harald -- Pardon, Els -- Valant, Celine -- Sexton, Patrick M -- Christopoulos, Arthur -- Felder, Christian C -- Gmeiner, Peter -- Steyaert, Jan -- Weis, William I -- Garcia, K Christopher -- Wess, Jurgen -- Kobilka, Brian K -- GM08311806/GM/NIGMS NIH HHS/ -- NS02847123/NS/NINDS NIH HHS/ -- T32 GM008294/GM/NIGMS NIH HHS/ -- U19 GM106990/GM/NIGMS NIH HHS/ -- Howard Hughes Medical Institute/ -- Intramural NIH HHS/ -- England -- Nature. 2013 Dec 5;504(7478):101-6. doi: 10.1038/nature12735. Epub 2013 Nov 20.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉1] Department of Molecular and Cellular Physiology, Stanford University School of Medicine, 279 Campus Drive, Stanford, California 94305, USA [2].〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/24256733" target="_blank"〉PubMed〈/a〉
    Keywords: Allosteric Regulation ; Binding Sites ; Cytoplasm/metabolism ; Humans ; Isoxazoles/chemistry/metabolism ; *Models, Molecular ; Protein Binding ; Protein Structure, Tertiary ; Quaternary Ammonium Compounds/chemistry/metabolism ; Receptors, Muscarinic/*chemistry/*metabolism
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  • 23
    Publication Date: 2013-10-08
    Description: In most eukaryotic cells microtubules undergo post-translational modifications such as acetylation of alpha-tubulin on lysine 40, a widespread modification restricted to a subset of microtubules that turns over slowly. This subset of stable microtubules accumulates in cell protrusions and regulates cell polarization, migration and invasion. However, mechanisms restricting acetylation to these microtubules are unknown. Here we report that clathrin-coated pits (CCPs) control microtubule acetylation through a direct interaction of the alpha-tubulin acetyltransferase alphaTAT1 (refs 8, 9) with the clathrin adaptor AP2. We observe that about one-third of growing microtubule ends contact and pause at CCPs and that loss of CCPs decreases lysine 40 acetylation levels. We show that alphaTAT1 localizes to CCPs through a direct interaction with AP2 that is required for microtubule acetylation. In migrating cells, the polarized orientation of acetylated microtubules correlates with CCP accumulation at the leading edge, and interaction of alphaTAT1 with AP2 is required for directional migration. We conclude that microtubules contacting CCPs become acetylated by alphaTAT1. In migrating cells, this mechanism ensures the acetylation of microtubules oriented towards the leading edge, thus promoting directional cell locomotion and chemotaxis.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3970258/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3970258/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Montagnac, Guillaume -- Meas-Yedid, Vannary -- Irondelle, Marie -- Castro-Castro, Antonio -- Franco, Michel -- Shida, Toshinobu -- Nachury, Maxence V -- Benmerah, Alexandre -- Olivo-Marin, Jean-Christophe -- Chavrier, Philippe -- R01 GM089933/GM/NIGMS NIH HHS/ -- England -- Nature. 2013 Oct 24;502(7472):567-70. doi: 10.1038/nature12571. Epub 2013 Oct 6.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉1] Institut Curie, Research Center, 75005 Paris, France [2] Membrane and Cytoskeleton Dynamics, CNRS UMR 144, 75005 Paris, France.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/24097348" target="_blank"〉PubMed〈/a〉
    Keywords: Acetylation ; Acetyltransferases/*metabolism ; Adaptor Protein Complex 2/metabolism ; Biocatalysis ; Cell Movement ; Clathrin/*metabolism ; Coated Pits, Cell-Membrane/enzymology/*metabolism ; HeLa Cells ; Humans ; Microtubules/chemistry/*metabolism ; Protein Binding ; Tubulin/metabolism
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  • 24
    Publication Date: 2013-08-27
    Description: The KCNH voltage-dependent potassium channels (ether-a-go-go, EAG; EAG-related gene, ERG; EAG-like channels, ELK) are important regulators of cellular excitability and have key roles in diseases such as cardiac long QT syndrome type 2 (LQT2), epilepsy, schizophrenia and cancer. The intracellular domains of KCNH channels are structurally distinct from other voltage-gated channels. The amino-terminal region contains an eag domain, which is composed of a Per-Arnt-Sim (PAS) domain and a PAS-cap domain, whereas the carboxy-terminal region contains a cyclic nucleotide-binding homology domain (CNBHD), which is connected to the pore through a C-linker domain. Many disease-causing mutations localize to these specialized intracellular domains, which underlie the unique gating and regulation of KCNH channels. It has been suggested that the eag domain may regulate the channel by interacting with either the S4-S5 linker or the CNBHD. Here we present a 2 A resolution crystal structure of the eag domain-CNBHD complex of the mouse EAG1 (also known as KCNH1) channel. It displays extensive interactions between the eag domain and the CNBHD, indicating that the regulatory mechanism of the eag domain primarily involves the CNBHD. Notably, the structure reveals that a number of LQT2 mutations at homologous positions in human ERG, in addition to cancer-associated mutations in EAG channels, localize to the eag domain-CNBHD interface. Furthermore, mutations at the interface produced marked effects on channel gating, demonstrating the important physiological role of the eag domain-CNBHD interaction. Our structure of the eag domain-CNBHD complex of mouse EAG1 provides unique insights into the physiological and pathophysiological mechanisms of KCNH channels.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3910112/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3910112/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Haitin, Yoni -- Carlson, Anne E -- Zagotta, William N -- F32 HL095241/HL/NHLBI NIH HHS/ -- R01 EY010329/EY/NEI NIH HHS/ -- Howard Hughes Medical Institute/ -- England -- Nature. 2013 Sep 19;501(7467):444-8. doi: 10.1038/nature12487. Epub 2013 Aug 25.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Department of Physiology and Biophysics, University of Washington School of Medicine, Seattle, Washington 98195, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23975098" target="_blank"〉PubMed〈/a〉
    Keywords: Animals ; Binding Sites ; Crystallography, X-Ray ; Ether-A-Go-Go Potassium Channels/*chemistry/genetics/*metabolism ; Humans ; Mice ; Models, Molecular ; Nucleotides, Cyclic/metabolism ; Protein Binding ; Protein Structure, Tertiary ; Static Electricity
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  • 25
    Publication Date: 2013-07-19
    Description: Binding of the glucagon peptide to the glucagon receptor (GCGR) triggers the release of glucose from the liver during fasting; thus GCGR plays an important role in glucose homeostasis. Here we report the crystal structure of the seven transmembrane helical domain of human GCGR at 3.4 A resolution, complemented by extensive site-specific mutagenesis, and a hybrid model of glucagon bound to GCGR to understand the molecular recognition of the receptor for its native ligand. Beyond the shared seven transmembrane fold, the GCGR transmembrane domain deviates from class A G-protein-coupled receptors with a large ligand-binding pocket and the first transmembrane helix having a 'stalk' region that extends three alpha-helical turns above the plane of the membrane. The stalk positions the extracellular domain (~12 kilodaltons) relative to the membrane to form the glucagon-binding site that captures the peptide and facilitates the insertion of glucagon's amino terminus into the seven transmembrane domain.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3820480/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3820480/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Siu, Fai Yiu -- He, Min -- de Graaf, Chris -- Han, Gye Won -- Yang, Dehua -- Zhang, Zhiyun -- Zhou, Caihong -- Xu, Qingping -- Wacker, Daniel -- Joseph, Jeremiah S -- Liu, Wei -- Lau, Jesper -- Cherezov, Vadim -- Katritch, Vsevolod -- Wang, Ming-Wei -- Stevens, Raymond C -- F32 DK088392/DK/NIDDK NIH HHS/ -- P50 GM073197/GM/NIGMS NIH HHS/ -- P50GM073197/GM/NIGMS NIH HHS/ -- U54 GM094586/GM/NIGMS NIH HHS/ -- U54 GM094618/GM/NIGMS NIH HHS/ -- Y1-CO-1020/CO/NCI NIH HHS/ -- Y1-GM-1104/GM/NIGMS NIH HHS/ -- England -- Nature. 2013 Jul 25;499(7459):444-9. doi: 10.1038/nature12393. Epub 2013 Jul 17.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Department of Integrative Structural and Computational Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, California 92037, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23863937" target="_blank"〉PubMed〈/a〉
    Keywords: Amino Acid Sequence ; Binding Sites ; Cell Membrane/metabolism ; Crystallography, X-Ray ; Glucagon/chemistry/metabolism ; Humans ; Ligands ; Models, Molecular ; Molecular Sequence Data ; Mutagenesis, Site-Directed ; Protein Binding ; Protein Structure, Tertiary ; Receptors, CXCR4/chemistry/classification ; Receptors, Glucagon/*chemistry/*classification/genetics/metabolism
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  • 26
    Publication Date: 2013-09-10
    Description: Faithful propagation of DNA methylation patterns during DNA replication is critical for maintaining cellular phenotypes of individual differentiated cells. Although it is well established that Uhrf1 (ubiquitin-like with PHD and ring finger domains 1; also known as Np95 and ICBP90) specifically binds to hemi-methylated DNA through its SRA (SET and RING finger associated) domain and has an essential role in maintenance of DNA methylation by recruiting Dnmt1 to hemi-methylated DNA sites, the mechanism by which Uhrf1 coordinates the maintenance of DNA methylation and DNA replication is largely unknown. Here we show that Uhrf1-dependent histone H3 ubiquitylation has a prerequisite role in the maintenance DNA methylation. Using Xenopus egg extracts, we successfully reproduce maintenance DNA methylation in vitro. Dnmt1 depletion results in a marked accumulation of Uhrf1-dependent ubiquitylation of histone H3 at lysine 23. Dnmt1 preferentially associates with ubiquitylated H3 in vitro though a region previously identified as a replication foci targeting sequence. The RING finger mutant of Uhrf1 fails to recruit Dnmt1 to DNA replication sites and maintain DNA methylation in mammalian cultured cells. Our findings represent the first evidence, to our knowledge, of the mechanistic link between DNA methylation and DNA replication through histone H3 ubiquitylation.〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Nishiyama, Atsuya -- Yamaguchi, Luna -- Sharif, Jafar -- Johmura, Yoshikazu -- Kawamura, Takeshi -- Nakanishi, Keiko -- Shimamura, Shintaro -- Arita, Kyohei -- Kodama, Tatsuhiko -- Ishikawa, Fuyuki -- Koseki, Haruhiko -- Nakanishi, Makoto -- England -- Nature. 2013 Oct 10;502(7470):249-53. doi: 10.1038/nature12488. Epub 2013 Sep 8.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Department of Cell Biology, Graduate School of Medical Sciences, Nagoya City University, Nagoya 467-8601, Japan. anishiya@med.nagoya-cu.ac.jp〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/24013172" target="_blank"〉PubMed〈/a〉
    Keywords: Animals ; Cell Line ; DNA Methylation/genetics/*physiology ; DNA Replication/genetics/*physiology ; HEK293 Cells ; HeLa Cells ; Histones/*metabolism ; Humans ; Mice ; Ovum/chemistry ; Protein Binding ; Ubiquitin-Protein Ligases/genetics/*metabolism ; Ubiquitination ; Xenopus Proteins/genetics/*metabolism ; Xenopus laevis/*genetics/*metabolism
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  • 27
    Publication Date: 2013-10-29
    Description: Members of the CD36 superfamily of scavenger receptor proteins are important regulators of lipid metabolism and innate immunity. They recognize normal and modified lipoproteins, as well as pathogen-associated molecular patterns. The family consists of three members: SR-BI (which delivers cholesterol to the liver and steroidogenic organs and is a co-receptor for hepatitis C virus), LIMP-2/LGP85 (which mediates lysosomal delivery of beta-glucocerebrosidase and serves as a receptor for enterovirus 71 and coxsackieviruses) and CD36 (a fatty-acid transporter and receptor for phagocytosis of effete cells and Plasmodium-infected erythrocytes). Notably, CD36 is also a receptor for modified lipoproteins and beta-amyloid, and has been implicated in the pathogenesis of atherosclerosis and of Alzheimer's disease. Despite their prominent roles in health and disease, understanding the function and abnormalities of the CD36 family members has been hampered by the paucity of information about their structure. Here we determine the crystal structure of LIMP-2 and infer, by homology modelling, the structure of SR-BI and CD36. LIMP-2 shows a helical bundle where beta-glucocerebrosidase binds, and where ligands are most likely to bind to SR-BI and CD36. Remarkably, the crystal structure also shows the existence of a large cavity that traverses the entire length of the molecule. Mutagenesis of SR-BI indicates that the cavity serves as a tunnel through which cholesterol(esters) are delivered from the bound lipoprotein to the outer leaflet of the plasma membrane. We provide evidence supporting a model whereby lipidic constituents of the ligands attached to the receptor surface are handed off to the membrane through the tunnel, accounting for the selective lipid transfer characteristic of SR-BI and CD36.〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Neculai, Dante -- Schwake, Michael -- Ravichandran, Mani -- Zunke, Friederike -- Collins, Richard F -- Peters, Judith -- Neculai, Mirela -- Plumb, Jonathan -- Loppnau, Peter -- Pizarro, Juan Carlos -- Seitova, Alma -- Trimble, William S -- Saftig, Paul -- Grinstein, Sergio -- Dhe-Paganon, Sirano -- MOP-102474/Canadian Institutes of Health Research/Canada -- MOP-126069/Canadian Institutes of Health Research/Canada -- Wellcome Trust/United Kingdom -- England -- Nature. 2013 Dec 5;504(7478):172-6. doi: 10.1038/nature12684. Epub 2013 Oct 27.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Cell Biology Program, The Hospital for Sick Children, Toronto M5G 1X8, Canada.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/24162852" target="_blank"〉PubMed〈/a〉
    Keywords: Animals ; Antigens, CD36/*metabolism ; CHO Cells ; Cricetulus ; HeLa Cells ; Humans ; Lysosome-Associated Membrane Glycoproteins/*chemistry/metabolism ; *Models, Molecular ; Protein Binding ; Protein Structure, Tertiary
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  • 28
    Publication Date: 2013-06-21
    Description: Allostery is an intrinsic property of many globular proteins and enzymes that is indispensable for cellular regulatory and feedback mechanisms. Recent theoretical and empirical observations indicate that allostery is also manifest in intrinsically disordered proteins, which account for a substantial proportion of the proteome. Many intrinsically disordered proteins are promiscuous binders that interact with multiple partners and frequently function as molecular hubs in protein interaction networks. The adenovirus early region 1A (E1A) oncoprotein is a prime example of a molecular hub intrinsically disordered protein. E1A can induce marked epigenetic reprogramming of the cell within hours after infection, through interactions with a diverse set of partners that include key host regulators such as the general transcriptional coactivator CREB binding protein (CBP), its paralogue p300, and the retinoblastoma protein (pRb; also called RB1). Little is known about the allosteric effects at play in E1A-CBP-pRb interactions, or more generally in hub intrinsically disordered protein interaction networks. Here we used single-molecule fluorescence resonance energy transfer (smFRET) to study coupled binding and folding processes in the ternary E1A system. The low concentrations used in these high-sensitivity experiments proved to be essential for these studies, which are challenging owing to a combination of E1A aggregation propensity and high-affinity binding interactions. Our data revealed that E1A-CBP-pRb interactions have either positive or negative cooperativity, depending on the available E1A interaction sites. This striking cooperativity switch enables fine-tuning of the thermodynamic accessibility of the ternary versus binary E1A complexes, and may permit a context-specific tuning of associated downstream signalling outputs. Such a modulation of allosteric interactions is probably a common mechanism in molecular hub intrinsically disordered protein function.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3718496/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3718496/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Ferreon, Allan Chris M -- Ferreon, Josephine C -- Wright, Peter E -- Deniz, Ashok A -- CA96865/CA/NCI NIH HHS/ -- GM066833/GM/NIGMS NIH HHS/ -- R01 CA096865/CA/NCI NIH HHS/ -- R01 GM066833/GM/NIGMS NIH HHS/ -- England -- Nature. 2013 Jun 20;498(7454):390-4. doi: 10.1038/nature12294.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Department of Integrative Structural and Computational Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, California 92037, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23783631" target="_blank"〉PubMed〈/a〉
    Keywords: Adenovirus E1A Proteins/*chemistry/*metabolism ; *Allosteric Regulation ; Amino Acid Motifs ; Animals ; Anisotropy ; CREB-Binding Protein/chemistry/metabolism ; Fluorescence Resonance Energy Transfer ; Humans ; Mice ; Models, Molecular ; Protein Binding ; Protein Folding ; Protein Structure, Tertiary ; Retinoblastoma Protein/chemistry/metabolism ; Thermodynamics ; p300-CBP Transcription Factors/chemistry
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  • 29
    Publication Date: 2013-06-25
    Description: Glutamate transporters are integral membrane proteins that catalyse neurotransmitter uptake from the synaptic cleft into the cytoplasm of glial cells and neurons. Their mechanism of action involves transitions between extracellular (outward)-facing and intracellular (inward)-facing conformations, whereby substrate binding sites become accessible to either side of the membrane. This process has been proposed to entail transmembrane movements of three discrete transport domains within a trimeric scaffold. Using single-molecule fluorescence resonance energy transfer (smFRET) imaging, we have directly observed large-scale transport domain movements in a bacterial homologue of glutamate transporters. We find that individual transport domains alternate between periods of quiescence and periods of rapid transitions, reminiscent of bursting patterns first recorded in single ion channels using patch-clamp methods. We propose that the switch to the dynamic mode in glutamate transporters is due to separation of the transport domain from the trimeric scaffold, which precedes domain movements across the bilayer. This spontaneous dislodging of the substrate-loaded transport domain is approximately 100-fold slower than subsequent transmembrane movements and may be rate determining in the transport cycle.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3829612/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3829612/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Akyuz, Nurunisa -- Altman, Roger B -- Blanchard, Scott C -- Boudker, Olga -- 5U54GM087519/GM/NIGMS NIH HHS/ -- R01 NS064357/NS/NINDS NIH HHS/ -- R01NS064357/NS/NINDS NIH HHS/ -- U54 GM087519/GM/NIGMS NIH HHS/ -- England -- Nature. 2013 Oct 3;502(7469):114-8. doi: 10.1038/nature12265. Epub 2013 Jun 23.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Department of Physiology and Biophysics, Weill Cornell Medical College, 1300 York Avenue, New York, New York 10064, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23792560" target="_blank"〉PubMed〈/a〉
    Keywords: Amino Acid Transport System X-AG/*chemistry/genetics/*metabolism ; Aspartic Acid/chemistry ; Biological Transport ; Fluorescence Resonance Energy Transfer ; *Models, Molecular ; Mutation ; Protein Binding ; Protein Structure, Tertiary ; Pyrococcus horikoshii/chemistry/genetics/*metabolism ; Sodium/chemistry
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  • 30
    Publication Date: 2013-03-29
    Description: Electron transfer reactions are essential for life because they underpin oxidative phosphorylation and photosynthesis, processes leading to the generation of ATP, and are involved in many reactions of intermediary metabolism. Key to these roles is the formation of transient inter-protein electron transfer complexes. The structural basis for the control of specificity between partner proteins is lacking because these weak transient complexes have remained largely intractable for crystallographic studies. Inter-protein electron transfer processes are central to all of the key steps of denitrification, an alternative form of respiration in which bacteria reduce nitrate or nitrite to N2 through the gaseous intermediates nitric oxide (NO) and nitrous oxide (N2O) when oxygen concentrations are limiting. The one-electron reduction of nitrite to NO, a precursor to N2O, is performed by either a haem- or copper-containing nitrite reductase (CuNiR) where they receive an electron from redox partner proteins a cupredoxin or a c-type cytochrome. Here we report the structures of the newly characterized three-domain haem-c-Cu nitrite reductase from Ralstonia pickettii (RpNiR) at 1.01 A resolution and its M92A and P93A mutants. Very high resolution provides the first view of the atomic detail of the interface between the core trimeric cupredoxin structure of CuNiR and the tethered cytochrome c domain that allows the enzyme to function as an effective self-electron transfer system where the donor and acceptor proteins are fused together by genomic acquisition for functional advantage. Comparison of RpNiR with the binary complex of a CuNiR with a donor protein, AxNiR-cytc551 (ref. 6), and mutagenesis studies provide direct evidence for the importance of a hydrogen-bonded water at the interface in electron transfer. The structure also provides an explanation for the preferential binding of nitrite to the reduced copper ion at the active site in RpNiR, in contrast to other CuNiRs where reductive inactivation occurs, preventing substrate binding.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3672994/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3672994/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Antonyuk, Svetlana V -- Han, Cong -- Eady, Robert R -- Hasnain, S Samar -- 097826/Z/11/Z/Wellcome Trust/United Kingdom -- BB/G005869/1/Biotechnology and Biological Sciences Research Council/United Kingdom -- England -- Nature. 2013 Apr 4;496(7443):123-6. doi: 10.1038/nature11996. Epub 2013 Mar 27.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Molecular Biophysics Group, Institute of Integrative Biology, Faculty of Health and Life Sciences, University of Liverpool, Liverpool L69 7ZX, UK.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23535590" target="_blank"〉PubMed〈/a〉
    Keywords: Azurin/chemistry/metabolism ; Catalytic Domain ; Copper/chemistry/metabolism ; Cytochromes c/chemistry/metabolism ; *Electron Transport ; Hydrogen Bonding ; Models, Molecular ; Mutant Proteins/chemistry/genetics/metabolism ; Nitrite Reductases/*chemistry/genetics/*metabolism ; Nitrites/chemistry/metabolism ; Protein Binding ; Protein Structure, Tertiary ; Protons ; Ralstonia pickettii/*enzymology ; Water/chemistry/metabolism
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  • 31
    Publication Date: 2013-03-05
    Description: P-type ATPases are ATP-powered ion pumps that establish ion concentration gradients across biological membranes, and are distinct from other ATPases in that the reaction cycle includes an autophosphorylation step. The best studied is Ca(2+)-ATPase from muscle sarcoplasmic reticulum (SERCA1a), a Ca(2+) pump that relaxes muscle cells after contraction, and crystal structures have been determined for most of the reaction intermediates. An important outstanding structure is that of the E1 intermediate, which has empty high-affinity Ca(2+)-binding sites ready to accept new cytosolic Ca(2+). In the absence of Ca(2+) and at pH 7 or higher, the ATPase is predominantly in E1, not in E2 (low affinity for Ca(2+)), and if millimolar Mg(2+) is present, one Mg(2+) is expected to occupy one of the Ca(2+)-binding sites with a millimolar dissociation constant. This Mg(2+) accelerates the reaction cycle, not permitting phosphorylation without Ca(2+) binding. Here we describe the crystal structure of native SERCA1a (from rabbit) in this E1.Mg(2+) state at 3.0 A resolution in addition to crystal structures of SERCA1a in E2 free from exogenous inhibitors, and address the structural basis of the activation signal for phosphoryl transfer. Unexpectedly, sarcolipin, a small regulatory membrane protein of Ca(2+)-ATPase, is bound, stabilizing the E1.Mg(2+) state. Sarcolipin is a close homologue of phospholamban, which is a critical mediator of beta-adrenergic signal in Ca(2+) regulation in heart (for reviews, see, for example, refs 8-10), and seems to play an important role in muscle-based thermogenesis. We also determined the crystal structure of recombinant SERCA1a devoid of sarcolipin, and describe the structural basis of inhibition by sarcolipin/phospholamban. Thus, the crystal structures reported here fill a gap in the structural elucidation of the reaction cycle and provide a solid basis for understanding the physiological regulation of the calcium pump.〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Toyoshima, Chikashi -- Iwasawa, Shiho -- Ogawa, Haruo -- Hirata, Ayami -- Tsueda, Junko -- Inesi, Giuseppe -- England -- Nature. 2013 Mar 14;495(7440):260-4. doi: 10.1038/nature11899. Epub 2013 Mar 3.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Institute of Molecular and Cellular Biosciences, The University of Tokyo, Bunkyo-ku, Tokyo 113-0032, Japan. ct@iam.u-tokyo.ac.jp〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23455422" target="_blank"〉PubMed〈/a〉
    Keywords: Animals ; Binding Sites/drug effects ; Calcium-Binding Proteins/pharmacology ; Cell Membrane/metabolism ; Crystallography, X-Ray ; Magnesium/chemistry/*metabolism/pharmacology ; Models, Molecular ; Muscle Proteins/*chemistry/*metabolism/pharmacology ; Phosphorylation ; Protein Binding ; Protein Conformation/drug effects ; Proteolipids/*chemistry/*metabolism/pharmacology ; Rabbits ; Sarcoplasmic Reticulum Calcium-Transporting ATPases/antagonists & ; inhibitors/*chemistry/*metabolism
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  • 32
    Publication Date: 2013-09-06
    Description: The ability to design proteins with high affinity and selectivity for any given small molecule is a rigorous test of our understanding of the physiochemical principles that govern molecular recognition. Attempts to rationally design ligand-binding proteins have met with little success, however, and the computational design of protein-small-molecule interfaces remains an unsolved problem. Current approaches for designing ligand-binding proteins for medical and biotechnological uses rely on raising antibodies against a target antigen in immunized animals and/or performing laboratory-directed evolution of proteins with an existing low affinity for the desired ligand, neither of which allows complete control over the interactions involved in binding. Here we describe a general computational method for designing pre-organized and shape complementary small-molecule-binding sites, and use it to generate protein binders to the steroid digoxigenin (DIG). Of seventeen experimentally characterized designs, two bind DIG; the model of the higher affinity binder has the most energetically favourable and pre-organized interface in the design set. A comprehensive binding-fitness landscape of this design, generated by library selections and deep sequencing, was used to optimize its binding affinity to a picomolar level, and X-ray co-crystal structures of two variants show atomic-level agreement with the corresponding computational models. The optimized binder is selective for DIG over the related steroids digitoxigenin, progesterone and beta-oestradiol, and this steroid binding preference can be reprogrammed by manipulation of explicitly designed hydrogen-bonding interactions. The computational design method presented here should enable the development of a new generation of biosensors, therapeutics and diagnostics.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3898436/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3898436/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Tinberg, Christine E -- Khare, Sagar D -- Dou, Jiayi -- Doyle, Lindsey -- Nelson, Jorgen W -- Schena, Alberto -- Jankowski, Wojciech -- Kalodimos, Charalampos G -- Johnsson, Kai -- Stoddard, Barry L -- Baker, David -- P41 GM103533/GM/NIGMS NIH HHS/ -- R01 GM049857/GM/NIGMS NIH HHS/ -- T32 HG000035/HG/NHGRI NIH HHS/ -- T32 HG00035/HG/NHGRI NIH HHS/ -- England -- Nature. 2013 Sep 12;501(7466):212-6. doi: 10.1038/nature12443. Epub 2013 Sep 4.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Department of Biochemistry, University of Washington, Seattle, Washington 98195, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/24005320" target="_blank"〉PubMed〈/a〉
    Keywords: Binding Sites ; Biotechnology ; *Computer Simulation ; Crystallography, X-Ray ; Digoxigenin/chemistry/*metabolism ; *Drug Design ; Estradiol/chemistry/metabolism ; Ligands ; Models, Molecular ; Progesterone/chemistry/metabolism ; Protein Binding ; Proteins/*chemistry/*metabolism ; Reproducibility of Results ; Substrate Specificity
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  • 33
    Publication Date: 2013-04-23
    Description: The functions of G-protein-coupled receptors (GPCRs) are primarily mediated and modulated by three families of proteins: the heterotrimeric G proteins, the G-protein-coupled receptor kinases (GRKs) and the arrestins. G proteins mediate activation of second-messenger-generating enzymes and other effectors, GRKs phosphorylate activated receptors, and arrestins subsequently bind phosphorylated receptors and cause receptor desensitization. Arrestins activated by interaction with phosphorylated receptors can also mediate G-protein-independent signalling by serving as adaptors to link receptors to numerous signalling pathways. Despite their central role in regulation and signalling of GPCRs, a structural understanding of beta-arrestin activation and interaction with GPCRs is still lacking. Here we report the crystal structure of beta-arrestin-1 (also called arrestin-2) in complex with a fully phosphorylated 29-amino-acid carboxy-terminal peptide derived from the human V2 vasopressin receptor (V2Rpp). This peptide has previously been shown to functionally and conformationally activate beta-arrestin-1 (ref. 5). To capture this active conformation, we used a conformationally selective synthetic antibody fragment (Fab30) that recognizes the phosphopeptide-activated state of beta-arrestin-1. The structure of the beta-arrestin-1-V2Rpp-Fab30 complex shows marked conformational differences in beta-arrestin-1 compared to its inactive conformation. These include rotation of the amino- and carboxy-terminal domains relative to each other, and a major reorientation of the 'lariat loop' implicated in maintaining the inactive state of beta-arrestin-1. These results reveal, at high resolution, a receptor-interacting interface on beta-arrestin, and they indicate a potentially general molecular mechanism for activation of these multifunctional signalling and regulatory proteins.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3654799/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3654799/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Shukla, Arun K -- Manglik, Aashish -- Kruse, Andrew C -- Xiao, Kunhong -- Reis, Rosana I -- Tseng, Wei-Chou -- Staus, Dean P -- Hilger, Daniel -- Uysal, Serdar -- Huang, Li-Yin -- Paduch, Marcin -- Tripathi-Shukla, Prachi -- Koide, Akiko -- Koide, Shohei -- Weis, William I -- Kossiakoff, Anthony A -- Kobilka, Brian K -- Lefkowitz, Robert J -- GM072688/GM/NIGMS NIH HHS/ -- GM087519/GM/NIGMS NIH HHS/ -- HL 075443/HL/NHLBI NIH HHS/ -- HL16037/HL/NHLBI NIH HHS/ -- HL70631/HL/NHLBI NIH HHS/ -- NS028471/NS/NINDS NIH HHS/ -- P41 RR011823/RR/NCRR NIH HHS/ -- R01 HL016037/HL/NHLBI NIH HHS/ -- R01 HL070631/HL/NHLBI NIH HHS/ -- R01 NS028471/NS/NINDS NIH HHS/ -- U01 GM094588/GM/NIGMS NIH HHS/ -- U54 GM074946/GM/NIGMS NIH HHS/ -- Howard Hughes Medical Institute/ -- England -- Nature. 2013 May 2;497(7447):137-41. doi: 10.1038/nature12120. Epub 2013 Apr 21.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Department of Medicine, Duke University Medical Center, Durham, North Carolina 27710, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23604254" target="_blank"〉PubMed〈/a〉
    Keywords: Animals ; Arrestins/*chemistry/immunology/*metabolism ; Crystallography, X-Ray ; Humans ; Immunoglobulin Fab Fragments/chemistry/immunology/metabolism ; Models, Molecular ; Phosphopeptides/*chemistry/*metabolism ; Phosphorylation ; Protein Binding ; Protein Conformation ; Protein Stability ; Rats ; Receptors, Vasopressin/*chemistry ; Rotation
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  • 34
    Publication Date: 2013-01-11
    Description: Insulin receptor signalling has a central role in mammalian biology, regulating cellular metabolism, growth, division, differentiation and survival. Insulin resistance contributes to the pathogenesis of type 2 diabetes mellitus and the onset of Alzheimer's disease; aberrant signalling occurs in diverse cancers, exacerbated by cross-talk with the homologous type 1 insulin-like growth factor receptor (IGF1R). Despite more than three decades of investigation, the three-dimensional structure of the insulin-insulin receptor complex has proved elusive, confounded by the complexity of producing the receptor protein. Here we present the first view, to our knowledge, of the interaction of insulin with its primary binding site on the insulin receptor, on the basis of four crystal structures of insulin bound to truncated insulin receptor constructs. The direct interaction of insulin with the first leucine-rich-repeat domain (L1) of insulin receptor is seen to be sparse, the hormone instead engaging the insulin receptor carboxy-terminal alpha-chain (alphaCT) segment, which is itself remodelled on the face of L1 upon insulin binding. Contact between insulin and L1 is restricted to insulin B-chain residues. The alphaCT segment displaces the B-chain C-terminal beta-strand away from the hormone core, revealing the mechanism of a long-proposed conformational switch in insulin upon receptor engagement. This mode of hormone-receptor recognition is novel within the broader family of receptor tyrosine kinases. We support these findings by photo-crosslinking data that place the suggested interactions into the context of the holoreceptor and by isothermal titration calorimetry data that dissect the hormone-insulin receptor interface. Together, our findings provide an explanation for a wealth of biochemical data from the insulin receptor and IGF1R systems relevant to the design of therapeutic insulin analogues.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3793637/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3793637/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Menting, John G -- Whittaker, Jonathan -- Margetts, Mai B -- Whittaker, Linda J -- Kong, Geoffrey K-W -- Smith, Brian J -- Watson, Christopher J -- Zakova, Lenka -- Kletvikova, Emilia -- Jiracek, Jiri -- Chan, Shu Jin -- Steiner, Donald F -- Dodson, Guy G -- Brzozowski, Andrzej M -- Weiss, Michael A -- Ward, Colin W -- Lawrence, Michael C -- DK13914/DK/NIDDK NIH HHS/ -- DK20595/DK/NIDDK NIH HHS/ -- DK40949/DK/NIDDK NIH HHS/ -- R01 DK040949/DK/NIDDK NIH HHS/ -- UL1 TR000439/TR/NCATS NIH HHS/ -- Biotechnology and Biological Sciences Research Council/United Kingdom -- England -- Nature. 2013 Jan 10;493(7431):241-5. doi: 10.1038/nature11781.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Walter and Eliza Hall Institute of Medical Research, 1G Royal Parade, Parkville, Victoria 3052, Australia.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23302862" target="_blank"〉PubMed〈/a〉
    Keywords: Animals ; Binding Sites ; Calorimetry ; Cattle ; Cell Line ; Crystallography, X-Ray ; Humans ; Insulin/*chemistry/*metabolism ; Leucine/metabolism ; Ligands ; Models, Molecular ; Protein Binding ; Protein Structure, Secondary ; Receptor, Insulin/*chemistry/*metabolism ; Reproducibility of Results
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  • 35
    Publication Date: 2013-09-27
    Description: The appropriate timing of flowering is crucial for plant reproductive success. It is therefore not surprising that intricate genetic networks have evolved to perceive and integrate both endogenous and environmental signals, such as carbohydrate and hormonal status, photoperiod and temperature. In contrast to our detailed understanding of the vernalization pathway, little is known about how flowering time is controlled in response to changes in the ambient growth temperature. In Arabidopsis thaliana, the MADS-box transcription factor genes FLOWERING LOCUS M (FLM) and SHORT VEGETATIVE PHASE (SVP) have key roles in this process. FLM is subject to temperature-dependent alternative splicing. Here we report that the two main FLM protein splice variants, FLM-beta and FLM-delta, compete for interaction with the floral repressor SVP. The SVP-FLM-beta complex is predominately formed at low temperatures and prevents precocious flowering. By contrast, the competing SVP-FLM-delta complex is impaired in DNA binding and acts as a dominant-negative activator of flowering at higher temperatures. Our results show a new mechanism that controls the timing of the floral transition in response to changes in ambient temperature. A better understanding of how temperature controls the molecular mechanisms of flowering will be important to cope with current changes in global climate.〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Pose, David -- Verhage, Leonie -- Ott, Felix -- Yant, Levi -- Mathieu, Johannes -- Angenent, Gerco C -- Immink, Richard G H -- Schmid, Markus -- England -- Nature. 2013 Nov 21;503(7476):414-7. doi: 10.1038/nature12633. Epub 2013 Sep 25.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉1] Max Planck Institute for Developmental Biology, Department of Molecular Biology, Spemannstr. 35, 72076 Tubingen, Germany [2] Instituto de Hortofruticultura Subtropical y Mediterranea, Universidad de Malaga-Consejo Superior de Investigaciones Cientificas, Departamento de Biologia Molecular y Bioquimica, Facultad de Ciencias, Universidad de Malaga, 29071 Malaga, Spain (D.P.); Department of Organismic and Evolutionary Biology, Harvard University, 16 Divinity Avenue, Cambridge, Massachusetts 02138, USA (L.Y.); Boyce Thompson Institute for Plant Research, Tower Road, Ithaca, New York 14853-1801, USA (J.M.).〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/24067612" target="_blank"〉PubMed〈/a〉
    Keywords: Alternative Splicing/*genetics ; Arabidopsis/genetics/*physiology ; Arabidopsis Proteins/chemistry/genetics/*metabolism ; DNA-Binding Proteins/chemistry/genetics/metabolism ; Flowers/genetics/*physiology ; Gene Expression Regulation, Plant ; MADS Domain Proteins/chemistry/genetics/*metabolism ; Plants, Genetically Modified ; Protein Binding ; Protein Isoforms/chemistry/genetics/*metabolism ; *Temperature ; Time Factors ; Transcription Factors/metabolism
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  • 36
    Publication Date: 2013-12-20
    Description: Nucleosomes are decorated with numerous post-translational modifications capable of influencing many DNA processes. Here we describe a new class of histone modification, methylation of glutamine, occurring on yeast histone H2A at position 105 (Q105) and human H2A at Q104. We identify Nop1 as the methyltransferase in yeast and demonstrate that fibrillarin is the orthologue enzyme in human cells. Glutamine methylation of H2A is restricted to the nucleolus. Global analysis in yeast, using an H2AQ105me-specific antibody, shows that this modification is exclusively enriched over the 35S ribosomal DNA transcriptional unit. We show that the Q105 residue is part of the binding site for the histone chaperone FACT (facilitator of chromatin transcription) complex. Methylation of Q105 or its substitution to alanine disrupts binding to FACT in vitro. A yeast strain mutated at Q105 shows reduced histone incorporation and increased transcription at the ribosomal DNA locus. These features are phenocopied by mutations in FACT complex components. Together these data identify glutamine methylation of H2A as the first histone epigenetic mark dedicated to a specific RNA polymerase and define its function as a regulator of FACT interaction with nucleosomes.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3901671/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3901671/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Tessarz, Peter -- Santos-Rosa, Helena -- Robson, Sam C -- Sylvestersen, Kathrine B -- Nelson, Christopher J -- Nielsen, Michael L -- Kouzarides, Tony -- 092096/Wellcome Trust/United Kingdom -- A10827/Cancer Research UK/United Kingdom -- BB/K017438/1/Biotechnology and Biological Sciences Research Council/United Kingdom -- England -- Nature. 2014 Jan 23;505(7484):564-8. doi: 10.1038/nature12819. Epub 2013 Dec 18.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉1] Gurdon Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QN, UK [2] Department of Pathology, University of Cambridge, Tennis Court Road, Cambridge CB2 1QN, UK. ; Department of Proteomics, The Novo Nordisk Foundation Center for Protein Research, Faculty of Health Sciences, University of Copenhagen, Blegdamsvej 3B, DK-2200 Copenhagen, Denmark. ; 1] Gurdon Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QN, UK [2] Department of Pathology, University of Cambridge, Tennis Court Road, Cambridge CB2 1QN, UK [3] Department of Biochemistry and Microbiology, University of Victoria, 3800 Finnerty Road, Victoria, British Columbia V8P 5C2, Canada.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/24352239" target="_blank"〉PubMed〈/a〉
    Keywords: Alanine/genetics/metabolism ; Amino Acid Motifs ; Amino Acid Sequence ; Binding Sites ; Cell Nucleolus/metabolism ; Chromatin/genetics ; Chromosomal Proteins, Non-Histone/metabolism ; DNA, Ribosomal/genetics ; Epistasis, Genetic ; Glutamine/*metabolism ; Histones/*chemistry/*metabolism ; Humans ; Methylation ; Methyltransferases/metabolism ; Molecular Chaperones/metabolism ; Molecular Sequence Data ; Multiprotein Complexes/metabolism ; Nuclear Proteins/metabolism ; Nucleosomes/metabolism ; Protein Binding ; Protein Processing, Post-Translational ; RNA/metabolism ; RNA Polymerase I/*metabolism ; Ribonucleoproteins, Small Nucleolar/metabolism ; Saccharomyces cerevisiae/enzymology/genetics/metabolism ; Saccharomyces cerevisiae Proteins/metabolism ; Substrate Specificity ; Transcription, Genetic
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  • 37
    Publication Date: 2013-12-24
    Description: Acyl carrier protein (ACP) transports the growing fatty acid chain between enzymatic domains of fatty acid synthase (FAS) during biosynthesis. Because FAS enzymes operate on ACP-bound acyl groups, ACP must stabilize and transport the growing lipid chain. ACPs have a central role in transporting starting materials and intermediates throughout the fatty acid biosynthetic pathway. The transient nature of ACP-enzyme interactions impose major obstacles to obtaining high-resolution structural information about fatty acid biosynthesis, and a new strategy is required to study protein-protein interactions effectively. Here we describe the application of a mechanism-based probe that allows active site-selective covalent crosslinking of AcpP to FabA, the Escherichia coli ACP and fatty acid 3-hydroxyacyl-ACP dehydratase, respectively. We report the 1.9 A crystal structure of the crosslinked AcpP-FabA complex as a homodimer in which AcpP exhibits two different conformations, representing probable snapshots of ACP in action: the 4'-phosphopantetheine group of AcpP first binds an arginine-rich groove of FabA, then an AcpP helical conformational change locks AcpP and FabA in place. Residues at the interface of AcpP and FabA are identified and validated by solution nuclear magnetic resonance techniques, including chemical shift perturbations and residual dipolar coupling measurements. These not only support our interpretation of the crystal structures but also provide an animated view of ACP in action during fatty acid dehydration. These techniques, in combination with molecular dynamics simulations, show for the first time that FabA extrudes the sequestered acyl chain from the ACP binding pocket before dehydration by repositioning helix III. Extensive sequence conservation among carrier proteins suggests that the mechanistic insights gleaned from our studies may be broadly applicable to fatty acid, polyketide and non-ribosomal biosynthesis. Here the foundation is laid for defining the dynamic action of carrier-protein activity in primary and secondary metabolism, providing insight into pathways that can have major roles in the treatment of cancer, obesity and infectious disease.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4437705/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4437705/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Nguyen, Chi -- Haushalter, Robert W -- Lee, D John -- Markwick, Phineus R L -- Bruegger, Joel -- Caldara-Festin, Grace -- Finzel, Kara -- Jackson, David R -- Ishikawa, Fumihiro -- O'Dowd, Bing -- McCammon, J Andrew -- Opella, Stanley J -- Tsai, Shiou-Chuan -- Burkart, Michael D -- GM095970/GM/NIGMS NIH HHS/ -- GM100305/GM/NIGMS NIH HHS/ -- P41 EB002031/EB/NIBIB NIH HHS/ -- R01 GM066978/GM/NIGMS NIH HHS/ -- R01 GM095970/GM/NIGMS NIH HHS/ -- R01 GM100305/GM/NIGMS NIH HHS/ -- Howard Hughes Medical Institute/ -- England -- Nature. 2014 Jan 16;505(7483):427-31. doi: 10.1038/nature12810. Epub 2013 Dec 22.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉1] Departments of Molecular Biology and Biochemistry, Chemistry, and Pharmaceutical Sciences, University of California, Irvine, California 92697, USA [2]. ; 1] Department of Chemistry and Biochemistry, University of California, San Diego, La Jolla, California 92093, USA [2]. ; 1] Department of Chemistry and Biochemistry, University of California, San Diego, La Jolla, California 92093, USA [2] San Diego Supercomputer Center, La Jolla, California 92093, USA [3] Howard Hughes Medical Institute, La Jolla, California 92093, USA. ; Departments of Molecular Biology and Biochemistry, Chemistry, and Pharmaceutical Sciences, University of California, Irvine, California 92697, USA. ; Department of Chemistry and Biochemistry, University of California, San Diego, La Jolla, California 92093, USA. ; 1] Department of Chemistry and Biochemistry, University of California, San Diego, La Jolla, California 92093, USA [2] Howard Hughes Medical Institute, La Jolla, California 92093, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/24362570" target="_blank"〉PubMed〈/a〉
    Keywords: Acyl Carrier Protein/*chemistry/*metabolism ; Binding Sites ; Catalytic Domain ; Cross-Linking Reagents/chemistry ; Crystallography, X-Ray ; Escherichia coli/*chemistry ; Fatty Acid Synthase, Type II/chemistry/metabolism ; Fatty Acids/*biosynthesis ; Histidine/metabolism ; Hydro-Lyases/chemistry/metabolism ; Magnetic Resonance Spectroscopy ; Models, Molecular ; Molecular Dynamics Simulation ; Protein Binding ; Protein Interaction Maps
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  • 38
    Publication Date: 2013-12-03
    Description: Cohesion between sister chromatids, mediated by the chromosomal cohesin complex, is a prerequisite for faithful chromosome segregation in mitosis. Cohesin also has vital roles in DNA repair and transcriptional regulation. The ring-shaped cohesin complex is thought to encircle sister DNA strands, but its molecular mechanism of action is poorly understood and the biochemical reconstitution of cohesin activity in vitro has remained an unattained goal. Here we reconstitute cohesin loading onto DNA using purified fission yeast cohesin and its loader complex, Mis4(Scc2)-Ssl3(Scc4) (Schizosaccharomyces pombe gene names appear throughout with their more commonly known Saccharomyces cerevisiae counterparts added in superscript). Incubation of cohesin with DNA leads to spontaneous topological loading, but this remains inefficient. The loader contacts cohesin at multiple sites around the ring circumference, including the hitherto enigmatic Psc3(Scc3) subunit, and stimulates cohesin's ATPase, resulting in efficient topological loading. The in vitro reconstitution of cohesin loading onto DNA provides mechanistic insight into the initial steps of the establishment of sister chromatid cohesion and other chromosomal processes mediated by cohesin.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3907785/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3907785/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Murayama, Yasuto -- Uhlmann, Frank -- A3592/Cancer Research UK/United Kingdom -- England -- Nature. 2014 Jan 16;505(7483):367-71. doi: 10.1038/nature12867. Epub 2013 Dec 1.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Chromosome Segregation Laboratory, Cancer Research UK London Research Institute, 44 Lincoln's Inn Fields, London WC2A 3LY, UK.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/24291789" target="_blank"〉PubMed〈/a〉
    Keywords: Adenosine Triphosphatases/metabolism ; Cell Cycle Proteins/*chemistry/*metabolism ; Chromatids/genetics/metabolism ; Chromosomal Proteins, Non-Histone/*chemistry/*metabolism ; DNA/*chemistry/*metabolism ; DNA-Binding Proteins/metabolism ; *Nucleic Acid Conformation ; Protein Binding ; Schizosaccharomyces/cytology/*genetics/*metabolism ; Schizosaccharomyces pombe Proteins/metabolism
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  • 39
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    Unknown
    Nature Publishing Group (NPG)
    Publication Date: 2013-11-29
    Description: 〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Reardon, Sara -- England -- Nature. 2013 Nov 28;503(7477):449-50. doi: 10.1038/503449a.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/24284710" target="_blank"〉PubMed〈/a〉
    Keywords: *Computational Biology ; *Computer Simulation ; Databases, Chemical ; Drug Discovery/*methods ; Humans ; Models, Molecular ; Molecular Targeted Therapy/*methods ; Pharmaceutical Preparations/chemistry/*metabolism ; Protein Binding ; Proteins/chemistry/*metabolism
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  • 40
    Publication Date: 2013-06-22
    Description: Of the 132 people known to have been infected with H7N9 influenza viruses in China, 37 died, and many were severely ill. Infection seems to have involved contact with infected poultry. We have examined the receptor-binding properties of this H7N9 virus and compared them with those of an avian H7N3 virus. We find that the human H7 virus has significantly higher affinity for alpha-2,6-linked sialic acid analogues ('human receptor') than avian H7 while retaining the strong binding to alpha-2,3-linked sialic acid analogues ('avian receptor') characteristic of avian viruses. The human H7 virus does not, therefore, have the preference for human versus avian receptors characteristic of pandemic viruses. X-ray crystallography of the receptor-binding protein, haemagglutinin (HA), in complex with receptor analogues indicates that both human and avian receptors adopt different conformations when bound to human H7 HA than they do when bound to avian H7 HA. Human receptor bound to human H7 HA exits the binding site in a different direction to that seen in complexes formed by HAs from pandemic viruses and from an aerosol-transmissible H5 mutant. The human-receptor-binding properties of human H7 probably arise from the introduction of two bulky hydrophobic residues by the substitutions Gln226Leu and Gly186Val. The former is shared with the 1957 H2 and 1968 H3 pandemic viruses and with the aerosol-transmissible H5 mutant. We conclude that the human H7 virus has acquired some of the receptor-binding characteristics that are typical of pandemic viruses, but its retained preference for avian receptor may restrict its further evolution towards a virus that could transmit efficiently between humans, perhaps by binding to avian-receptor-rich mucins in the human respiratory tract rather than to cellular receptors.〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Xiong, Xiaoli -- Martin, Stephen R -- Haire, Lesley F -- Wharton, Stephen A -- Daniels, Rodney S -- Bennett, Michael S -- McCauley, John W -- Collins, Patrick J -- Walker, Philip A -- Skehel, John J -- Gamblin, Steven J -- MC_U117584222/Medical Research Council/United Kingdom -- MC_U117585868/Medical Research Council/United Kingdom -- U117512723/PHS HHS/ -- U117570592/PHS HHS/ -- U117584222/PHS HHS/ -- U117585868/PHS HHS/ -- England -- Nature. 2013 Jul 25;499(7459):496-9. doi: 10.1038/nature12372.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉MRC National Institute for Medical Research, The Ridgeway, Mill Hill, London NW71AA, UK.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23787694" target="_blank"〉PubMed〈/a〉
    Keywords: Animals ; Binding Sites ; Birds/metabolism/virology ; Crystallography, X-Ray ; Hemagglutinin Glycoproteins, Influenza Virus/chemistry/metabolism ; Humans ; Influenza A Virus, H7N3 Subtype/metabolism ; Influenza A virus/chemistry/isolation & purification/*metabolism ; Influenza, Human/*virology ; Models, Molecular ; Mucins/chemistry/metabolism ; N-Acetylneuraminic Acid/analogs & derivatives/chemistry/*metabolism ; Protein Binding ; Protein Conformation ; Receptors, Virus/chemistry/*metabolism
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  • 41
    Publication Date: 2013-07-03
    Description: Transcription factors are frequently altered in leukaemia through chromosomal translocation, mutation or aberrant expression. AML1-ETO, a fusion protein generated by the t(8;21) translocation in acute myeloid leukaemia, is a transcription factor implicated in both gene repression and activation. AML1-ETO oligomerization, mediated by the NHR2 domain, is critical for leukaemogenesis, making it important to identify co-regulatory factors that 'read' the NHR2 oligomerization and contribute to leukaemogenesis. Here we show that, in human leukaemic cells, AML1-ETO resides in and functions through a stable AML1-ETO-containing transcription factor complex (AETFC) that contains several haematopoietic transcription (co)factors. These AETFC components stabilize the complex through multivalent interactions, provide multiple DNA-binding domains for diverse target genes, co-localize genome wide, cooperatively regulate gene expression, and contribute to leukaemogenesis. Within the AETFC complex, AML1-ETO oligomerization is required for a specific interaction between the oligomerized NHR2 domain and a novel NHR2-binding (N2B) motif in E proteins. Crystallographic analysis of the NHR2-N2B complex reveals a unique interaction pattern in which an N2B peptide makes direct contact with side chains of two NHR2 domains as a dimer, providing a novel model of how dimeric/oligomeric transcription factors create a new protein-binding interface through dimerization/oligomerization. Intriguingly, disruption of this interaction by point mutations abrogates AML1-ETO-induced haematopoietic stem/progenitor cell self-renewal and leukaemogenesis. These results reveal new mechanisms of action of AML1-ETO, and provide a potential therapeutic target in t(8;21)-positive acute myeloid leukaemia.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3732535/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3732535/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Sun, Xiao-Jian -- Wang, Zhanxin -- Wang, Lan -- Jiang, Yanwen -- Kost, Nils -- Soong, T David -- Chen, Wei-Yi -- Tang, Zhanyun -- Nakadai, Tomoyoshi -- Elemento, Olivier -- Fischle, Wolfgang -- Melnick, Ari -- Patel, Dinshaw J -- Nimer, Stephen D -- Roeder, Robert G -- CA113872/CA/NCI NIH HHS/ -- CA129325/CA/NCI NIH HHS/ -- CA163086/CA/NCI NIH HHS/ -- CA166835/CA/NCI NIH HHS/ -- R01 CA163086/CA/NCI NIH HHS/ -- R01 CA166835/CA/NCI NIH HHS/ -- UL1 RR024143/RR/NCRR NIH HHS/ -- UL1RR024143/RR/NCRR NIH HHS/ -- England -- Nature. 2013 Aug 1;500(7460):93-7. doi: 10.1038/nature12287. Epub 2013 Jun 30.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Laboratory of Biochemistry and Molecular Biology, The Rockefeller University, New York, New York 10065, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23812588" target="_blank"〉PubMed〈/a〉
    Keywords: Amino Acid Motifs ; Amino Acid Sequence ; Animals ; Binding Sites ; Cell Division ; Cell Line, Tumor ; *Cell Transformation, Neoplastic/genetics ; Core Binding Factor Alpha 2 Subunit/chemistry/*metabolism ; Hematopoietic Stem Cells/cytology/metabolism/pathology ; Humans ; Leukemia, Myeloid, Acute/genetics/*metabolism/*pathology ; Mice ; Models, Molecular ; Molecular Sequence Data ; Multiprotein Complexes/chemistry/*metabolism ; Oncogene Proteins, Fusion/chemistry/*metabolism ; Point Mutation ; Protein Binding ; Protein Multimerization ; Protein Stability ; Protein Structure, Tertiary ; Transcription Factors/*metabolism
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  • 42
    Publication Date: 2013-03-15
    Description: The hepatocyte nuclear factor 4alpha (HNF-4alpha; also known as NR2A1) is a member of the nuclear receptor (NR) family of transcription factors, which have conserved DNA-binding domains and ligand-binding domains. HNF-4alpha is the most abundant DNA-binding protein in the liver, where some 40% of the actively transcribed genes have a HNF-4alpha response element. These regulated genes are largely involved in the hepatic gluconeogenic program and lipid metabolism. In the pancreas HNF-4alpha is also a master regulator, controlling an estimated 11% of islet genes. HNF-4alpha protein mutations are linked to maturity-onset diabetes of the young, type 1 (MODY1) and hyperinsulinaemic hypoglycaemia. Previous structural analyses of NRs, although productive in elucidating the structure of individual domains, have lagged behind in revealing the connectivity patterns of NR domains. Here we describe the 2.9 A crystal structure of the multidomain human HNF-4alpha homodimer bound to its DNA response element and coactivator-derived peptides. A convergence zone connects multiple receptor domains in an asymmetric fashion, joining distinct elements from each monomer. An arginine target of PRMT1 methylation protrudes directly into this convergence zone and sustains its integrity. A serine target of protein kinase C is also responsible for maintaining domain-domain interactions. These post-translational modifications lead to changes in DNA binding by communicating through the tightly connected surfaces of the quaternary fold. We find that some MODY1 mutations, positioned on the ligand-binding domain and hinge regions of the receptor, compromise DNA binding at a distance by communicating through the interjunctional surfaces of the complex. The overall domain representation of the HNF-4alpha homodimer is different from that of the PPAR-gamma-RXR-alpha heterodimer, even when both NR complexes are assembled on the same DNA element. Our findings suggest that unique quaternary folds and interdomain connections in NRs could be exploited by small-molecule allosteric modulators that affect distal functions in these polypeptides.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3606643/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3606643/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Chandra, Vikas -- Huang, Pengxiang -- Potluri, Nalini -- Wu, Dalei -- Kim, Youngchang -- Rastinejad, Fraydoon -- R01 DK094147/DK/NIDDK NIH HHS/ -- R01 DK097475/DK/NIDDK NIH HHS/ -- England -- Nature. 2013 Mar 21;495(7441):394-8. doi: 10.1038/nature11966. Epub 2013 Mar 13.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Metabolic Signaling and Disease Program, Sanford-Burnham Medical Research Institute, Orlando, Florida 32827, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23485969" target="_blank"〉PubMed〈/a〉
    Keywords: Hepatocyte Nuclear Factor 4/*chemistry/genetics/metabolism ; Humans ; Hypoglycemia/genetics ; *Models, Molecular ; Mutation ; Point Mutation ; Protein Binding ; Protein Structure, Quaternary ; Protein Structure, Tertiary
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  • 43
    Publication Date: 2013-02-01
    Description: 〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Foulquier, Sebastien -- Steckelings, Ulrike Muscha -- Unger, Thomas -- England -- Nature. 2013 Jan 31;493(7434):S9. doi: 10.1038/493S9a.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉CARIM-School for Cardiovascular Diseases, Maastricht University, The Netherlands. s.foulquier@maastrichtuniversity.nl〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23364772" target="_blank"〉PubMed〈/a〉
    Keywords: Animals ; Cardiovascular Diseases/*metabolism/physiopathology/prevention & control ; Humans ; Protein Binding ; Receptor, Angiotensin, Type 1/metabolism ; Receptor, Angiotensin, Type 2/metabolism ; Renin-Angiotensin System/*physiology
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  • 44
    Publication Date: 2013-10-22
    Description: A large number of cis-regulatory sequences have been annotated in the human genome, but defining their target genes remains a challenge. One strategy is to identify the long-range looping interactions at these elements with the use of chromosome conformation capture (3C)-based techniques. However, previous studies lack either the resolution or coverage to permit a whole-genome, unbiased view of chromatin interactions. Here we report a comprehensive chromatin interaction map generated in human fibroblasts using a genome-wide 3C analysis method (Hi-C). We determined over one million long-range chromatin interactions at 5-10-kb resolution, and uncovered general principles of chromatin organization at different types of genomic features. We also characterized the dynamics of promoter-enhancer contacts after TNF-alpha signalling in these cells. Unexpectedly, we found that TNF-alpha-responsive enhancers are already in contact with their target promoters before signalling. Such pre-existing chromatin looping, which also exists in other cell types with different extracellular signalling, is a strong predictor of gene induction. Our observations suggest that the three-dimensional chromatin landscape, once established in a particular cell type, is relatively stable and could influence the selection or activation of target genes by a ubiquitous transcription activator in a cell-specific manner.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3838900/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3838900/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Jin, Fulai -- Li, Yan -- Dixon, Jesse R -- Selvaraj, Siddarth -- Ye, Zhen -- Lee, Ah Young -- Yen, Chia-An -- Schmitt, Anthony D -- Espinoza, Celso A -- Ren, Bing -- P50 GM085764/GM/NIGMS NIH HHS/ -- P50 GM085764-03/GM/NIGMS NIH HHS/ -- T32 GM008666/GM/NIGMS NIH HHS/ -- U01 ES017166/ES/NIEHS NIH HHS/ -- England -- Nature. 2013 Nov 14;503(7475):290-4. doi: 10.1038/nature12644. Epub 2013 Oct 20.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉1] Ludwig Institute for Cancer Research, 9500 Gilman Drive, La Jolla, California 92093, USA [2].〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/24141950" target="_blank"〉PubMed〈/a〉
    Keywords: Cell Line ; Chromatin/chemistry/genetics/*metabolism ; *Chromosome Mapping ; Enhancer Elements, Genetic/physiology ; Gene Expression Regulation ; *Genome, Human ; Humans ; Imaging, Three-Dimensional ; Promoter Regions, Genetic/physiology ; Protein Binding ; Signal Transduction ; Tumor Necrosis Factor-alpha/metabolism
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  • 45
    Publication Date: 2013-02-12
    Description: Molecular control of the pluripotent state is thought to reside in a core circuitry of master transcription factors including the homeodomain-containing protein NANOG, which has an essential role in establishing ground state pluripotency during somatic cell reprogramming. Whereas the genomic occupancy of NANOG has been extensively investigated, comparatively little is known about NANOG-associated proteins and their contribution to the NANOG-mediated reprogramming process. Using enhanced purification techniques and a stringent computational algorithm, we identify 27 high-confidence protein interaction partners of NANOG in mouse embryonic stem cells. These consist of 19 previously unknown partners of NANOG that have not been reported before, including the ten-eleven translocation (TET) family methylcytosine hydroxylase TET1. We confirm physical association of NANOG with TET1, and demonstrate that TET1, in synergy with NANOG, enhances the efficiency of reprogramming. We also find physical association and reprogramming synergy of TET2 with NANOG, and demonstrate that knockdown of TET2 abolishes the reprogramming synergy of NANOG with a catalytically deficient mutant of TET1. These results indicate that the physical interaction between NANOG and TET1/TET2 proteins facilitates reprogramming in a manner that is dependent on the catalytic activity of TET1/TET2. TET1 and NANOG co-occupy genomic loci of genes associated with both maintenance of pluripotency and lineage commitment in embryonic stem cells, and TET1 binding is reduced upon NANOG depletion. Co-expression of NANOG and TET1 increases 5-hydroxymethylcytosine levels at the top-ranked common target loci Esrrb and Oct4 (also called Pou5f1), resulting in priming of their expression before reprogramming to naive pluripotency. We propose that TET1 is recruited by NANOG to enhance the expression of a subset of key reprogramming target genes. These results provide an insight into the reprogramming mechanism of NANOG and uncover a new role for 5-methylcytosine hydroxylases in the establishment of naive pluripotency.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3606645/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3606645/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Costa, Yael -- Ding, Junjun -- Theunissen, Thorold W -- Faiola, Francesco -- Hore, Timothy A -- Shliaha, Pavel V -- Fidalgo, Miguel -- Saunders, Arven -- Lawrence, Moyra -- Dietmann, Sabine -- Das, Satyabrata -- Levasseur, Dana N -- Li, Zhe -- Xu, Mingjiang -- Reik, Wolf -- Silva, Jose C R -- Wang, Jianlong -- 079249/Wellcome Trust/United Kingdom -- 086692/Wellcome Trust/United Kingdom -- 095645/Wellcome Trust/United Kingdom -- 1R01-GM095942-01A1/GM/NIGMS NIH HHS/ -- BB/H008071/1/Biotechnology and Biological Sciences Research Council/United Kingdom -- G0700098/Medical Research Council/United Kingdom -- R01 GM095942/GM/NIGMS NIH HHS/ -- R01 HL112294/HL/NHLBI NIH HHS/ -- WT079249/Wellcome Trust/United Kingdom -- WT086692MA/Wellcome Trust/United Kingdom -- Biotechnology and Biological Sciences Research Council/United Kingdom -- Medical Research Council/United Kingdom -- England -- Nature. 2013 Mar 21;495(7441):370-4. doi: 10.1038/nature11925. Epub 2013 Feb 10.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Wellcome Trust-Medical Research Council Cambridge Stem Cell Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QR, UK.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23395962" target="_blank"〉PubMed〈/a〉
    Keywords: Animals ; Cellular Reprogramming/*physiology ; DNA-Binding Proteins/genetics/*metabolism ; Embryonic Stem Cells ; Gene Expression Regulation, Developmental ; Genome ; Homeodomain Proteins/genetics/*metabolism ; Mice ; Protein Binding ; Proto-Oncogene Proteins/genetics/*metabolism
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  • 46
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    Nature Publishing Group (NPG)
    Publication Date: 2013-03-23
    Description: 〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Patton, E Elizabeth -- Harrington, Lea -- G120/875/Medical Research Council/United Kingdom -- MC_PC_U127585840/Medical Research Council/United Kingdom -- MC_U127585840/Medical Research Council/United Kingdom -- England -- Nature. 2013 Mar 21;495(7441):320-1. doi: 10.1038/495320a.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23518559" target="_blank"〉PubMed〈/a〉
    Keywords: Gene Expression Regulation, Neoplastic ; Humans ; Mutation ; Neoplasms/enzymology/*genetics ; Protein Binding ; Telomerase/metabolism
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  • 47
    Publication Date: 2013-07-16
    Description: Folate receptors (FRalpha, FRbeta and FRgamma) are cysteine-rich cell-surface glycoproteins that bind folate with high affinity to mediate cellular uptake of folate. Although expressed at very low levels in most tissues, folate receptors, especially FRalpha, are expressed at high levels in numerous cancers to meet the folate demand of rapidly dividing cells under low folate conditions. The folate dependency of many tumours has been therapeutically and diagnostically exploited by administration of anti-FRalpha antibodies, high-affinity antifolates, folate-based imaging agents and folate-conjugated drugs and toxins. To understand how folate binds its receptors, we determined the crystal structure of human FRalpha in complex with folic acid at 2.8 A resolution. FRalpha has a globular structure stabilized by eight disulphide bonds and contains a deep open folate-binding pocket comprised of residues that are conserved in all receptor subtypes. The folate pteroate moiety is buried inside the receptor, whereas its glutamate moiety is solvent-exposed and sticks out of the pocket entrance, allowing it to be conjugated to drugs without adversely affecting FRalpha binding. The extensive interactions between the receptor and ligand readily explain the high folate-binding affinity of folate receptors and provide a template for designing more specific drugs targeting the folate receptor system.〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Chen, Chen -- Ke, Jiyuan -- Zhou, X Edward -- Yi, Wei -- Brunzelle, Joseph S -- Li, Jun -- Yong, Eu-Leong -- Xu, H Eric -- Melcher, Karsten -- R01 DK071662/DK/NIDDK NIH HHS/ -- R01 GM102545/GM/NIGMS NIH HHS/ -- England -- Nature. 2013 Aug 22;500(7463):486-9. doi: 10.1038/nature12327. Epub 2013 Jul 14.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Program for Structural Biology and Drug Discovery, Van Andel Research Institute, 333 Bostwick Avenue North East, Grand Rapids, Michigan 49503, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23851396" target="_blank"〉PubMed〈/a〉
    Keywords: Binding Sites/genetics ; Crystallography, X-Ray ; Folate Receptor 1/*chemistry/genetics/*metabolism ; Folic Acid/chemistry/*metabolism ; Humans ; Ligands ; Models, Molecular ; Mutation ; Protein Binding ; Structure-Activity Relationship
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  • 48
    Publication Date: 2013-11-08
    Description: Acyl carrier protein represents one of the most highly conserved proteins across all domains of life and is nature's way of transporting hydrocarbon chains in vivo. Notably, type II acyl carrier proteins serve as a crucial interaction hub in primary cellular metabolism by communicating transiently between partner enzymes of the numerous biosynthetic pathways. However, the highly transient nature of such interactions and the inherent conformational mobility of acyl carrier protein have stymied previous attempts to visualize structurally acyl carrier protein tied to an overall catalytic cycle. This is essential to understanding a fundamental aspect of cellular metabolism leading to compounds that are not only useful to the cell, but also of therapeutic value. For example, acyl carrier protein is central to the biosynthesis of the lipid A (endotoxin) component of lipopolysaccharides in Gram-negative microorganisms, which is required for their growth and survival, and is an activator of the mammalian host's immune system, thus emerging as an important therapeutic target. During lipid A synthesis (Raetz pathway), acyl carrier protein shuttles acyl intermediates linked to its prosthetic 4'-phosphopantetheine group among four acyltransferases, including LpxD. Here we report the crystal structures of three forms of Escherichia coli acyl carrier protein engaging LpxD, which represent stalled substrate and liberated products along the reaction coordinate. The structures show the intricate interactions at the interface that optimally position acyl carrier protein for acyl delivery and that directly involve the pantetheinyl group. Conformational differences among the stalled acyl carrier proteins provide the molecular basis for the association-dissociation process. An unanticipated conformational shift of 4'-phosphopantetheine groups within the LpxD catalytic chamber shows an unprecedented role of acyl carrier protein in product release.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3947097/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3947097/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Masoudi, Ali -- Raetz, Christian R H -- Zhou, Pei -- Pemble, Charles W 4th -- AI-055588/AI/NIAID NIH HHS/ -- GM-51310/GM/NIGMS NIH HHS/ -- P30 CA014236/CA/NCI NIH HHS/ -- R01 AI055588/AI/NIAID NIH HHS/ -- R01 GM051310/GM/NIGMS NIH HHS/ -- England -- Nature. 2014 Jan 16;505(7483):422-6. doi: 10.1038/nature12679. Epub 2013 Nov 6.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Department of Biochemistry, Duke University Medical Center, Durham, North Carolina 27710, USA. ; 1] Department of Biochemistry, Duke University Medical Center, Durham, North Carolina 27710, USA [2]. ; 1] Human Vaccine Institute, Duke University Medical Center, Durham, North Carolina 27710, USA [2] Duke Macromolecular Crystallography Center, Duke University Medical Center, Durham, North Carolina 27710, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/24196711" target="_blank"〉PubMed〈/a〉
    Keywords: Acyl Carrier Protein/*chemistry/*metabolism ; Acyltransferases/chemistry/metabolism ; *Biocatalysis ; Crystallography, X-Ray ; Escherichia coli/*chemistry ; Hydrolysis ; Lipid A/*biosynthesis/metabolism ; Models, Molecular ; Protein Binding ; Protein Conformation
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  • 49
    Publication Date: 2013-05-03
    Description: MicroRNAs (miRNAs) are generated by two-step processing to yield small RNAs that negatively regulate target gene expression at the post-transcriptional level. Deregulation of miRNAs has been linked to diverse pathological processes, including cancer. Recent studies have also implicated miRNAs in the regulation of cellular response to a spectrum of stresses, such as hypoxia, which is frequently encountered in the poorly angiogenic core of a solid tumour. However, the upstream regulators of miRNA biogenesis machineries remain obscure, raising the question of how tumour cells efficiently coordinate and impose specificity on miRNA expression and function in response to stresses. Here we show that epidermal growth factor receptor (EGFR), which is the product of a well-characterized oncogene in human cancers, suppresses the maturation of specific tumour-suppressor-like miRNAs in response to hypoxic stress through phosphorylation of argonaute 2 (AGO2) at Tyr 393. The association between EGFR and AGO2 is enhanced by hypoxia, leading to elevated AGO2-Y393 phosphorylation, which in turn reduces the binding of Dicer to AGO2 and inhibits miRNA processing from precursor miRNAs to mature miRNAs. We also identify a long-loop structure in precursor miRNAs as a critical regulatory element in phospho-Y393-AGO2-mediated miRNA maturation. Furthermore, AGO2-Y393 phosphorylation mediates EGFR-enhanced cell survival and invasiveness under hypoxia, and correlates with poorer overall survival in breast cancer patients. Our study reveals a previously unrecognized function of EGFR in miRNA maturation and demonstrates how EGFR is likely to function as a regulator of AGO2 through novel post-translational modification. These findings suggest that modulation of miRNA biogenesis is important for stress response in tumour cells and has potential clinical implications.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3717558/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3717558/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Shen, Jia -- Xia, Weiya -- Khotskaya, Yekaterina B -- Huo, Longfei -- Nakanishi, Kotaro -- Lim, Seung-Oe -- Du, Yi -- Wang, Yan -- Chang, Wei-Chao -- Chen, Chung-Hsuan -- Hsu, Jennifer L -- Wu, Yun -- Lam, Yung Carmen -- James, Brian P -- Liu, Xiuping -- Liu, Chang-Gong -- Patel, Dinshaw J -- Hung, Mien-Chie -- CA099031/CA/NCI NIH HHS/ -- CA109311/CA/NCI NIH HHS/ -- CA16672/CA/NCI NIH HHS/ -- P01 CA099031/CA/NCI NIH HHS/ -- P30 CA016672/CA/NCI NIH HHS/ -- R01 CA109311/CA/NCI NIH HHS/ -- England -- Nature. 2013 May 16;497(7449):383-7. doi: 10.1038/nature12080. Epub 2013 May 1.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Department of Molecular and Cellular Oncology, The University of Texas MD Anderson Cancer Center, 1515 Holcombe Boulevard, Houston, Texas 77030, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23636329" target="_blank"〉PubMed〈/a〉
    Keywords: Argonaute Proteins/*chemistry/*metabolism ; Breast Neoplasms/genetics/metabolism/mortality/pathology ; Cell Hypoxia/genetics/*physiology ; Cell Line, Tumor ; Cell Survival ; Female ; Gene Expression Regulation, Neoplastic ; Humans ; MicroRNAs/biosynthesis/chemistry/genetics/*metabolism ; Neoplasm Invasiveness ; Nucleic Acid Conformation ; Phosphorylation ; Phosphotyrosine/metabolism ; Prognosis ; Protein Binding ; RNA Precursors/chemistry/genetics/metabolism ; Receptor, Epidermal Growth Factor/*metabolism ; Ribonuclease III/metabolism ; Survival Analysis
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  • 50
    Publication Date: 2013-11-29
    Description: N(6)-methyladenosine (m(6)A) is the most prevalent internal (non-cap) modification present in the messenger RNA of all higher eukaryotes. Although essential to cell viability and development, the exact role of m(6)A modification remains to be determined. The recent discovery of two m(6)A demethylases in mammalian cells highlighted the importance of m(6)A in basic biological functions and disease. Here we show that m(6)A is selectively recognized by the human YTH domain family 2 (YTHDF2) 'reader' protein to regulate mRNA degradation. We identified over 3,000 cellular RNA targets of YTHDF2, most of which are mRNAs, but which also include non-coding RNAs, with a conserved core motif of G(m(6)A)C. We further establish the role of YTHDF2 in RNA metabolism, showing that binding of YTHDF2 results in the localization of bound mRNA from the translatable pool to mRNA decay sites, such as processing bodies. The carboxy-terminal domain of YTHDF2 selectively binds to m(6)A-containing mRNA, whereas the amino-terminal domain is responsible for the localization of the YTHDF2-mRNA complex to cellular RNA decay sites. Our results indicate that the dynamic m(6)A modification is recognized by selectively binding proteins to affect the translation status and lifetime of mRNA.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3877715/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3877715/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Wang, Xiao -- Lu, Zhike -- Gomez, Adrian -- Hon, Gary C -- Yue, Yanan -- Han, Dali -- Fu, Ye -- Parisien, Marc -- Dai, Qing -- Jia, Guifang -- Ren, Bing -- Pan, Tao -- He, Chuan -- GM071440/GM/NIGMS NIH HHS/ -- GM088599/GM/NIGMS NIH HHS/ -- K01 HG006699/HG/NHGRI NIH HHS/ -- R01 GM071440/GM/NIGMS NIH HHS/ -- R01 GM088599/GM/NIGMS NIH HHS/ -- England -- Nature. 2014 Jan 2;505(7481):117-20. doi: 10.1038/nature12730. Epub 2013 Nov 27.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Department of Chemistry and Institute for Biophysical Dynamics, The University of Chicago, 929 East 57th Street, Chicago, Illinois 60637, USA. ; Ludwig Institute for Cancer Research, Department of Cellular and Molecular Medicine, UCSD Moores Cancer Center and Institute of Genome Medicine, University of California, San Diego School of Medicine, 9500 Gilman Drive, La Jolla, California 92093-0653, USA. ; Department of Biochemistry and Molecular Biology and Institute for Biophysical Dynamics, The University of Chicago, 929 East 57th Street, Chicago, Illinois 60637, USA. ; 1] Department of Chemistry and Institute for Biophysical Dynamics, The University of Chicago, 929 East 57th Street, Chicago, Illinois 60637, USA [2] Department of Chemical Biology and Synthetic and Functional Biomolecules Center, College of Chemistry and Molecular Engineering, Peking University, Beijing 100871, China.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/24284625" target="_blank"〉PubMed〈/a〉
    Keywords: Adenosine/*analogs & derivatives/metabolism/pharmacology ; Base Sequence ; DNA-Binding Proteins/genetics ; HeLa Cells ; Humans ; Nucleotide Motifs ; Organelles/genetics/metabolism ; Protein Binding ; Protein Biosynthesis ; *RNA Stability/drug effects ; RNA Transport ; RNA, Messenger/*chemistry/*metabolism ; RNA, Untranslated/chemistry/metabolism ; RNA-Binding Proteins/chemistry/classification/*metabolism ; Substrate Specificity
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  • 51
    Publication Date: 2013-11-19
    Description: Splicing of precursor messenger RNA (pre-mRNA) in eukaryotic cells is carried out by the spliceosome, which consists of five small nuclear ribonucleoproteins (snRNPs) and a number of accessory factors and enzymes. Each snRNP contains a ring-shaped subcomplex of seven proteins and a specific RNA molecule. The U6 snRNP contains a unique heptameric Lsm protein complex, which specifically recognizes the U6 small nuclear RNA at its 3' end. Here we report the crystal structures of the heptameric Lsm complex, both by itself and in complex with a 3' fragment of U6 snRNA, at 2.8 A resolution. Each of the seven Lsm proteins interacts with two neighbouring Lsm components to form a doughnut-shaped assembly, with the order Lsm3-2-8-4-7-5-6. The four uridine nucleotides at the 3' end of U6 snRNA are modularly recognized by Lsm3, Lsm2, Lsm8 and Lsm4, with the uracil base specificity conferred by a highly conserved asparagine residue. The uracil base at the extreme 3' end is sandwiched by His 36 and Arg 69 from Lsm3, through pi-pi and cation-pi interactions, respectively. The distinctive end-recognition of U6 snRNA by the Lsm complex contrasts with RNA binding by the Sm complex in the other snRNPs. The structural features and associated biochemical analyses deepen mechanistic understanding of the U6 snRNP function in pre-mRNA splicing.〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Zhou, Lijun -- Hang, Jing -- Zhou, Yulin -- Wan, Ruixue -- Lu, Guifeng -- Yin, Ping -- Yan, Chuangye -- Shi, Yigong -- England -- Nature. 2014 Feb 6;506(7486):116-20. doi: 10.1038/nature12803. Epub 2013 Nov 17.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉1] Ministry of Education Key Laboratory of Protein Science, Tsinghua University, Beijing 100084, China [2] Tsinghua-Peking Joint Center for Life Sciences, Center for Structural Biology, School of Life Sciences and School of Medicine, Tsinghua University, Beijing 100084, China [3]. ; 1] Tsinghua-Peking Joint Center for Life Sciences, Center for Structural Biology, School of Life Sciences and School of Medicine, Tsinghua University, Beijing 100084, China [2] State Key Laboratory of Bio-membrane and Membrane Biotechnology, Tsinghua University, Beijing 100084, China [3]. ; Ministry of Education Key Laboratory of Protein Science, Tsinghua University, Beijing 100084, China. ; State Key Laboratory of Bio-membrane and Membrane Biotechnology, Tsinghua University, Beijing 100084, China. ; Tsinghua-Peking Joint Center for Life Sciences, Center for Structural Biology, School of Life Sciences and School of Medicine, Tsinghua University, Beijing 100084, China. ; 1] Tsinghua-Peking Joint Center for Life Sciences, Center for Structural Biology, School of Life Sciences and School of Medicine, Tsinghua University, Beijing 100084, China [2] State Key Laboratory of Bio-membrane and Membrane Biotechnology, Tsinghua University, Beijing 100084, China. ; 1] Ministry of Education Key Laboratory of Protein Science, Tsinghua University, Beijing 100084, China [2] Tsinghua-Peking Joint Center for Life Sciences, Center for Structural Biology, School of Life Sciences and School of Medicine, Tsinghua University, Beijing 100084, China.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/24240276" target="_blank"〉PubMed〈/a〉
    Keywords: Amino Acid Sequence ; Asparagine/chemistry ; Base Sequence ; Crystallography, X-Ray ; Histidine/chemistry ; Models, Molecular ; Molecular Sequence Data ; Multiprotein Complexes/*chemistry/metabolism ; Protein Binding ; Protein Structure, Quaternary ; RNA, Small Nuclear/*chemistry/*genetics/metabolism ; RNA-Binding Proteins/*chemistry/metabolism ; Ribonucleoproteins, Small Nuclear/chemistry/metabolism ; Saccharomyces cerevisiae/chemistry ; Saccharomyces cerevisiae Proteins/*chemistry/metabolism ; Uracil/chemistry/metabolism
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  • 52
    Publication Date: 2013-10-25
    Description: Many secretory proteins are targeted by signal sequences to a protein-conducting channel, formed by prokaryotic SecY or eukaryotic Sec61 complexes, and are translocated across the membrane during their synthesis. Crystal structures of the inactive channel show that the SecY subunit of the heterotrimeric complex consists of two halves that form an hourglass-shaped pore with a constriction in the middle of the membrane and a lateral gate that faces the lipid phase. The closed channel has an empty cytoplasmic funnel and an extracellular funnel that is filled with a small helical domain, called the plug. During initiation of translocation, a ribosome-nascent chain complex binds to the SecY (or Sec61) complex, resulting in insertion of the nascent chain. However, the mechanism of channel opening during translocation is unclear. Here we have addressed this question by determining structures of inactive and active ribosome-channel complexes with cryo-electron microscopy. Non-translating ribosome-SecY channel complexes derived from Methanocaldococcus jannaschii or Escherichia coli show the channel in its closed state, and indicate that ribosome binding per se causes only minor changes. The structure of an active E. coli ribosome-channel complex demonstrates that the nascent chain opens the channel, causing mostly rigid body movements of the amino- and carboxy-terminal halves of SecY. In this early translocation intermediate, the polypeptide inserts as a loop into the SecY channel with the hydrophobic signal sequence intercalated into the open lateral gate. The nascent chain also forms a loop on the cytoplasmic surface of SecY rather than entering the channel directly.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3948209/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3948209/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Park, Eunyong -- Menetret, Jean-Francois -- Gumbart, James C -- Ludtke, Steven J -- Li, Weikai -- Whynot, Andrew -- Rapoport, Tom A -- Akey, Christopher W -- GM052586/GM/NIGMS NIH HHS/ -- GM067887/GM/NIGMS NIH HHS/ -- GM080139/GM/NIGMS NIH HHS/ -- GM45377/GM/NIGMS NIH HHS/ -- K99 HL097083/HL/NHLBI NIH HHS/ -- R00 HL097083/HL/NHLBI NIH HHS/ -- R01 GM045377/GM/NIGMS NIH HHS/ -- R01 GM052586/GM/NIGMS NIH HHS/ -- R01 GM067887/GM/NIGMS NIH HHS/ -- R01 GM080139/GM/NIGMS NIH HHS/ -- R01 HL121718/HL/NHLBI NIH HHS/ -- Howard Hughes Medical Institute/ -- England -- Nature. 2014 Feb 6;506(7486):102-6. doi: 10.1038/nature12720. Epub 2013 Oct 23.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Department of Cell Biology and Howard Hughes Medical Institute, Harvard Medical School, 240 Longwood Avenue, Boston, Massachusetts 02115, USA. ; Department of Physiology and Biophysics, Boston University School of Medicine, 700 Albany Street, Boston, Massachusetts 02118-2526, USA. ; School of Physics, Georgia Institute of Technology, Atlanta, Georgia 30332, USA. ; National Center for Macromolecular Imaging, Verna and Marrs McLean Department of Biochemistry and Molecular Biology, Baylor College of Medicine, 1 Baylor Plaza, Houston, Texas 77030, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/24153188" target="_blank"〉PubMed〈/a〉
    Keywords: Cryoelectron Microscopy ; Escherichia coli/*chemistry/ultrastructure ; Escherichia coli Proteins/chemistry/isolation & ; purification/*metabolism/*ultrastructure ; Methanocaldococcus/*chemistry/ultrastructure ; Models, Molecular ; Multiprotein Complexes/chemistry/isolation & ; purification/metabolism/ultrastructure ; Peptides/chemistry/metabolism ; Protein Binding ; *Protein Biosynthesis ; Protein Subunits/chemistry/isolation & purification/metabolism ; Protein Transport ; Ribosomes/chemistry/*metabolism/*ultrasonography
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