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  • 1
    Electronic Resource
    Electronic Resource
    Magnitude and significance of NAD turnover in human cell line D98/AH2 (1976)
    [s.l.] : Nature Publishing Group
    Nature 259 (1976), S. 695-696 
    Details
    ISSN: 1476-4687
    Source: Nature Archives 1869 - 2009
    Topics: Biology , Chemistry and Pharmacology , Medicine , Natural Sciences in General , Physics
    Notes: [Auszug] What is the magnitude in vivo of the reactions which consume NAD? We present here data for D98/AH2, a human cell line derived from HeLa9 which answers this question and which makes possible calculation of the fraction of NAD biosynthesis that compensates for breakdown and the fraction that expands ...
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1038/259695a0
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  • 2
    Electronic Resource
    Electronic Resource
    Replication of Escherichia coli requires DNA polymerase I (1974)
    [s.l.] : Nature Publishing Group
    Nature 250 (1974), S. 513-514 
    Details
    ISSN: 1476-4687
    Source: Nature Archives 1869 - 2009
    Topics: Biology , Chemistry and Pharmacology , Medicine , Natural Sciences in General , Physics
    Notes: [Auszug] Among many DNA polymerase I mutants of E. coli we have isolated one (BT 4113) which is conditional lethal at elevated temperature because of a temperature-sensitive DNA polymerase I. A bacterial culture was mutagenised by N-methyl-nitrosoguanidine and screened for pol A mutants using a new replica ...
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1038/250513a0
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  • 3
    Electronic Resource
    Electronic Resource
    Pyridine nucleotide metabolism in imaginal discs of Drosophila melanogaster (1976)
    Springer
    Biochemical genetics 14 (1976), S. 197-207 
    Details
    ISSN: 1573-4927
    Keywords: pyridine nucleotide metabolism ; imaginal discs ; NAD turnover ; ecdysterone
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract The pyridine nucleotide metabolism of imaginal discs of Drosophila melanogaster has been studied in vitro by incubating discs with labeled nicotinic acid in the presence and absence of ecdysterone. The major labeled compounds found within the discs are NAD, NADP, and nicotinic acid. There is preferential uptake of nicotinamide over nicotinic acid, although the Priess-Handler pathway is used exclusively. The presence of ecdysterone produces a small increase in the NADP/NAD ratio, and an increase in NAD synthesis, probably to compensate for increased NAD turnover.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00484760
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  • 4
    Electronic Resource
    Electronic Resource
    Electrophoresis of the nucleic acidsContribution No. 3108.A preliminary report of this work has appeared elsewhere. (1964)
    New York : Wiley-Blackwell
    Biopolymers 2 (1964), S. 245-257 
    Details
    ISSN: 0006-3525
    Keywords: Chemistry ; Polymer and Materials Science
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology
    Notes: A zone electrophoresis apparatus using ultraviolet, optics has been constructed to study nucleic acids at concentration less than 0.004%. Native DNA has a mobility about 15% higher than denatured DNA over a range of conditions of pH and ionic strength. DNA's from different sources have closely similar mobilities. A study of a molecular weight series of DNA indicates that the mobility is constant in the molecular weight range of 2.5 × 105 to 1.3 × 108. DNA mobilities change in the expected way with pH but the fractional change in mobility is less than the change in charge calculated by titration curves. A small decrease in mobility accompanies an increase in ionic strength. RNA's from various sources have mobilities slightly lower than denatured DNA except for s-RNA which travels slightly faster.
    Additional Material: 4 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/bip.1964.360020306
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  • 5
    Electronic Resource
    Electronic Resource
    Pyridine nucleotide metabolism in mammalian cells in culture (1973)
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 82 (1973), S. 165-179 
    Details
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The biosynthesis of pyridine nucleotides has been examined in a number of mammalian cell lines in culture. In all lines examined, nicotinamide is incorporated by a biochemical pathway distinct from the Preiss-Handler pathway for nicotinic acid.In at least the human cell line D98/AH2, there is no detectable endogenous synthesis of the pyridine ring from tryptophan. Although most cell lines examined (hamster BHK 21/13, mouse L929 and human D98/AH2) use either nicotinic acid or nicotinamide as a precursor for DPN and TPN, two mouse cell lines, 3T3-4E and LM CIID, are unable to utilize nicotinic acid as a source of the pyridine ring.If nicotinic acid is present in the medium, substantial amounts of intracellular desamido DPN accumulate suggesting that the last step (desamido DPN→DPN) is limiting in the Preiss-Handler pathway. With nicotinamide, the only compound which accumulates in substantial amounts apart from DPN and TPN is nicotinamide ribose; there is no detectable NMN. The results of pulse-labeling experiments suggest that nicotinamide ribose may be an intermediate in the nicotinamide pathway.Following growth of D98/AH2 cells in high concentrations of niacin, biosynthesis of DPN from nicotinamide was completely inhibited for at least six hours. The converse experiment revealed no inhibition of niacin incorporation. This observation suggests that a niacin pathway intermediate, which present evidence indicates is desamido-DPN. can inhibit nicotinamide utilization.Newly synthesized DPN turns over with a half-life of two hours in azaserine-treated D98/AH2 cells. In the absence of azaserine, the nicotinamide moiety of newly synthesized DPN is lost from D98/AH2 cells to the medium with a half-life of eight hours. About 80% of the nicotinamide is lost to medium as nicotinamide ribose.
    Additional Material: 9 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/jcp.1040820205
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  • 6
    Electronic Resource
    Electronic Resource
    Turnover of nicotinamide adenine dinucleotide in cultures of human cells (1976)
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 88 (1976), S. 207-217 
    Details
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The rate of turnover of nicotinamide adenine dinucleotide (NAD) in the human cell line, D98/AH2, has been estimated by measuring the rates of entry into and exit from NAD molecules of 14C-adenine. In one set of experiments, cells were labeled by growth in medium containing 14C-adenine for six hours and then shifted to medium without labeled adenine. The loss of 14C-adenine from the adenine nucleotide and pyridine nucleotide pools was measured, and the data were analyzed using an analytical treatment which corrects for the relatively slow turnover of precursor pools. The loss of 14C-adenine from the NAD pool and from the precursor ATP pool could be related to the absolute rate of NAD breakdown. Under the experimental conditions used, the rate of NAD turnover ranged from 83,000 to 126,000 molecules per second per cell. In a complementary experiment cells were grown in the presence of unlabeled adenine, then shifted into medium containing 14C-adenine and the rate of entry of 14C-adenine into adenine and pyridine nucleotides was measured. The data were treated using a similar analysis to relate the rate of entry of 14C-adenine into NAD and the precursor ATP pools to the absolute turnover rate of NAD. This analysis gave a value for NAD turnover of 78,000 molecules per second per cell in excellent agreement with results from the pulse-chase experiments. The results from both types of experiment indicate that within D98/AH2 cells the half-life of an intact NAD molecule is 60 ± 18 minutes. Thus, in a human D98/AH2 cell growing with a generation time of 24 hours, NAD is turning over at twice the rate found in Escherichia coli with a generation time of half an hour.
    Additional Material: 5 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/jcp.1040880210
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  • 7
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    Metabolic model for diversity-generating biosynthesis (2016)
    National Academy of Sciences
    Details
    Publication Date: 2016-02-01
    Description: A conventional metabolic pathway leads to a specific product. In stark contrast, there are diversity-generating metabolic pathways that naturally produce different chemicals, sometimes of great diversity. We demonstrate that for one such pathway,tru, each ensuing metabolic step is slower, in parallel with the increasing potential chemical divergence generated as the pathway proceeds. Intermediates are long lived and accumulate progressively, in contrast with conventional metabolic pathways, in which the first step is rate-limiting and metabolic intermediates are short-lived. Understanding these fundamental differences enables several different practical applications, such as combinatorial biosynthesis, some of which we demonstrate here. We propose that these principles may provide a unifying framework underlying diversity-generating metabolism in many different biosynthetic pathways.
    Print ISSN: 0027-8424
    Electronic ISSN: 1091-6490
    Topics: Biology , Medicine , Natural Sciences in General
    Published by National Academy of Sciences
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  • 8
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    Specialized insulin is used for chemical warfare by fish-hunting cone snails (2015)
    National Academy of Sciences
    Details
    Publication Date: 2015-01-20
    Description: More than 100 species of venomous cone snails (genus Conus) are highly effective predators of fish. The vast majority of venom components identified and functionally characterized to date are neurotoxins specifically targeted to receptors, ion channels, and transporters in the nervous system of prey, predators, or competitors. Here we describe a venom component targeting energy metabolism, a radically different mechanism. Two fish-hunting cone snails, Conus geographus and Conus tulipa, have evolved specialized insulins that are expressed as major components of their venoms. These insulins are distinctive in having much greater similarity to fish insulins than to the molluscan hormone and are unique in that posttranslational modifications characteristic of conotoxins (hydroxyproline, γ-carboxyglutamate) are present. When injected into fish, the venom insulin elicits hypoglycemic shock, a condition characterized by dangerously low blood glucose. Our evidence suggests that insulin is specifically used as a weapon for prey capture by a subset of fish-hunting cone snails that use a net strategy to capture prey. Insulin appears to be a component of the nirvana cabal, a toxin combination in these venoms that is released into the water to disorient schools of small fish, making them easier to engulf with the snail’s distended false mouth, which functions as a net. If an entire school of fish simultaneously experiences hypoglycemic shock, this should directly facilitate capture by the predatory snail.
    Print ISSN: 0027-8424
    Electronic ISSN: 1091-6490
    Topics: Biology , Medicine , Natural Sciences in General
    Published by National Academy of Sciences
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  • 9
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    Insights into the origins of fish hunting in venomous cone snails from studies of Conus tessulatus (2015)
    National Academy of Sciences
    Details
    Publication Date: 2015-04-06
    Description: Prey shifts in carnivorous predators are events that can initiate the accelerated generation of new biodiversity. However, it is seldom possible to reconstruct how the change in prey preference occurred. Here we describe an evolutionary “smoking gun” that illuminates the transition from worm hunting to fish hunting among marine cone snails, resulting in the adaptive radiation of fish-hunting lineages comprising ∼100 piscivorous Conus species. This smoking gun is δ-conotoxin TsVIA, a peptide from the venom of Conus tessulatus that delays inactivation of vertebrate voltage-gated sodium channels. C. tessulatus is a species in a worm-hunting clade, which is phylogenetically closely related to the fish-hunting cone snail specialists. The discovery of a δ-conotoxin that potently acts on vertebrate sodium channels in the venom of a worm-hunting cone snail suggests that a closely related ancestral toxin enabled the transition from worm hunting to fish hunting, as δ-conotoxins are highly conserved among fish hunters and critical to their mechanism of prey capture; this peptide, δ-conotoxin TsVIA, has striking sequence similarity to these δ-conotoxins from piscivorous cone snail venoms. Calcium-imaging studies on dissociated dorsal root ganglion (DRG) neurons revealed the peptide’s putative molecular target (voltage-gated sodium channels) and mechanism of action (inhibition of channel inactivation). The results were confirmed by electrophysiology. This work demonstrates how elucidating the specific interactions between toxins and receptors from phylogenetically well-defined lineages can uncover molecular mechanisms that underlie significant evolutionary transitions.
    Print ISSN: 0027-8424
    Electronic ISSN: 1091-6490
    Topics: Biology , Medicine , Natural Sciences in General
    Published by National Academy of Sciences
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  • 10
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    Conotoxin κM-RIIIJ, a tool targeting asymmetric heteromeric Kv1 channels (2018)
    National Academy of Sciences
    Details
    Publication Date: 2018-12-28
    Description: The vast complexity of native heteromeric K+ channels is largely unexplored. Defining the composition and subunit arrangement of individual subunits in native heteromeric K+ channels and establishing their physiological roles is experimentally challenging. Here we systematically explored this “zone of ignorance” in molecular neuroscience. Venom components, such as peptide toxins, appear to have evolved to modulate physiologically relevant targets by discriminating among closely related native ion channel complexes. We provide proof-of-principle for this assertion by demonstrating that κM-conotoxin RIIIJ (κM-RIIIJ) from Conus radiatus precisely targets “asymmetric” Kv channels composed of three Kv1.2 subunits and one Kv1.1 or Kv1.6 subunit with 100-fold higher apparent affinity compared with homomeric Kv1.2 channels. Our study shows that dorsal root ganglion (DRG) neurons contain at least two major functional Kv1.2 channel complexes: a heteromer, for which κM-RIIIJ has high affinity, and a putative Kv1.2 homomer, toward which κM-RIIIJ is less potent. This conclusion was reached by (i) covalent linkage of members of the mammalian Shaker-related Kv1 family to Kv1.2 and systematic assessment of the potency of κM-RIIIJ block of heteromeric K+ channel-mediated currents in heterologous expression systems; (ii) molecular dynamics simulations of asymmetric Kv1 channels providing insights into the molecular basis of κM-RIIIJ selectivity and potency toward its targets; and (iii) evaluation of calcium responses of a defined population of DRG neurons to κM-RIIIJ. Our study demonstrates that bioactive molecules present in venoms provide essential pharmacological tools that systematically target specific heteromeric Kv channel complexes that operate in native tissues.
    Print ISSN: 0027-8424
    Electronic ISSN: 1091-6490
    Topics: Biology , Medicine , Natural Sciences in General
    Published by National Academy of Sciences
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