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  • Column liquid chromatography  (157)
  • Saccharomyces cerevisiae
  • Springer  (238)
  • Institute of Physics
  • 1995-1999  (64)
  • 1990-1994  (174)
  • 1940-1944
  • 1999  (64)
  • 1993  (174)
Collection
Keywords
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  • 1995-1999  (64)
  • 1990-1994  (174)
  • 1940-1944
Year
  • 1
    Electronic Resource
    Electronic Resource
    Identification of a fungal triacetylfusarinine C siderophore transport gene (TAF1) in Saccharomyces cerevisiae as a member of the major facilitator superfamily (1999)
    Springer
    BioMetals 12 (1999), S. 301-306 
    Details
    ISSN: 1572-8773
    Keywords: iron ; siderophores ; transport ; Saccharomyces cerevisiae ; fungi
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract Transport proteins of microorganisms may either belong to the ATP-binding cassette (ABC) superfamily or to the major facilitator (MFS)-superfamily. MFS transporters are single-polypeptide membrane transporters that transport small molecules via uniport, symport or antiport mechanisms in response to a chemiosmotic gradient. Although Saccharomyces cerevisiae is a non-siderophore producer, various bacterial and fungal siderophores can be utilized as an iron source. From yeast genome sequencing data six genes of the unknown major facilitator (UMF) family were known of which YEL065w Sce was recently identified as a transporter for the bacterial siderophore ferrioxamine B (Sit1p). The present investigation shows that another UMF gene, YHL047c Sce, encodes a transporter for the fungal siderophore triacetylfusarinine C. The gene YHL047c Sce (designated TAF1) was disrupted using the kanMX disruption module in a fet3 background (strain DEY 1394 Δfet3), possessing a defect in the high affinity ferrous iron transport. Growth promotion assays and transport experiments with 55Fe-labelled triacetylfusarinine C showed a complete loss of iron utilization and uptake in the disrupted strain, indicating that TAF1 is the gene for the fungal triacetylfusarinine transport in Saccharomyces cerevisiae and possibly in other siderophore producing fungi.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1009252118050
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  • 2
    Electronic Resource
    Electronic Resource
    Interaction of Saccharomyces cerevisiae with gold: toxicity and accumulation (1999)
    Springer
    BioMetals 12 (1999), S. 289-294 
    Details
    ISSN: 1572-8773
    Keywords: accumulation ; gold ; proton efflux ; Saccharomyces cerevisiae ; toxicity
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract This paper examines the effects of ionic gold on Saccharomyces cerevisiae, as determined by long-term (growth in gold-containing media) and short-term interactions (H+ efflux activity). An increasing gold concentration inhibited growth and at 〈0.2 mM Au, growth was not observed. Transmission electron microscopy revealed no differences in ultrastructure but fine electron dense particles were observed in unstained preparations from gold-containing medium. After glucose addition (to 10mM) to starved suspensions of S. cerevisiae, glucose-dependent reduction of external pH occurred as the cells extruded protons. In the presence of increasing gold concentrations, the lag time before proton extrusion did not change but the rate and duration decreased significantly with a marked influence on proton efflux rate being observed at ≤ 10 μM. Extension of preincubation time of yeast cells in gold-containing medium resulted in a decreasing proton efflux rate and colloidal phase formation in the cell suspensions, the time between gold addition and the beginning of colloidal phase formation depending on the gold concentration used. Both Ca and Mg enhanced the inhibitory effect of gold on the yeast cells with Ca showing a stronger inhibitory effect than Mg.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1009210101628
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  • 3
    Electronic Resource
    Electronic Resource
    Cloning and expression of Hormoconis resinae glucoamylase P cDNA in Saccharomyces cerevisiae (1993)
    Springer
    Current genetics 24 (1993), S. 38-44 
    Details
    ISSN: 1432-0983
    Keywords: Glucoamylase ; Gene cloning ; Hormoconis resinae ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A cDNA coding for glucoamylase P of Hormoconis resinae was cloned using a synthetic oligonucleotide probe coding for a peptide fragment of the purified enzyme and polyclonal anti-glucoamylase antibodies. Nucleotide-sequence analysis revealed an open reading frame of 1848 base pairs coding for a protein of 616 amino-acid residues. Comparison with other fungal glucoamylase amino-acid sequences showed homologies of 37–48%. The glucoamylase cDNA, when introduced into Saccharomyces cerevisiae under the control of the yeast ADC1 promoter, directed the secretion of active glucoamylase P into the growth medium.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00324663
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  • 4
    Electronic Resource
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    Normal mitochondrial structure and genome maintenance in yeast requires the dynamin-like product of the MGM1 gene (1993)
    Springer
    Current genetics 24 (1993), S. 141-148 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Dynamin ; Mitochondria ; GTP binding protein
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The isolation and characterization of MGM1, and yeast gene with homology to members of the dynamin gene family, is described. The MGM1 gene is located on the right arm of chromosome XV between STE4 and PTP2. Sequence analysis revealed a single open reading frame of 902 residues capable of encoding a protein with an approximate molecular mass of 101 kDa. Loss of MGM1 resulted in slow growth on rich medium, failure to grow on non-fermentable carbon sources, and loss of mitochondrial DNA. The mitochondria also appeared abnormal when visualized with an antibody to a mitochondrial-matrix marker. MGM1 encodes a dynamin-like protein involved in the propagation of functional mitochondria in yeast.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00324678
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  • 5
    Electronic Resource
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    Molecular cloning of the PEL1 gene of Saccharomyces cerevisiae that is essential for the viability of petite mutants (1993)
    Springer
    Current genetics 24 (1993), S. 307-312 
    Details
    ISSN: 1432-0983
    Keywords: Growth control ; Genetic mapping ; Molecular cloning ; Nucleo-mitochondrial interaction ; Saccharomyces cerevisiae ; Viability of petites
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The PEL1 gene of Saccharomyces cerevisiae is essential for the cell viability of mitochondrial petite mutants, for the ability to utilize glycerol and ethanol on synthetic medium, and for cell growth at higher temperatures. By tetrad analysis the gene was assigned to chromosome III, centromere proximal of LEU2. The PEL1 gene has been isolated and cloned by the complementation of a pel1 mutation. The molecular analysis of the chromosomal insert carrying PEL1 revealed that this gene corresponds to the YCL4W open reading frame on the complete DNA sequence of chromosome III. The putative Pel1 protein is characterized by a low molecular weight of approximately 17 kDa, a low codon adaptation index, and a high leucine content.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00336781
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  • 6
    Electronic Resource
    Electronic Resource
    Sequence analysis of a Papaver somniferum L. mitochondrial DNA fragment promoting autonomous plasmid replication in Saccharomyces cerevisiae and Kluyveromyces lactis (1993)
    Springer
    Current genetics 24 (1993), S. 366-367 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Papaver somniferum L. ; ARS ; Mitochondrial DNA
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The minimal fragment of mitochondrial DNA from Papaver somniferum L. (poppy) able to promote autonomous plasmid replication in the yeast Saccharomyces cerevisiae was sequenced. Sequence analysis of the 917-bp MK4/8 DNA fragment revealed a high AT content, and the presence of two 12-bp sequences differing from the ARS core consensus of S. cerevisiae only by a T and C insertion, respectively. The mitochondrial insert contains a further six 11-bp sequences with one mismatch to the S. cerevisiae core consensus, more then 20 related sequences with two base pair exchanges, numerous direct and inverted repeats, and many copies of a sequence motif called the ARS box. The original 4.2-kb mitochondrial DNA fragment, as well as the minimal 917-bp subfragment in vector pFL1-E (a variant of YIP5, lacking an origin of replication in yeast), were then tested for their ability to replicate autonomously in another fungus, Kluyveromyces lactis.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00336790
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  • 7
    Electronic Resource
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    The ogd1 and kgd1 mutants lacking 2-oxoglutarate dehydrogenase activity in yeast are allelic and can be differentiated by the cloned amber suppressor (1993)
    Springer
    Current genetics 24 (1993), S. 377-381 
    Details
    ISSN: 1432-0983
    Keywords: 2-Oxoglutarate dehydrogenase ; Molecular cloning ; Saccharomyces cerevisiae ; Sequencing ; Suppressor ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The activity of mitochondrial 2-oxoglutarate dehydrogenase in S. cerevisiae can be impaired either by the ogd1 or the kgd1 mutation. The OGD1 gene and two suppressor genes were isolated by complementation of the ogd1 mutant. The complementation of the kdg1 mutant by the OGD1 gene, an allelism test, and meiotic mapping, revealed that the ogd1 and kgd1 mutations are allelic. The two mutations were differentiated by the cloned suppressor gene which was able to partially complement ogd1, but not kgd1. The molecular analysis of the suppressor gene revealed its identity with the natural tRNA CAG Gln gene found in the upstream region of URA10.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00351844
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  • 8
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    Use of reporter genes for the isolation and characterisation of different classes of sporulation mutants in the yeast Saccharomyces cerevisiae (1993)
    Springer
    Current genetics 24 (1993), S. 451-454 
    Details
    ISSN: 1432-0983
    Keywords: Yeast ; Saccharomyces cerevisiae ; Sporulation mutants ; Reporter genes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Reporter genes consisting of sporulation-specific promoters fused to lacZ were used as markers to monitor the sporulation pathway of the yeast Saccharomyces cerevisiae. Strains transformed with these lacZ gene fusions expressed β-galactosidase (assayable on plates using the substrate 5-bromo-4-chloro-3-indolyl-β-D-galactopyranoside, X-gal) in a sporulation-dependent manner. Mutagenesis experiments performed on transformed strains resulted in the recovery of a number of novel sporulation mutants. Three classes of mutants were obtained: those which overexpressed the reporter gene under sporulation conditions, those which did not express the gene under any conditions, and those which expressed the gene in vegetative cells not undergoing sporulation. On the basis of the blue colony-colour produced in the presence of X-gal these have been described as superblue, white, and blue vegetative mutants, respectively. These were further characterised using earlier reporter genes and other marker systems. This study established that the multicopy reporter plasmids chosen do not interfere with sporulation; they are valid tools for monitoring the pathway and they provide a way to isolate mutations not readily selected by other markers.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00351856
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  • 9
    Electronic Resource
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    Physical mapping of the MEL gene family in Saccharomyces cerevisiae (1993)
    Springer
    Current genetics 24 (1993), S. 461-464 
    Details
    ISSN: 1432-0983
    Keywords: Chromosome fragmentation ; MEL gene family ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Nine members, MEL2–MEL10, of the MEL gene family coding for α-galactosidase were physically mapped to the ends of the chromosomes by chromosome fragmentation. Genetic mapping of the genes supported the location of all the MEL genes in the left arm of their resident chromosomes.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00351706
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  • 10
    Electronic Resource
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    Parameters affecting lithium acetate-mediated transformation of Saccharomyces cerevisiae and development of a rapid and simplified procedure (1993)
    Springer
    Current genetics 24 (1993), S. 455-459 
    Details
    ISSN: 1432-0983
    Keywords: Yeast ; Saccharomyces cerevisiae ; Transformation ; Plasmid
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We have compared a number of procedures for the transformation of whole cells of the yeast Saccharomyces cerevisiae and assessed the effects of dimethylsulphoxide (DMSO) or ethanol, both of which have been reported to enhance transformation efficiency. We find that simplified methods benefit from the addition of one of these compounds, and although differences are observed between strains as to the more beneficial reagent, peak transformation efficiency is, in general obtained with 10% DMSO or 10% EtOH. Increases of between six- and 50-fold are observed, despite a reduction in cell viability, and at this concentration the two compounds are not additive in their effects. The optimum level appears to depend on a balance between improved DNA uptake and reduced cell viability. As a result of this work we present a straightforward and rapid transformation procedure.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00351857
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  • 11
    Electronic Resource
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    Cysteine uptake by Saccharomyces cerevisiae is accomplished by multiple permeases (1999)
    Springer
    Current genetics 35 (1999), S. 609-617 
    Details
    ISSN: 1432-0983
    Keywords: Key words Cysteine uptake ; Amino-acid permeases ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Uptake by Saccharomyces cerevisiae of the sulphur-containing amino acid L-cysteine was found to be non-saturable under various conditions, and uptake kinetics suggested the existence of two or more transport systems in addition to the general amino-acid permease, Gap1p. Overexpression studies identified BAP2, BAP3, AGP1 and GNP1 as genes encoding transporters of cysteine. Uptake studies with disruption mutants confirmed this, and identified two additional genes for transporters of cysteine, TAT1 and TAT2, both very homologous to BAP2, BAP3, AGP1 and GNP1. While Gap1p and Agp1p appear to be the main cysteine transporters on the non-repressing nitrogen source proline, Bap2p, Bap3p, Tat1p, Tat2p, Agp1p and Gnp1p are all important for cysteine uptake on ammonium-based medium. Furthermore, whereas Bap2p, Bap3p, Tat1p and Tat2p seem most important under amino acid-rich conditions, Agp1p contributes significantly when only ammonium is present, and Gnp1p only contributes under the latter condition.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s002940050459
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  • 12
    Electronic Resource
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    Starvation in yeast increases non-adaptive mutation (1999)
    Springer
    Current genetics 35 (1999), S. 77-81 
    Details
    ISSN: 1432-0983
    Keywords: Key words Adaptive mutations ; 6-N-hydroxylaminopurine ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The frequency of reversion in a histidine-requiring mutant of Saccharomyces cerevisiae increases about ten-fold in stationary cells during histidine starvation. Histidine starvation enhances a similar frequency of reversion in a tryptophan-requiring mutant. Starvation, therefore, enhances mutation frequencies in a non-adaptive manner. The base analogue 6-N-hydroxylaminopurine (HAP) added prior to plating on medium with limited histidine strongly increases reversion of the histidine mutant. HAP-induced reversion increases further in stationary starving cells with the same kinetics as that which increases spontaneous reversion. Adding HAP to the stationary starving cells does not produce any effect.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s002940050435
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  • 13
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    Strand interruptions confer strand preference during intracellular correction of a plasmid-borne mismatch in Saccharomyces cerevisiae (1999)
    Springer
    Current genetics 35 (1999), S. 499-505 
    Details
    ISSN: 1432-0983
    Keywords: Key words Heteroduplex repair ; Strand discrimina-tion ; Strand interruptions ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Site-directed mutagenesis was used to construct yeast centromere plasmids in which a strand nick or gap could be placed 5′ or 3′, on either strand, to a reporter gene (SUP4-o) carrying defined base mismatches. The plasmids were then transformed into yeast cells and the direction and efficiency of mismatch repair were assayed by scoring colouring of the transformant colonies. Strands that were nicked were consistently corrected more often than intact strands, but the effect was very small. However, placement of a small gap at the same positions as the nicks resulted in a marked increase in selection for the gapped strand and an enhanced efficiency of mismatch repair. Both the preference for the gapped strand and correction of the mismatch were offset by deletion of the mismatch repair gene PMS1. Together, the results suggest that strand interruptions can direct intracellular mismatch correction of plasmid-borne base mispairs in yeast.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s002940050445
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  • 14
    Electronic Resource
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    Analysis of the genes activated by the FLO8 gene in Saccharomyces cerevisiae (1999)
    Springer
    Current genetics 36 (1999), S. 256-261 
    Details
    ISSN: 1432-0983
    Keywords: Key wordsFLO8 ; Transcriptional regulation ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract It is thought that the FLO8 gene encodes a transcriptional activator of the dominant flocculation gene FLO1 in Saccharomycescerevisiae. To determine other genes which are regulated by FLO8, a detailed comparison of the transcripts from the FLO8 and Δflo8 strains was carried out. In addition to the FLO1 gene, it was found that transcription of the FLO11 and STA1 genes is positively regulated by FLO8. In flo8 strains, not only transcripts of the FLO11, STA1, and FLO1 genes but also invasive growth, extracellular glucoamylase production, and flocculation were undetected. From these results, it is suggested that FLO8 regulates these characteristics via the transcriptional regulation of the FLO11, STA1, and FLO1 genes.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s002940050498
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  • 15
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    Evolutionary conservation of genomic sequences related to the GGP1 gene encoding a yeast GPI-anchored glycoprotein (1993)
    Springer
    Current genetics 23 (1993), S. 19-21 
    Details
    ISSN: 1432-0983
    Keywords: Glycosylphosphatidylinositol anchored-protein ; Southern analysis ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The GGP1 gene encodes the only GPI-anchored glycoprotein (gp115) that has been purified todate in the budding yeast Saccharomyces cerevisiae. It is a single-copy gene whose deduced amino-acid sequence shares no significant homology to any other known protein. In this paper we report a Southern hybridization analysis of genomic DNA from different eukaryotic organisms to identify homologues of the GGP1 gene. We have analyzed DNA prepared from a unicellular green alga (Chlamydomonas eugametos), from two distantly related yeast species (Candida cylindracea and Schizosaccharomyces pombe), and from the common bean Phasoleus vulgaris. The moderate stringency of the experimental conditions and the high specificity of the probes used indicate that a single-copy of GGP1-related sequences exists in all these eukaryotic organisms. The chromosomal localization of the GGP1 gene in S. cerevisiae has also been determined.
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    URL: http://dx.doi.org/10.1007/BF00336744
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  • 16
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    The STA2 and MEL1 genes of Saccharomyces cerevisiae are idiomorphic (1993)
    Springer
    Current genetics 23 (1993), S. 92-94 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Gene mapping ; Idiomorphism
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The STA2 (glucoamylase) gene of Saccharomyces cerevisiae has been mapped close to the end of the left arm of chromosome II. Meiotic analysis of a cross between a haploid strain containing STA2, and another strain carrying the melibiase gene MEL1 (which is known to be at the end of the left arm of chromosome II) produced parental ditype tetrads only. Since there is no significant DNA sequence similarity between the STA2 and MEL1 genes, or their respective flanking regions, we conclude that these two genes are carried by separate non-hybridizing sequences of chromosomal DNA, either of which can reside at the end of the left arm of chromosome II. By analogy with the mating-type locus of Neurospora crassa, we suggest that the STA2 and MEL1 genes are idiomorphs with respect to one another.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00336753
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  • 17
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    Molecular cloning of the yeast OPI3 gene as a high copy number suppressor of the cho2 mutation (1993)
    Springer
    Current genetics 23 (1993), S. 95-101 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Phospholipid synthesis ; Phospholipid-N-methyltransferase ; Mutant ; Over-expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract By functional complementation of the auxotrophic requirements for choline of a cdg1, cho2 double-mutant, by transformation with a genomic DNA library in a high copy number plasmid, two different types of complementing DNA inserts were identified. One type of insert was earlier shown to represent the CHO2 structural gene. In this report we describe the molecular and biochemical characterization of the second type of complementing activity. The transcript encoded by the cloned gene was about 1000-nt in length and was regulated in response to the soluble phospholipid precursors, inositol and choline. A gene disruption resulted in no obvious growth phenotype at 23°C or 30°C, but in a lack of growth at 37°C in the presence of monomethylethanolamine. Null-mutants exhibited an inositol-secretion phenotype, indicative of mutations in the lipid biosynthetic pathway. Complementation analysis, biochemical analysis of the phospholipid methylation pathway in vivo, and comparison of the restriction pattern of the cloned gene to published sequences, unequivocally identified the cloned gene as the OPI3 gene, encoding phospholipid-N-methyltransferase in yeast. When present in multiple copies the OPI3 gene efficiently suppresses the phospholipid methylation defect of a cho2 mutation. As a result of impaired synthesis of phosphatidylcholine, the INO1-deregulation phenotype is abolished in cho2 mutants transformed with the OPI3 gene on a high copy number plasmid. Taken together, these data demonstrate a significantly overlapping specificity of the OPI3 gene product for three sequential phospholipid methylation reactions in the de novo Ptd-Cho biosynthetic pathway.
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    URL: http://dx.doi.org/10.1007/BF00352006
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  • 18
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    Epitope-tagging vectors designed for yeast (1993)
    Springer
    Current genetics 23 (1993), S. 181-183 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; c-myc epitope ; Fusion
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract In order to facilitate the process of epitope-tagging of yeast proteins, we have constructed two Saccharomyces cerevisiae-Escherichia coli shuttle vectors that allow fusion of a sequence encoding an epitope of the human c-myc protein at the 3′ end of any gene. An example of the use of this technique is presented.
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    URL: http://dx.doi.org/10.1007/BF00352019
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  • 19
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    Genetic and molecular analysis of REC114, an early meiotic recombination gene in yeast (1993)
    Springer
    Current genetics 23 (1993), S. 295-304 
    Details
    ISSN: 1432-0983
    Keywords: Meiosis ; Meiotic recombination ; Saccharomyces cerevisiae ; REC114
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Four new meiotic recombination genes were previously isolated by selecting for mutations that rescue the meiotic lethality of rad52 spo13 strains. One of these genes, REC114, is described here, and the data confirm that REC114 is a meiosis-specific recombination gene with no detectable function in mitosis. REC114 is located on chromosome XIII approximately 4,9 cM from CIN4. The nucleotide sequence reveals an open reading frame of 1262 bp, consensus intron splice sites close to the 3′ end, and indicates that the second exon codes for only seven amino acids. In the promoter region, a URS1 consensus sequence (TGGGCGGCTA), identical to the URS1 found in the promoter of SPO16, is present 93 bp upstream of the translation start site. Northern-blot hybridization demonstrates that REC114 is transcribed only during meiosis and that it is not expressed in the absence of the IME1 gene product, even when IME2 is constitutively expressed.
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    URL: http://dx.doi.org/10.1007/BF00310890
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  • 20
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    Lack of correlation between trehalase activation and trehalose-6 phosphate synthase deactivation in cAMP-altered mutants of Saccharomyces cerevisiae (1993)
    Springer
    Current genetics 23 (1993), S. 382-387 
    Details
    ISSN: 1432-0983
    Keywords: Trehalase ; Trehalose-6-P synthase ; cAMP mutants ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The rise in cAMP level that follows the addition of glucose or 2,4-dinitrophenol (DNP) to stationaryphase cells of Saccharomyces cerevisiae was accompanied by a marked activation of trehalase (3-fold increase) and a concomitant deactivation of trehalose-6 phosphate synthase (50% of the basal levels). In glucose-grown exponential cells, which are deficient in glucose-induced cAMP signalling, the addition of glucose also prompted a decrease in trehalose-6 phosphate synthase, but had no effect on trehalase activity. Mutants defective in the RAS-adenylate cyclase pathway (ras1 ras2 bcy1 strain), as well as mutants containing greatly reduced protein kinase activity either cAMP-dependent (tpk w1 BCY1 strains) or cAMP-independent (tpk1 w1 bcy1 strains), were unable to show glucose- or DNP-induced trehalase activation but still displayed a clear decrease in trehalose-6 phosphate synthase activity upon addition of these compounds. These data suggest that the activity of trehalose-6 phosphate synthase, as opposed to that of trehalase, is not controlled by the cAMP signalling pathway “in vivo”. Trehalose-6 phosphate synthase was competitively inhibited by glucose (Ki=15 mM) and resulted unaffected by ATP in assays performed “in vitro”.
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    URL: http://dx.doi.org/10.1007/BF00312622
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  • 21
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    Structure and regulation of the isocitrate lyase gene ICL1 from the yeast Saccharomyces cerevisiae (1993)
    Springer
    Current genetics 23 (1993), S. 375-381 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Isocitrate lyase ; Gene regulation ; Ethanol induction
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The ICL1 gene encoding the isocitrate lyase from Saccharomyces cerevisiae was cloned and sequenced. A reading frame of 557 amino acids showing significant similarity to isocitrate lyases from seven other species could be identified. Construction of icl1 null mutants led to growth defects on C2 carbon sources while utilization of sugars or C3 substrates remained unaffected. Using an ICL1-lacZ fusion integrated at the ICL1 locus, a more than 200-fold induction of β-galactosidase activity was observed after growth on ethanol when compared with glucose-repressed conditions. A preliminary analysis of the ICL1 upstream region identified a 364-bp fragment necessary and sufficient for this regulatory phenotype. Sequence motifs also present in the upstream regions of co-regulated genes were found within this region.
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    URL: http://dx.doi.org/10.1007/BF00312621
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  • 22
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    Phenotypic identification of amplifications of the ADH4 and CUP1 genes of Saccharomyces cerevisiae (1993)
    Springer
    Current genetics 23 (1993), S. 392-396 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Gene amplification ; ADH4 ; CUP1
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Primary gene amplification, i.e., mutation from one gene copy to multiple gene copies per genome, is important in genomic evolution, as a means of producing anti-cancer drug resistance, and is associated with the progression of tumor malignancy. Primary amplification has not been studied in normal eukaryotic cells because amplifications are extremely rare in these cells. A system has been developed to phenotypically identify co-amplifications of the ADH4 and CUP1 genes of Saccharomyces cerevisiae and 21 independent spontaneous amplifications have been isolated.
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    URL: http://dx.doi.org/10.1007/BF00312624
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    Donation of information to the unbroken chromosome in double-strand break repair (1993)
    Springer
    Current genetics 23 (1993), S. 414-422 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Donation ; Gene conversion ; Double-strand break repair ; Heteroduplex DNA
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We have used transformation of yeast with lincarized plasmids to study the transfer of information to the unbroken chromosome during double-strand break repair. Using a strain which carried the wild-type HIS3 allele, and a linearized plasmid which carried a mutant his3 allele, we have obtained His- transformants. In these, double-strand break repair has resulted in precise transfer of genetic information from the plasmid to the chromosome. Such repair events, we suggest, are gene conversions which entail the formation of heteroduplex DNA on the (unbroken) chromosome. If this suggestion is correct, our results reflect the spatial distribution of such heteroduplex DNA. Transfer of information from the plasmid to the chromosome was obtained at a maximal frequency of 1.5% of the repair events, and showed a dependence with distance. Transformation to His- was also obtained with a 2-kbp insertion and with a deletion of 200 bp. The latter results suggest that gene conversion of large heterologies can occur via repair of a heteroduplex DNA intermediate.
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    URL: http://dx.doi.org/10.1007/BF00312628
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    MCB elements and the regulation of DNA replication genes in yeast (1993)
    Springer
    Current genetics 24 (1993), S. 185-192 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Cell cycle ; Transcription ; DNA replication
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract In eukaryotic organisms, genes involved in DNA replication are often subject to some form of cell cycle control. In the yeast Saccharomyces cerevisiae, most of the DNA replication genes that have been characterized to date are regulated at the transcriptional level during G1 to S phase transition. A cis-acting element termed the MluI cell cycle box (or MCB) conveys this pattern of regulation and is common among more than 20 genes involved in DNA synthesis and repair. Recent findings indicate that the MCB element is well conserved among fungi and may play a role in controlling entry into the cell division cycle. It is evident from studies in higher systems, however, that transcriptional regulation is not the only form of control that governs the cell-cycle-dependent expression of DNA replication genes. Moreover, it is unclear why this general pattern of regulation exists for so many of these genes in various eukaryotic systems. This review summarizes recent studies of the MCB element in yeast and briefly discusses the purpose of regulating DNA replication genes in the eukaryotic cell cycle.
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    URL: http://dx.doi.org/10.1007/BF00351790
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    Pentose-phosphate pathway in Saccharomyces cerevisiae: analysis of deletion mutants for transketolase, transaldolase, and glucose 6-phosphate dehydrogenase (1993)
    Springer
    Current genetics 24 (1993), S. 373-376 
    Details
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Pentose-phosphate pathway ; Transketolase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Deletion mutants for the yeast transketolase gene TKL1 were constructed by gene replacement. Transketolase activity was below the level of detection in mutant crude extracts. Transketolase protein could be detected as a single protein band of the expected size by Western-blot analysis in wild-type strains but not in the delection mutant. Deletion of TKL1 led to a reduced but distinct growth in synthetic medium without an aromatic amino-acid supplement. We also isolated double and triple mutants for transketolase (tkl1), transaldolase (tal1), and glucose 6-phosphate dehydrogenase (zwf1) by crossing the different mutants. A tal1 tkl1 double mutant grew nearly like wild-type in rich medium. Only the tkl1 zwf1 double and the tal1 tkl1 zwf1 triple mutant grew more slowly than the wild-type in rich medium. This growth defect could be partly alleviated by the addition of xylulose but not ribose. The triple mutant still grew slowly on a synthetic mineral salts medium without a supplement of aromatic amino acids. This suggests the existence of an alternative but limited source of pentose phosphates and erythrose 4-phosphate in the tkl1 zwf1 double mutants. Hybridization with low stringency showed the existence of a sequence with homology to transketolase, possibly a second gene.
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    Mutant allele pso7-1, that sensitizes Saccharomyces cerevisiae to photoactivated psoralen, is allelic with COX11, encoding a protein indispensable for a functional cytochrome c oxidase (1999)
    Springer
    Current genetics 36 (1999), S. 124-129 
    Details
    ISSN: 1432-0983
    Keywords: Key words Psoralen sensitivity ; Cytochrome oxidase ; Saccharomyces cerevisiae ; Oxidative stress
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The yeast gene PSO7 was cloned from a genomic library by complementation of the pso7-1 mutant's sensitivity phenotype to 4-nitroquinoline-1-oxide (4NQO). Sequence analysis revealed that PSO7 is allelic to the 1.1-kb ORF of the yeast gene COX11 which is located on chromosome XVI and encodes a protein of 28-kDa localized in the inner mitochondrial membrane. Allelism of PSO7/COX11 was verified by non-complementation of 4NQO-sensitivity in diploids homo- and hetero-allelic for the pso7-1 and cox11::TRP1 mutant alleles. Sensitivity to 4NQO was the same in exponentially growing cells of the pso7-1 mutant and the cox11::TRP1 disruptant. Allelism of COX11 and PSO7 indicates that the pso7 mutant's sensitivity to photoactivated 3-carbethoxypsoralen and to 4NQO is not caused by defective DNA repair, but rather is due to an altered metabolism of the pro-mutagen 4NQO in the absence of cytochrome oxidase (Cox) in pso7-1/cox11::TRP1 mutants/disruptants. Lack of Cox might also lead to a higher reactivity of the active oxygen species produced by photoactivated 3-carbethoxypsoralen. The metabolic state of the cells is important for their sensitivity phenotype since the largest enhancement of sensitivity to 4NQO between wild-type (WT) and the pso7 mutant occurs in exponentially growing cells, while cells in stationary phase or growing cells in phosphate buffer have the same 4NQO resistance, irrespective of their WT/mutant status. Strains containing the pso7-1 or cox11::TRP1 mutant allele were also sensitive to the oxidative stress-generating agents H2O2 and paraquat. Mutant pso7-1, as well as disruptant cox11::TRP1, harboured mitochondria that in comparison to WT contained less than 5% and no detectable Cox activity, respectively.
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    Induction of pyruvate decarboxylase in glycolysis mutants of Saccharomyces cerevisiae correlates with the concentrations of three-carbon glycolytic metabolites (1993)
    Springer
    Archives of microbiology 160 (1993), S. 324-328 
    Details
    ISSN: 1432-072X
    Keywords: Saccharomyces cerevisiae ; Pyruvate decarboxylase ; Pyruvate kinase ; Signalling ; Glycolysis mutants
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Pyruvate decarboxylase, PDCase, activity in wild-type yeast cells growing on ethanol is quite low but increases up to tenfold upon addition of glucose, less with galactose and only slightly with glycerol. PDCase levels in glycolysis mutant strains growing on ethanol or acetate were higher than in the wild-type strain. These levels correlated with the sum of the concentrations of three-carbon glycolytic metabolites. The highest accumulation was observed in a fructose bisphosphate aldolase deletion mutant concomintant with the highest PDCase activity wild-type level. On the other hand, the PDCase levels in the different mutants again correlated with the sum of the concentrations of the three-carbon glycolytic metabolites. This was interpreted to mean that full induction of PDCase activity requires the accumulation of hexose-and triosephosphates.
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    Characterization of acetyl-CoA: l-lysine N6-acetyltransferase, which catalyses the first step of carbon catabolism from lysine in Saccharomyces cerevisiae (1993)
    Springer
    Archives of microbiology 160 (1993), S. 397-400 
    Details
    ISSN: 1432-072X
    Keywords: Saccharomyces cerevisiae ; Acetyl-CoA ; l-Lysine N6 ; acetytransferase ; Lysine catabolism
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The carbon catabolism of l-lysine starts in Saccharomyces cerevisiae with acetylation by an acetyl-CoA: l-lysine N6-acetyltransferase. The enzyme is strongly induced in cells grown on l-lysine as sole carbon source and has been purified about 530-fold. Its activity was specific for acetyl-CoA and, in addition to l-lysine, 5-hydroxylysine and thialysine act as acetyl acceptor. The following apparent Michaelis constants were determined: acetyl-CoA 0.8 mM, l-lysine 5.8 mM, dl-5-hydroxylysine 2.8 mM, l-thialysine 100 mM. The enzyme had a maximum activity at pH 8.5 and 37°C. Its molecular mass, estimated by sodium dodecyl sulphate-polyacrylamide gel electrophoresis, was 52 kDa. Since the native molecular mass, determined by gel filtration, was 48 kDa, the enzyme is a monomer.
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    Comparative effects of Saccharomyces cerevisiae cultivation under copper stress on the activity and kinetic parameters of plasma-membrane-bound H+-ATPases PMA1 and PMA2 (1999)
    Springer
    Archives of microbiology 171 (1999), S. 273-278 
    Details
    ISSN: 1432-072X
    Keywords: Key words Plasma membrane H+-ATPase ; PMA1 ; ATPase ; PMA2 ATPase ; Saccharomyces cerevisiae ; Copper stress ; Copper tolerance
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The major yeast plasma membrane H+-ATPase is encoded by the essential PMA 1 gene. The PMA 2 gene encodes an H+-ATPase that is functionally interchangeable with the one encoded by PMA 1 , but it is expressed at a much lower level than the PMA 1 gene and it is not essential. Using genetically manipulated strains of Saccharomyces cerevisiae that exclusively synthesize PMA1 ATPase or PMA2 ATPase under control of the PMA1 promoter, we found that yeast cultivation under mild copper stress leads to a similar activation of PMA2 and PMA1 isoforms. At high inhibitory copper concentrations (close to the maximum that allowed growth), ATPase activity was reduced from maximal levels; this decrease in activity was less important for PMA2 ATPase than for PMA1 ATPase. The higher tolerance to high copper stress of the artificial strain synthesizing PMA2 ATPase exclusively, as compared to that synthesizing solely PMA1 ATPase, correlated both with the lower sensitivity of PMA2 ATPase to the deleterious effects of copper in vivo and with its higher apparent affinity for MgATP, and suggests that plasma membrane H+-ATPase activity plays a role in yeast tolerance to copper.
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    A strange calmodulin of yeast (1999)
    Springer
    Molecular and cellular biochemistry 190 (1999), S. 47-54 
    Details
    ISSN: 1573-4919
    Keywords: calmodulin ; yeast calmodulin ; Ca2+ binding ; Ca2+ binding protein ; Saccharomyces cerevisiae ; interdomain interaction
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Calmodulin of Saccharomyces cerevisiae has different Ca2+ binding properties from other calmodulins. We previously reported that the maximum number of Ca2+ binding was 3 mol/mol and the fourth binding site was defective, which was different from 4 mol/mol for others. Their macroscopic dissociation constants suggested the cooperative three Ca2+ bindings rather than a pair of cooperative two Ca2+ bindings of ordinary calmodulin. Here we present evidence for yeast calmodulin showing the intramolecular close interaction between the N-terminal half domain and the C-terminal half domain, while the two domains of ordinary calmodulin are independent of each other. We will discuss the relationship of the shape and the shape change caused by the Ca2+ binding to the enzyme activation in yeast. The functional feature of calmodulin in yeast will also be considered, which might be different from the one of vertebrate calmodulin.
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    URL: http://dx.doi.org/10.1023/A:1006951710440
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    Intracellular pH inSchizosaccharomyces pombe — Comparison withSaccharomyces cerevisiae (1993)
    Springer
    Molecular and cellular biochemistry 124 (1993), S. 131-140 
    Details
    ISSN: 1573-4919
    Keywords: Schizosaccharomyces pombe ; Saccharomyces cerevisiae ; H+-ATPase ; intracellular pH ; carboxy-seminaphthorhodafluor-1
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract We examined cytoplasmic pH regulation inSchizosaccharomyces pombe andSaccharomyces cerevisiae using pH-sensitive fluorescent dyes. Of several different fluorescent compounds tested, carboxy-seminaphthorhodafluor-1 (C.SNARF-1) was the most effective. Leakage of C.SNARF-1 fromS. pombe was much slower than leakage fromC. cerevisiae. Using the pH-dependent fluorescence of C.SNARF-1 we showed that at an external pH of 7, mean resting internal pH was 7.0 forS. pombe and 6.6 forS. cerevisiae. We found that internal pH inS. pombe was maintained over a much narrower range in response to changes in external pH, especially at acidic pH. The addition of external glucose caused an intracellular alkalinization in both species, although the effect was much greater inS. cerevisiae than inS. pombe. The plasma membrane H+-ATPase inhibitor diethylstilbestrol reduced both the rate and extent of alkalinisation, with an IC50 of approximately 35 μM in both species. Amiloride also inhibited internal alkalinisation with IC50's of 745 μM forS. cerevisiae and 490 μM forS. pombe.
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    Characterization of Saccharomyces cerevisiae strains expressing ira1 mutant alleles modeled after disease-causing mutations in NF1 (1999)
    Springer
    Molecular and cellular biochemistry 202 (1999), S. 109-118 
    Details
    ISSN: 1573-4919
    Keywords: NF1 mutations ; IRA1 ; Saccharomyces cerevisiae ; RAS2 ; GAP activity
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The 2818 amino acids of neurofibromin, the product of the human NF1 gene, include a 230 amino acid Ras-GAP related domain (GRD). Functions which may be associated with the rest of the protein remain unknown. However, many NF1 mutations in neurofibromatosis 1 patients are found downstream of the GRD, suggesting that the C-terminal region of the protein is also functionally important. Since the C-terminal region of neurofibromin encompassing these mutations is homologous with the corresponding regions in the two Saccharomyces cerevisiae Ras-GAPs, Ira1p and Ira2p, we chose yeast as a model system for functional exploration of this region (Ira-C region). Three missense mutations that affect the Ira-C region of NF1 were used as a model for the mutagenesis of IRA1. The yeast phenotypes of heat shock sensitivity, iodine staining, sporulation efficiency, pseudohyphae formation, and GAP activity were scored. Even though none of the mutations directly affected the Ira1p-GRD, mutations at two of the three sites resulted in a decrease in the GAP activity present in ira1 cells. The third mutation appeared to disassociate the phenotypes of sporulation ability and GAP activity. This and other evidence suggest an effector function for Ira1p.
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    Direct observation of oxidative stress on the cell wall of Saccharomyces cerevisiae strains with atomic force microscopy (1999)
    Springer
    Molecular and cellular biochemistry 201 (1999), S. 17-24 
    Details
    ISSN: 1573-4919
    Keywords: Saccharomyces cerevisiae ; atomic force microscope ; bioscope ; organic synthesis ; molecular biology ; oxidative stress ; pore enlargement ; cell wall ; baker's yeast ; biotechnology
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract We imaged pores on the surface of the cell wall of three different industrial strains of Saccharomyces cerevisiae using atomic force microscopy. The pores could be enlarged using 10 mM diamide, an SH residue oxidant that attacks surface proteins. We found that two strains showed signs of oxidative damage via changes in density and diameter of the surface pores. We found that the German strain was resistant to diamide induced oxidative damage, even when the concentration of the oxidant was increased to 50 mM. The normal pore size found on the cell walls of American strains had diameters of about 200nm. Under conditions of oxidative stress the diameters changed to 400nm. This method may prove to be a useful rapid screening process (45-60 min) to determine which strains are oxidative resistant, as well as being able to screen for groups of yeast that are sensitive to oxidative stress. This rapid screening tool may have direct applications in molecular biology (transference of the genes to inside of living cells) and biotechnology (biotransformations reactions to produce chiral synthons in organic chemistry.
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    Antibodies raised against yeast HSP 104 cross-react with a heat-and abscisic acid-regulated polypeptide in rice (1993)
    Springer
    Plant molecular biology 22 (1993), S. 1177-1180 
    Details
    ISSN: 1573-5028
    Keywords: abscisic acid ; developmental regulation ; heat shock proteins ; Oryza sativa ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Antibodies raised against yeast heat shock protein (HSP) 104 recognized a heat-inducible polypeptide with a molecular mass of 110 kDa in shoot tissue of young rice seedlings. Root tissue of the same age showed no immuno-reaction with yeast HSP 104 antibodies. The 110 kDa polypeptide of rice was also shown to be abscisic acid-inducible in young seedlings. Though this polypeptide was seen to be constitutively present in the flag leaf of 90-day-old field-grown plant, it was not much affected by either heat shock or abscisic acid in this case.
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    Characterization of the Oxaloacetate Decarboxylase and Pyruvate Kinase-like Activities of Saccharomyces cerevisiae and Anaerobiospirillum succiniciproducens Phosphoenolpyruvate Carboxykinases (1999)
    Springer
    The protein journal 18 (1999), S. 659-664 
    Details
    ISSN: 1573-4943
    Keywords: Phosphoenolpyruvate carboxykinase ; oxaloacetate decarboxylase ; pyruvate kinase-like activity ; Anaerobiospirillum succiniciproducens ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Abstract Two members of the ATP-dependent class of phosphoenolpyruvate carboxykinases (PEPCKs) (Saccharomyces cerevisiae and Anaerobiospirillum succiniciproducens) have been comparatively studied with regard to their oxaloacetate (OAA) decarboxylase and pyruvate kinase-like activities. The pyruvate kinase-like activities were dependent on the presence of Mn2+; at the same concentrations Mg2+ was not effective. These activities were synergistically activated by a combination of both metal ions. V max for these activities in A. succiniciproducens and S. cerevisiae PEPCKs was 0.13% and 1.2% that of the principal reaction, respectively. The OAA decarboxylase activity was nucleotide independent and, with decreasing order of effectiveness, these activities were supported by Mn2+ and Mg2+. AMP is an activator of these reactions. V max for the OAA decarboxylase activities in A. succiniciproducens and S. cerevisiae PEPCKs was 4% and 0.2% that of the PEP-forming reaction, respectively.
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    Fluorescence staining of yeast cells permeabilized by killer toxin K1: Determination of optimum conditions (1993)
    Springer
    Journal of fluorescence 3 (1993), S. 241-244 
    Details
    ISSN: 1573-4994
    Keywords: Killer toxin K1 ; bromocresol purple staining ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Physics
    Notes: Abstract Optimal assay conditions were established for the previously described method used to determine the activity ofSaccharomyces cerevisiae pore-forming killer toxin K1. The method is based on cell staining with bromocresol purple. Sensitive cells ofS. cerevisiae from the early exponential phase under nongrowth conditions and in the presence of glucose were the most convenient for determining the killer toxin activity. Maximum killing war reached when the suspension was buffered with 10 mM citrate-phosphate at pH 4.6.
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    Genetic evidence for interaction between components of the yeast 26S proteasome: combination of a mutation in RPN12 (a lid component gene) with mutations in RPT1 (an ATPase gene) causes synthetic lethality (1999)
    Springer
    Molecular genetics and genomics 262 (1999), S. 145-153 
    Details
    ISSN: 1617-4623
    Keywords: Key words Proteasome ; Synthetic lethality ; Saccharomyces cerevisiae ; AAA-ATPase ; 19S Regulatory particle
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The 19S regulatory particle of the yeast 26S proteasome consists of six related ATPases (Rpt proteins) and at least 11 non-ATPase proteins (Rpn proteins). RPN12 (formerly NIN1) encodes an Rpn component of the 19S regulatory particle and is essential for growth. To determine which subunit(s) of the 26S proteasome interact(s) with Rpn12, we attempted to screen for mutations that cause synthetic lethality in the presence of the rpn12-1 (formerly nin1-1) mutation. Among the candidates recovered was a new allele of RPT1 (formerly CIM5). This mutant allele was designated rpt1-2; on its own this mutation caused no phenotypic change, whereas the rpn12-1 rpt1-2 double mutant was lethal, suggesting a strong interaction between Rpn12 and Rpt1. The site of the rpt1-2 mutation was determined by DNA sequencing of the RPT1 locus retrieved from the mutant, and a single nucleotide alteration was found. This changes amino acid 446 of the RPT1 product from alanine to valine. The alanine residue is conserved in all Rpt proteins, except Rpt5, but no function has yet been assigned to the region that contains it. We propose that this region is necessary for Rpt1 to interact with Rpn12. The terminal phenotype of the rpn12-1 rpt1-2 double mutant was not cell cycle specific, suggesting that in the double mutant cells the function of the 26S proteasome is completely eliminated, thereby inducing multiple defects in cellular functions.
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    Cat8p, the activator of gluconeogenic genes in Saccharomyces cerevisiae, regulates carbon source-dependent expression of NADP-dependent cytosolic isocitrate dehydrogenase (Idp2p) and lactate permease (Jen1p) (1999)
    Springer
    Molecular genetics and genomics 262 (1999), S. 869-875 
    Details
    ISSN: 1617-4623
    Keywords: Key wordsCAT8 ; Transcriptional regulation ; IDP2 ; JEN1 ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The yeast transcriptional activator Cat8p has been identified as a factor that is essential for the derepression of genes involved in gluconeogenesis (like FBP1, PCK1, ACR1, ICL1 and MLS1) when only non-fermentable carbon sources are provided. Cat8p-dependent expression is mediated by cis-acting elements in the respective promoters, which are named UAS/CSREs (upstream activating sequence/carbon source responsive element). To establish whether the function of Cat8p is restricted to the activation of gluconeogenesis or is also involved in the regulation of a greater variety of genes, we investigated the transcriptional regulation of two genes, IDP2 and JEN1, which exhibit a similar expression pattern to gluconeogenic genes, although IDP2 at least is not linked directly to the gluconeogenic pathway. We identified functional UAS/CSRE elements in the promoters of both genes. Expression studies revealed that JEN1 is regulated negatively by the repressors Mig1p and Mig2p, and that Cat8p is needed for full derepression of the gene under non-fermentative growth conditions. Furthermore, we showed that Mig2p is also involved in the repression of CAT8 itself. The results presented in this study support a model in which Cat8p-dependent gene activation is not restricted to gluconeogenesis, but targets a wide variety of genes which are strongly derepressed under non-fermentative growth conditions.
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    Yeast genes GIS1–4: multicopy suppressors of the Gal− phenotype of snf1 mig1 srb8/10/11 cells (1999)
    Springer
    Molecular genetics and genomics 262 (1999), S. 589-599 
    Details
    ISSN: 1617-4623
    Keywords: Key words Ras/cAMP pathway ; Saccharomyces cerevisiae ; Snf1 ; Mig1 ; Mediator
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Cyclin C and the cyclin C-dependent protein kinase are associated with the RNA polymerase II Mediator complex, which regulates initiation of transcription in response to signals from activators and repressors bound to upstream promoter elements. Disruption of the corresponding genes, SRB11 and SRB10, in budding yeast causes a reduction in expression of the GAL genes, which is particularly pronounced in a mig1 snf1 background. We have screened two yeast genomic libraries for genes that can suppress this phenotype when overexpressed. Seven suppressor genes were identified, GIS1–7. GIS1 encodes one of two related zinc-finger proteins, which also share two other highly conserved domains present in several eukaryotic transcription factors. GIS2 encodes a homologue of the mammalian CNBP and fission yeast Byr3 proteins. GIS3 and GIS4 predict proteins with no obvious similarities to any known proteins. GIS5–7 are identical to the previously described genes PDE2, SGE1 and TUB3, respectively. None of the suppressor genes seem to be involved in Mediator function. Instead, we find that the GIS1, GIS2 and GIS4 genes interact with the CDC25 gene, indicating a possible involvement of these genes in the RAS/cAMP signaling pathway.
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    URL: http://dx.doi.org/10.1007/s004380051121
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    Growth-related expression of ribosomal protein genes in Saccharomyces cerevisiae (1993)
    Springer
    Molecular genetics and genomics 239 (1993), S. 196-204 
    Details
    ISSN: 1617-4623
    Keywords: Saccharomyces cerevisiae ; Ribosomal protein genes ; Transcription activation ; cAMP ; Growth control
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The rate of ribosomal protein gene (rp-gene) transcription in yeast is accurately adjusted to the cellular requirement for ribosomes under various growth conditions. However, the molecular mechanisms underlying this co-ordinated transcriptional control have not yet been elucidated. Transcriptional activation of rp-genes is mediated through two different multifunctional trans-acting factors, ABF1 and RAP1. In this report, we demonstrate that changes in cellular rp-mRNA levels during varying growth conditions are not parallelled by changes in the in vitro binding capacity of ABF1 or RAP1 for their cognate sequences. In addition, the nutritional upshift response of rp-genes observed after addition of glucose to a culture growing on a non-fermentative carbon source turns out not to be the result of increased expression of the ABF1 and RAP1 genes or of elevated DNA-binding activity of these factors. Therefore, growth rate-dependent transcription regulation of rp-genes is most probably not mediated by changes in the efficiency of binding of ABF1 and RAP1 to the upstream activation sites of these genes, but rather through other alterations in the efficiency of transcription activation. Furthermore, we tested the possibility that cAMP may play a role in elevating rp-gene expression during a nutritional shift-up. We found that the nutritional upshift response occurs normally in several mutants defective in cAMP metabolism.
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    URL: http://dx.doi.org/10.1007/BF00281618
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    Inducible removal of UV-induced pyrimidine dimers from transcriptionally active and inactive genes of Saccharomyces cerevisiae (1993)
    Springer
    Molecular genetics and genomics 239 (1993), S. 28-32 
    Details
    ISSN: 1617-4623
    Keywords: Saccharomyces cerevisiae ; UV damage ; Mating type ; Inducible repair
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The prior UV irradiation of α haploid Saccharomyces cerevisiae with a UV dose of 25 J/m2 substantially increases the repairability of damage subsequently induced by a UV dose of 70 J/m2 given 1 h after the first irradiation. This enhancement of repair is seen at both the MATa and HMLα loci, which are, respectively, transcriptionally active and inactive in α haploid cells. The presence in the medium of the protein synthesis inhibitor, cycloheximide in the period between the two irradiations eliminated this effect. Enhanced repair still occurred if cycloheximide was present only after the final UV irradiation. This indicated that the first result is not due to cycloheximide merely blocking the synthesis of repair enzymes associated with a hypothetical rapid turnover of such molecules. The enhanced repairability is not the result of changes in chromatin accessibility without protein synthesis, merely caused by the repair of the damage induced by the prior irradiation. The data clearly show that a UV-inducible removal of pyrimidine dimers has occurred which involves the synthesis of new proteins. The genes known to possess inducible promoters, and which are involved in excision are RAD2, RAD7, RAD16 and RAD23. Studies with the rad7 and rad16 mutants which are defective in the ability to repair HMLα and proficient in the repair' of MATα showed that in rad7, preirradiation enhanced the repair at MATα, whereas in rad16 this increased repair of MATα was absent. The preirradiation did not modify the inability to repair HMLα in either strain. Thus RAD16 has a role in this inducible repair. Inducible repair is also absent in a rad2 strain which cannot repair MATα or HMLα after a single UV dose.
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    URL: http://dx.doi.org/10.1007/BF00281597
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    Determination of intact molecular species of bovine milk 1,2-diacyl-sn-glycero-3-phosphocholine and 1,2-diacyl-sn-glycero-3-phosphoethanolamine by reversed phase HPLC, a multivariate optimization (1993)
    Springer
    Chromatographia 35 (1993), S. 517-523 
    Details
    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Bovine milk ; Multivariate optimization ; Glycerophospholipids ; Molecular species separation ; Chemometrics
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary The molecular species profile, representing the fatty acid combinations on the glycerol backbone, is the most fundamental description of an intact glycerophospholipid class and provides a key to understanding its functional characteristics. We have developed a powerful RP-HPLC method for the separation of molecular species, including so-called “critical pairs”, of natural mixtures of 1,2-diacyl-sn-glycero-3-phosphocholine 1,2-diacyl-sn-glycero-3-phosphoethanolamine from bovine milk and other natural sources. We have used a novel multivariate development and optimization strategy to locate the region of optimum separation conditions within a prechosen domain of the chromatographic parameter space.
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    URL: http://dx.doi.org/10.1007/BF02267910
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    Preparation and evaluation of cross-linked polyacrylate stationary phases for open tubular liquid chromatography (1993)
    Springer
    Chromatographia 35 (1993), S. 597-606 
    Details
    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Open tubular columns ; Cross-linked polyacrylate stationary phase ; Photopolymerization ; Antracene derivatives
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary Polyacrylate films containing a lauryl moiety were fabricated on about 12 μm I.D. fused silica capillaries by in-situ photopolymerization of acrylates. Relatively thick films up to 1 μm were obtained by the technique. The films were tested as retentive layers for reversed phase open tubular liquid chromatography (OTLC). Due to the presence of the incorporated lauryl moiety in the layer considerable retention could be achieved for polar compounds with aqueous mobile phases. The efficiency of the prepared columns approximates to the theoretical value. The polyacrylate films appear to be extremely stable even under strong basic conditions. The films swell considerably in the presence of nonpolar solvents resulting in more favourable retention and a significant improvement of the chromatographic performance.
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    URL: http://dx.doi.org/10.1007/BF02267923
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    Analysis of n-3 fatty acids in fish oils by high-performance liquid chromatography (1993)
    Springer
    Chromatographia 35 (1993), S. 627-630 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Fatty acid analysis ; Fish oils
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary A chromatographic method has been developed for the determination of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) in fish oil dietary supplements containing triglycerides rich in (n−3) polyunsaturated fatty acids (PUFAs). Following the ester saponification, free fatty acids were resolved by TLC into two fractions: i) a more mobile group of saturated and monounsaturated fatty acids and ii) a less mobile group containing PUFAs. The PUFA fraction was further analyzed by HPLC to determine the levels of EPA and DHA in the fish oil sample.
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    URL: http://dx.doi.org/10.1007/BF02267927
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    Determination of carbidopa and levodopa in human plasma by high-performance liquid chromatography with electrochemical detection (1993)
    Springer
    Chromatographia 35 (1993), S. 607-612 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Carbidopa and levodopa in plasma ; Electrochemical detection
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary A validated reversed-phase high-performance liquid chromatographic procedure employing electrochemical detection (LCEC) for the analysis of carbidopa and levodopa in human plasma is reported. The method is sensitive and specific with amperometric detection at a glassy carbon working electrode with Eapp=0.75 V vs. Ag/AgCl. The retention times of levodopa, internal standard, and carbidopa are 3.3, 4.5, and 9.7 minutes, respectively, with an overall chromatographic run time of 12.0 minutes. The peak height ratio versus plasma concentration is linear over the range of 5.0 to 500 ng/mL for each analyte and exhibits correlation coefficients of 0.9957 or better (n=9). The mean absolute recovery of carbidopa and levodopa using the described assay is 36.6 and 66.0%, respectively. The inter- and intra-day accuracy and precision are within 11.8% of the actual values for all concentrations. Also, due to the demonstrated instability of carbidopa and levodopa in plasma a procedure is provided to circumvent this. Blood collected in pre-treated Vacutainer tubes can be stored in an ice bath for up to 4 hours without any significant degradation, thereby providing a practical means for processing several clinical samples simultaneously.
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    URL: http://dx.doi.org/10.1007/BF02267924
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    High-performance liquid chromatographic separation of fullerene mixtures using a novel stationary phase (1993)
    Springer
    Chromatographia 35 (1993), S. 631-636 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Fullerenes ; FullereneSep® ; Separation factor ; HPLC-particle beam mass spectrometry
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary High-performance liquid chromatographic separation of fullerene mixtures in the range C60 to C100 can be achieved using a novel stationary phase which rivals the performance of the more expensive ‘Pirkle-type’ columns currently employed.
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    URL: http://dx.doi.org/10.1007/BF02267928
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    Spurious interferences in the ion chromatographic determination of trace elements in human hair (1993)
    Springer
    Chromatographia 35 (1993), S. 675-678 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Ion chromatography ; Trace metal determination ; Sample contamination ; Human hair
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary The dynamic ion-exchange chromatographic analysis of bivalent ions, such as Cu, Pb, Zn, Ni, Co and Mn in resistant organic matrices needs preliminary wet ashing with oxidizing acids (HNO3 and HClO4). Such treatment may lead to an increase in the amounts of metals present in the sample due to their release from containers or equipment used in an acidic environment and to metal impurities contained in the acids. The effect of these interferences in the determination of trace metals in human hair was studied.
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    URL: http://dx.doi.org/10.1007/BF02267936
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    Individual determination of ortho and non-ortho substituted polychlorobiphenyls (PCBs) in sediments by high performance liquid chromatographic pre-separation and gas chromatography/ECD detection (1993)
    Springer
    Chromatographia 36 (1993), S. 65-70 
    Details
    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; GC/ECD detection ; PCBs, ortho and non-ortho ; Chromatographic separation ; Congener determination ; FTIR identification
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary An analytical procedure for the individual determination of ortho and non-ortho PCB congeners in sediments, using high performance liquid chromatography (HPLC) preseparation and gas chromatography/ECD detection, is described. Gas chromatography/FTIR spectrometry (GC/FTIR) and gas chromatography/mass spectrometry (GC/MS) were employed for individual congener identification and determination. Sample extraction, clean-up of extract and selective elution procedures were optimized by using reference certified marine sediment samples. Recovery and precision were typically 83% and 16% respectively at 2 ng/g of total PCB content. The proposed procedure, tested by analyzing real sediment samples, showed a reproducibility better than 20% at 13 ng/g PCB level.
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    URL: http://dx.doi.org/10.1007/BF02263839
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    HPLC determination of pyroglutamic acid as a degradation product in parenteral amino acid formulations (1993)
    Springer
    Chromatographia 36 (1993), S. 75-78 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Glutamine ; Pyroglutamic acid ; Amino acids formulations
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary The aminoacid glutamine in aqueous solution and in conditions of high temperature and long term storage is partly transformed into pyroglutamic acid which exhibits potential neurotoxic effects. Commercially available aminoacid mixtures supplemented with glutamine are heat-sterilized and some losses of glutamine and formation of pyroglutamic acid may occur. The aim of the work was to set up an easy and reliable HPLC method which allows the determination of pyroglutamic acid as a degradation product of glutamine. The column was a 5 μm Hypersil ODS (100×4.6 mm) and the mobile phase 100% 0.007 M phosphate buffer pH 3.5. Stability studies in different conditions of temperature and time of storage were performed on aminoacid mixture available in the commerce.
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    URL: http://dx.doi.org/10.1007/BF02263841
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    Determination of picomole amounts of carbonyls as luminarin hydrazones by high-performance liquid chromatography with fluorescence detection (1993)
    Springer
    Chromatographia 36 (1993), S. 96-104 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Carbonyl compounds ; Luminarin hydrazides ; Fluorescence detection
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary Fluorogenic reagents (luminarin 3, luminarin 11 and luminarin 12), having a quinolizinocoumarin moiety as fluorophore and a carboxylic acid hydrazide function as reacting group, have been developed. These reagents were found to be highly sensitive fluorescence derivatization reagents for aldehydes and ketones in high-performance liquid chromatography. The reagents readily react with carbonyl compounds in aqueous sulphuric acid solution (0.1 M) at room temperature to produce the corresponding hydrazone derivatives, which can be separated on both reversed or normal-phase column. The structures of the derivatives were studied, together with their properties in reversed and normalphase chromatographic systems. UV absorbance, corrected fluorescence spectral data and quantum yields of luminarin 3, luminarin 11 and luminarin 12 are presented. The detection limits (signal to noise ratio=3) for aldehydes and ketones were in the sub-pmol range. Luminarin 3 was also applied to the determination of hydroxymethylfurfural (HMF) in orange juices and concentrates. The method for HMF involves the solid-liquid extraction of the juice by using a C-18 cartridge prior to derivatization and normal-phase separation of the derivative with fluorimetric detection at 387 nmex., 444 nm em. The calibration curve was linear for amounts of HMF ranging from 0.1 to 10 nmol. Intrarun relative standard deviation was 12.8% for 0.1 nmol and 2.6% for 1 nmol. Recovery studies indicated an average of 98.7±1.9% for juice concentrate and 99.8±3.2% for pasteurized juice.
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    Chemometric optimization in drug analysis by HPLC: A critical evaluation of the quality criteria used in the analysis of drug purity (1993)
    Springer
    Chromatographia 36 (1993), S. 120-124 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Method development ; Chemometrics ; Response criteria ; Analysis of drug purity
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary Optimization procedures require adequate response criteria to assess the quality of each chromatogram obtained during the process. The objective of this paper is to evaluate the possibility of using different resolution functions, in the chromatographic separation of a drug and its impurities (particularly the impurity just eluted after the drug). This study shows the limits of some resolution expressions. The interest of the simple Δt criterion, the difference between retention times, is presented in this paper.
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    Effect of the organic modifier on the retention of retinoids in reversed-phase liquid chromatography (1993)
    Springer
    Chromatographia 36 (1993), S. 147-151 
    Details
    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Polarity ; Retinoids
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary The retention of retinoids in reversed-phase liquid chromatography was studied using aqueous mobile phases of different composition (methanol 94–86% and acetonityrile 92–82%) at five temperatures (40–60 °C). With both organic modifiers the effect of the molecular structure increased as the water content and the polarity of the mobile phase increased. The temperature-dependence increased in the same manner with aqueous acetonitrile mobile phases. The π-π interactions between the retinoids and acetonitrile diminish when the water content of the mobile phase is increased, as happens also to the hydrophobic interactions with both organic modifiers. The net effect of these changes depends on the composition of the mobile phase. There was excellent correlation of retention with all polarity parameters studied(ϕ, P′, xe, xd, xn, E T N , δT, δ, δo and δd), when the calculations were made separately with methanol and acetonitrile. The volume fraction of the organic modifier, ϕ, was the only parameter describing the retention well in both organic modifiers simultaneously.
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    Determination of a new calcium antagonist in human plasma by liquid extraction enrichment and HPLC with UV detection (1993)
    Springer
    Chromatographia 36 (1993), S. 343-346 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Aromatic amines ; Arylbenzylamide methylthioether derivative ; SL 85.1016 ; Pharmacokinetics
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary An HPLC method has been developed for the determination of SL 85.1016, a new calcium antagonist arylbenzylamide methylthioether derivative. SL 85.1016 and the internal standard, SL 87.0210, are extracted from alkaline human plasma withn-hexane and back extracted into 0.05 M phosphate buffer (pH 2.5; 0.2 ml). Acetonitrile (50 μl) is added to the final aqueous extract in order to prevent absorption of the compounds of interest onto the walls of the glass tube; this solution then is partially processed by HPLC on a C18 column with UV detection (254 nm). The determination limit of the method is 2 ng.ml−1 of SL 85.1016 in human plasma; the response to the drug is linear in the range 2–200 ng.mg−1.
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    Analysis ofAllium sulfur amino acids by HPLC after derivatization (1993)
    Springer
    Chromatographia 36 (1993), S. 347-350 
    Details
    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Allium ; Derivatization ; S-alk(en)yl-L-cysteine sulfoxides ; Sulfur amino acids
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary Two pre-column derivatization procedures coupled with reversed phase HPLC have been compared for the analysis ofS-Alk(en)yl0L-cysteine sulfoxides in variousAllium species. In order to establish external standards some (+/-) sulfoxides were synthesized, using a new method to enhance asymmetric synthesis of the diastereoisomers. The first derivatization method is the formation ofo-phthaldialdehyde/tert.-butylthiol derivatives which can be analyzed using UV detection. The second, presently used for amino acid analysis, is the Waters Pico-Tag method, which employs phenylisothiocyanate as derivatization reagent. As the Pico-Tag method was found to be the most efficient for determination ofS-alk(en)yl-L-cysteine sulfoxides it was used to determine the alliin content of various samples of garlic.
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    Collaborative study of the analysis of chlortetracycline by liquid chromatography on octylsilyl silica gel (1993)
    Springer
    Chromatographia 37 (1993), S. 640-644 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Chlortetracycline ; Reversed-phase silical gel ; C8 bonded phase ; Collaborative study
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary An established method for the analysis of chlortetracycline by liquid chromatography using octylsilylated silica gel as the stationary phase was examined in a multicentre study involving five laboratories and a total of six columns. Three chlortetracycline hydrochloride samples were analysed. The main component and the impurities were determined. An analysis of variance, treating each column as a different laboratory, showed an absence of consistent between-laboratory bias and the presence of a significant laboratory-sample interaction. The repeatability and the reproducibility of the method, expressed as relative standard deviations of the result of the determination of chlortetracycline hydrochloride, were calculated to be 0.9% and 1.1% respectively.
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    Characterization of octadecylsilica stationary phases by spectroscopic methods (1993)
    Springer
    Chromatographia 37 (1993), S. 627-634 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; ODS phases ; NMR and FTIR characterization ; Stationary phase functionality
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary Two spectroscopic methods, solid state NMR and diffuse reflectance FTIR, have been used in conjunction with liquid chromatographicretention data to identify and distinguish the functionality of octadecylsilica (ODS) stationary phases. NMR and FTIR spectra are indicative of various features of the different functionalities of the phases and this information can be used to elucidate the configuration of the organosilyl moiety in the ODS on the silica surface and explain why DSS phases have different molecular shape (mainly planarity) recognition capability.
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    Polypyrrole-coated silica as a new stationary phase for liquid chromatography (1993)
    Springer
    Chromatographia 37 (1993), S. 423-428 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Polymer-modified silica ; Polypyrrole cloride ; Polypyrrole dodecylsulfate ; Small molecules ; Polyaromatic hydrocarbons
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary Polypyrrole chloride and polypyrrole dodecylsulfate coated-silica packing materials have been synthesized. Reverse-phase chromtographuy was employed to characterise both packings. A series of test compounds with known properties was used as molecular probes. They included benzene and derivatives, basic drugs, and polyaromatic hydrocarbons. A commericial C18 column was also used for the purpose of comparison in some cases.
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    Enantiomeric resolution ofN-alkyl-N-methylanilineN-oxides by high-performance liquid chromatography (1993)
    Springer
    Chromatographia 37 (1993), S. 487-491 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Chiralcel OD ; N-Alkyl-N-methylanilineN-oxide enantiomers ; Chiral nitrogen centre ; Flavin-containing monooxygenase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary A high-performance liquid chromatographic method for the resolution of the enantiomers of a series ofN-alkyl-N-methylanilineN-oxides is reported. The resolutions were achieved using a Chiralcel, OD chiral stationary-phase with a mobile-phase of hexane and ethanol in varying proportions. The chromatographic order of elution of the enantiomers ofN-ethyl-N-methylanilineN-oxide was determined to be (+)-(R) before (−)-(S).
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    Quantitative determination of sotolon in wines by high-performance liquid chromatography (1993)
    Springer
    Chromatographia 37 (1993), S. 539-542 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Sotolon ; Vin jaune ; Flor-sherry
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary Sotolon (4,5-dimethyl-3-hydroxy-2(5H)-furanone) is a key flavour compound in the french flor-sherry “Vin jaune”. This compound was determined quantitatively by extraction of 25 ml of wine on a XAD-4 resin, elution with diethyl ether, separation by HPLC on a Lichrospher 100 Diol column, elution with dichloromethane/hexane (60/40) and UV detection at 232 nm. The amount of sotolon in “Vin jaune” (120 to 268 μg/l) was related to the development of the yeast film over a period of 6 years. Only 6 to 51 μg/l were found in the “Vin de paille” which is made with overmaturated grapes of the same Savagnin vine-plant but without development of yeasts, and 80 to 140 μg/l in “Tokai” which are partly grown under a yeast film.
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    A micro-scale method employing surface plasmon resonance detection for the determination of conditions for immunoaffinity chromatography of proteins (1993)
    Springer
    Chromatographia 35 (1993), S. 45-49 
    Details
    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Immunoaffinity chromatography ; Proteins ; Surface plasmon resonance detection
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary The BIAcoreTM Biosensor System utilizes a detection principle known as Surface Plasmon Resonance (SPR) which, in simple terms, detects changes in the refractive index of a solution in contact with a gold film. The gold film is surface modified with carboxymethylated dextran to produce a hydrophillic matrix onto which macromolecules may be covalently immobilized. In this respect, the BIAcoreTM is similar to any other insoluble matrix, such as a chromatographic support. The SPR detection principle, however, allows one to directly ‘visualize’ the interaction under study, in real time and without the need for reporter molecules such as enzyme-labels. In addition, the very small sample requirements, automated robotics unit and ease of data analysis suggest the potential use of the BIAcoreTM instrumentation for assay development and possibly process development, especially where bio-specific interactions such as immunoaffinity chromatography are used as a step in the process. In this report, we demonstrate the use of the BIAcoreTM SPR detector for the micro-scale determination of conditions for immunoaffinity chromatography of soluble complementreceptor 1, sCR1.
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    Optimisation of post-column reaction detector for HPLC of explosives (1993)
    Springer
    Chromatographia 35 (1993), S. 5-12 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Post-column reaction detector ; Explosives
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary Improvements in selectivity and sensitivity in the analysis of common explosives, like nitrate esters, nitramines and nitroaromatic compounds can be achieved by post-column derivatisation in a two step reaction detector. The first step in derivatisation is the photolysis of the analytes with UV at 254 nm. The photo reactor consists of a crocheted 20 m Tefzel capillary, which is coiled around a low pressure mercury lamp. In second step the nitrite ion generated is subsequently detected by a colourimetric reaction. The azo dye formed can be selectively detected at 540 nm. Addition of alkali after chromatographic separation to prevent oxidation of initially formed nitrite to nitrate during photolysis leads to a complex multistage arrangement. However, the contribution to peak broadening by the reactor is negligible and it is possible to detect 25–50 ppb of nitramines and 30–100 ppb of nitrate esters. Another advantage of the method is the selective detection of nitro compounds, even in complex matrices. The trace analysis of explosives is of growing interest in forensic science as well as in environmental analysis. It has been shown [1] that explosives can easily be extracted from soil and debris by the use of supercritical carbon dioxide. The separation and determination of explosives by gas chromatography is hindered by their thermal instability. In HPLC only the nitro aromatic explosives can be detected with sufficient sensitivity. Other types of explosives like the esters of nitric acid or nitramines do not absorb sufficiently in the UV region for sensitive detection. It has been shown [2] that explosives are liable to photochemical decomposition in the UV region, resulting in nitrate and nitrite, which have been detected after separation by ion-pair chromatography with electrochemical detection. A more sensitive and selective detection of nitrite has been possible in flow injection analysis [3]. Here a modified Griess reaction has been used. In a first step nitrite ions are used to form the diazonium salt with sulfanilamide which is coupled in a second step with N-[naphthyl-(1)]-ethylene diamine (NED) to form a redviolet azo dye with an absorption maximum at 540 nm. The advantage of this method is selective detection in the visible region, where hardly any other organic components are detected, which might be present in a crude environmental sample. In this paper the transfer of the Griess reaction to post-column derivatisation in RP chromatography of explosives will be described, and the optimisation of trace analysis of these solutes will be discussed.
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    Optimizing separations in reversed-phase liquid chromatography by varying pH and solvent composition (1993)
    Springer
    Chromatographia 35 (1993), S. 18-32 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Reverse phase ; Optimization
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary An interpretive optimization procedure in which pH can be one of the variables is presented with the emphasis on optimizing separations. When varying the pH in reversed-phase liquid chromatography the retention of ionogenic solutes will change. Thus, the selectivity between ionogenic and neutral solutes or between ionogenic solutes mutually can be optimized. However, pH also greatly affects the efficiency (plate count) and peak shape (asymmetry). Optimum selectivity (i.e. large differences in retention times) may be observed under conditions where peaks are broad and asymmetrical. Thus, it is essential to simultaneously consider retention, peak width and peak shape and their effects on separation (effective resolution) in pH-optimization studies. A procedure in which this is done is presented and applied to optimizing the separation of a synthetic mixture of selected pharmaceuticals. After initial experiments to establish the parameter space (boundaries for pH and binary methanol — water composition), twelve experiments are performed according to a 3×4 experimental design. At each loaction the retention, peak height, peak area and peak symmetry are recorded for each solute. These data are then used to build models for each of the four characteristics and for each solute. From this set of models the response surface, describing the quality of separation as a function of pH and composition, can be calculated. A variety of optimization criteria (quantifying quality of separation) can be used. The optimum corresponds to the highest point on the response surface.
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    Separation and identification of fullerenes by liquid chromatography (1993)
    Springer
    Chromatographia 35 (1993), S. 38-44 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Fullerenes ; On-line and off-line identification
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary The separation of fullerenes with a monomeric octadecylsilica bonded phase using n-hexane or toluene/methanol mobile phase systems is described. Analytical and preparative separations, incorporating on-line UV/VIS spectral measurements, confirmed the existence of large fullerenes such as C76, C78 and C84. However, isomers of C78 and C84 were not conclusively found.
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    URL: http://dx.doi.org/10.1007/BF02278554
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    HPLC determination of volatile phenols in wines (1993)
    Springer
    Chromatographia 35 (1993), S. 50-54 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Wine ; Volatile phenols
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary An alternative to the traditional solvent extraction method used to extract and rapidly quantify ethyl-and vinylphenol and ethyl-and vinylgaiacol from wine is presented. The method is based on retention of volatile phenols on adsorbants. Among the tested resins, the most efficient, AG 2-X8 (anion exchange resin), worked as well with a synthetic solution as with wines. The percolation of clarified wine adjusted to pH 9 on this resin permits, in particular, the elimination of organic acids. Phenols are not eluted after rinsing the column with 1N HCl, but are eluted with methanol after this treatment. Good recovery (91 %) and good repeatability are observed. The eluate is directly analysed by HPLC on an RP18 column after two-fold dilution in water. The four volatile phenols were completely separated and detected by UV at 280 nm with high sensitivity (20–40 ppb). No interference with other compounds were noted in the different wines analysed.
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    URL: http://dx.doi.org/10.1007/BF02278556
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    Ion-pair LC of sugars at alkaline pH and pulsed electrochemical detection (1993)
    Springer
    Chromatographia 35 (1993), S. 61-66 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Sugars ; Ion-pair separation ; Pulsed electrochemical detection ; Hypercarb and PLRP-S
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary Sugars, sugar acids, amino sugars and oligomers are separated as ion-pairs with hydrophobic counter ions at alkaline pH using PLRP-S and Hypercarb as solid phases. Important parameters for regulation of retention and selectivity are nature and concentration of the counter ion, pH (hydroxide concentration) and temperature. Reversals in the elution order wer obtained in some cases. Oligosaccharides are highly retained in these systems. The addition of organic modifiers to the mobile phase for elution of the solutes were found to interfere with the pulsed electrochemical detection (PED). Anions added to the mobile phase compete with the solutes for ion-pair retention, hence, decreasing the capacity factors, and phosphate could be used for this purpose in the separation of maltooligomers (M2-M10) from corn syrup.
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    Ion-pair LC of carbohydrates at alkaline pH. Separation and retention behaviour of glycoconjugates (1993)
    Springer
    Chromatographia 35 (1993), S. 55-60 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Carbohydrates ; Ion-pair separation ; Glycoconjugates ; PRP-1 and PLRP-S
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary A new technique for the separation of carbohydrates as ion-pairs in strongly alkaline solution is presented. Carbohydrates are weakly acidic and partly present as anions at pH≥12 [1]. They are retained as ion-pairs on polymeric solid phases (PRP-1 and PLRP-S) with a hydrophobic quaternary ammonium counter ion present in the mobile phase. The effects of nature and concentration of mobile phase components on the retention of carbohydrates have been investigated and an ion-pair distribution model is proposed. The influence of temperature indicated no changes in retention mechanism with high counter ion concentration, but the resolution decreased with increasing temperature. Saccharides added to the mobile phase were shown to increase the retention and the selectivity.
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    The glycan patterns at the individual glycosylation sites in orosomucoid from allergic reaction patients (1993)
    Springer
    Chromatographia 35 (1993), S. 90-92 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; α1-acid glycoprotein ; Complex glycans ; Allergic reaction ; Terfenadine
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary Little is known about the alterations that have occurred at the individual glycosylation sites in allergy patients or how these glycosylation patterns may change after anti-allergy treatments. Using reverse-phase HPLC, we have quantitated the glycoforms present at the individual glycosylation sites on orosomucoid isolated from the sera of allergic reaction patients and an allergic reaction patient treated with the antihistamine Terfenadine. The glycan structures isolated from the five glycosylation sites for the individual taking Terfenadine were all within normal ranges. It is suggested that if the changes in glycosylation in OMD in the allergic state are functionally driven, then it should be possible to correlate biological activities with quantitative changes at the individual glycosylation sites, and hence further define the role of OMD in allergy and inflammation.
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    A comparison of temperature and mobile phase composition effects for bonded liquid crystal and polymeric stationary phases in HPLC (1993)
    Springer
    Chromatographia 35 (1993), S. 85-89 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Bonded liquid crystals ; Polymeric stationary phases ; Variable temperature studies
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary Using two polycyclyic aromatic hydrocarbons as solutes, a comparison is made between a bonded liquid crystal stationary phase and a conventional polymeric C-18 phase. The bonded nematic liquid crystal phase was the silanized form of 4-[4-(allyloxy)benzoyl-oxy]biphenyl and the polymeric phase was Vydac 201TP. Both phases display shape and planarity selectivity as indicated by the results of the variable temperature and mobile phase composition studies. The slot theory of retention can be used to explain these results. However, the liquid crystal phase is more sensitive to molecular geometry, probably due to its more ordered structure on the surface. Variable temperature experiments which compare retention during both heating and cooling provides additional support for this conclusion. With the polymeric bonded C-18 phase, each solute had identical retention at the same temperature during both the heating and cooling cycles. On the bonded liquid crystal phase, measurable differences in retention were observed at identical temperatures depending on whether the column was heated or cooled. This effect is attributed to a degree of partially reversible disordering which occurs as the column temperature was increased. However, conditioning with the appropriate mobile phase can restore the original retention characteristics of the bonded liquid crystal phase.
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    Chromatographic comparison of the relative Lewis acidities of alumina and zirconia (1993)
    Springer
    Chromatographia 35 (1993), S. 133-138 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Ligand exchange chromatography ; Zirconium oxide ; Aluminum oxide ; Lewis acids and bases
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary The chromatographic properties of alumina and zirconia are compared with respect to their relative Lewis acidities when parabenzoic acid derivatives are chromatographed in aqueous media. Weak Bronsted acids show similar capacity factors on alumina and zirconia. As solute Bronsted acidity increases, the capacity factor increases more significantly on the zirconia phase than on the alumina phase. Efficiencies, as determined by the dominant desorption rate constants, are generally comparable between the two phases with alumina showing slightly higher efficiencies, Secondary interactions between the solutes and the stationary phases are nearly absent on zirconia but are very apparent on the alumina phase.
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    Development and comparison of different fluorimetric HPLC-methods with standard methods for the determination of formaldehyde in the atmosphere (1993)
    Springer
    Chromatographia 35 (1993), S. 142-148 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Formaldehyde in air ; Fluorimetric detection ; Comparison of fluorescent derivatives
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary Three different fluorimetric methods for the measurement of atmospheric formaldehyde are described. The relatively unknown 2-diphenylacetylindan-1,3-dionhydrazine (DAIH) method was developed and optimized with an improved limit of detection of 2 ppbv corresponding to a sample volume of 7.5 L. Separation of lutidines necessary for the Nash method was successfully carried out and the separation time was reduced by a factor of 5. The detection limit of the Nash method was improved to 1 ppbv. In comparison, the N,N-diphenyl-1-naphthylamine-5-sulfonhydrazine (DNSH) method performed in a similar manner, reached an LOD of 0.5 ppbv. Finally, measurements made in candle smoke using the three different fluorimetric methods and three standard methods yielded very similar results.
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    HPLC determination of 18 β-glycyrrhetinic and glycyrrhizinic acids in toothpastes after solid phase extraction (1993)
    Springer
    Chromatographia 35 (1993), S. 167-172 
    Details
    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; 18 β-glycyrrhetinic acid ; Glycyrrhizinic acid ; Toothpastes ; Solid phase extraction (SPE)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary HPLC methods suitable for the selective and sensitive determination of 18 β-Glycyrrhetinic acid (GT) and Glycyrrhizinic acid (GZ) in toothpastes have been developed. The methods involve a preliminary quantitative solid phase extraction (C-18 sorbent) for sample clean-up and analyte concentration. Chromatographic separations were performed on reversed phase (C-8; C-18) columns using mixtures of methanol and phosphate buffers (pH 3.0) as the mobile phase under isocratic or gradient elution conditions. The method were applied successfully to the analysis of commercial toothpastes containing low levels (0.013–0.065%) of GT and GZ.
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    Direct enantiomeric analysis of amphetamine in plasma by simultaneous solid phase extraction and chiral derivatization (1993)
    Springer
    Chromatographia 35 (1993), S. 153-159 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Amphetamine in plasma ; Enantiomeric separation ; Solid phase derivatization
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary An analytical approach has been developed for the one step determination of enantiomeric amphetamine composition in plasma, using on-line, pre-column solid phase derivatization with reversed phase HPLC separation. The high molecular weight protein components were excluded by the small pore structure of the polymer and washed out of the reaction column before derivatization. Spiked amphetamine in human plasma was extracted and derivatized by the polystyrene based FMOC-L-prolyl solid phase reagent. The derivatized diastereomers were separated on a conventional ODS column with an ACN/H2O mobile phase. No kinetic resolution or racemization was observed in this solid phase derivatization. Calibration plots and reproducibility experiments were performed to demonstrate the validity of the new approach. Automation of the procedure provided a simple and reproducible method for direct chiral recognition in plasma samples.
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    Determination of trace amounts of morpholine and its thermal degradation products in boiler water by HPLC (1993)
    Springer
    Chromatographia 35 (1993), S. 173-176 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Pre-column derivatisation ; Morpholine and degradation products ; Boiler feed water
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary Morpholine and its amine-type thermal degradation products present in boiler feed water and steam condensate were derivatised with N-succinimidyl-p-nitrophenylacetate. These pre-column derivatives were determined by high-performance liquid chromatography with UV detection at 280 nm. The analytical column was Supelco-sil-ODS with an isocratic mobile phase. Morpholine and its breakdown products were monitored in the range 0.01–10 μg ml−1 with a relative standard deviation of 0.4–3.0%. Chromatographic analysis of boiler feed water and steam condensate samples collected from a boiler servicing a petroleum refinery is described.
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    An improved HPLC system for the analysis of photosynthetic pigments (1993)
    Springer
    Chromatographia 35 (1993), S. 189-192 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Carotenoid pigments ; Chlorophylls ; Photosynthestic tissue
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary A high performance liquid chromatography system for the analysis of photosynthetic pigments is presented. The method employs an octadecylsilica stationary phase, a programmed quaternary mobile phase consisting of mixtures of methanol, acetonitrile, water and hexane, and a photodiode array detector. Carotenoids and chlorophylls are rapidly analysed in a single chromatographic separation. Thecis-trans isomers of most carotenoids are separated by this method.
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    Determination of monensins A and B in the fermentation broth ofStreptomyces cinnamonensis by high performance liquid chromatography (1993)
    Springer
    Chromatographia 35 (1993), S. 206-208 
    Details
    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Monensins A and B ; Fermentation broth ; Streptomyces cinnamonensis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary High performance liquid chromatography on an octadecyl silica column has been used to determine both the monensin A: monensin B ratio and by the method of standard addition, the concentration of both in the fermentation broth ofStreptomyces cinnamonensis. Refractive index detection was preferred to ultraviolet owing to the presence of UV-absorbing components which could not be completely separated from the substances of interest. A linear relationship was obtained from the calibration data. The coefficients of variation for the estimation both of the ratio and the concentrations of the compounds were better then 5%. The estimated limit of detection for both substances was about 1 μg/ml. The results obtained from the determination of the ratios of monensins were compared with those obtained by chemical ionization mass spectrometry. Chromatographic separation of monensins on the silica gel column is also described.
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    HPLC determination of antioxidant synergists and ascorbic acid in some fatty pharmaceuticals, cosmetics and food (1993)
    Springer
    Chromatographia 35 (1993), S. 232-236 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Ascorbic acid ; Antioxidant synergists ; Food, pharmaceuticals and cosmetics
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary A specific and sensitive reverse-phase HPLC method for the quantitative determination of ascorbic acid and antioxidant synergists (1-tartaric acid, citric acid, lactic acid as lithium lactate and EDTA) in fatty pharmaceuticals, cosmetics and food has been developed. Two extraction procedures were used; treatment with hot water, and extraction with water from a hexane dilution of the product. No significant differences between the two procedures were found (p〈0.05), except for ascorbic acid. Quantitative determinations were performed using a C-18 column and sulfuric acid (pH 1.95) mobile phase. With detection, at 210 nm, lactic acid overlapped with ascorbic acid, but the former could be readily identified by TLC. Ascorbic acid was detected at 254 nm, when lactic acid (as lithium lactate) did not interfere in the analysis. Mean recoveries for tartaric, citric and lactic acids were in the range 96–101%.
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    A study of the use of microdialysis probes as a sampling unit in on-line bioprocess monitoring in conjunction with column liquid chromatography (1993)
    Springer
    Chromatographia 35 (1993), S. 285-289 
    Details
    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Microdialysis ; Fermentation monitoring
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary An automated on-line sampling and analytical set-up for the control of fermentations was studied incorporating a microdialysis probe as the sampling device. Applications to a penicillin broth and an ethanol fermentation were studied. Typical recovery values of carbohydrates were found to be close to 100% even after exposure of the microdialysis probe in the process for about 30 h.
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    The on-line determination of the enantiomeric purity of enzymatically produced chiral compounds (1993)
    Springer
    Chromatographia 35 (1993), S. 264-268 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Enantiomer separation ; Stereospecific synthesis ; Crown ether phase ; Immobilized enzyme reactor
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary A prototype of a coupled-column chromatographic system has been developed to determine the enantiomeric purity of enzymatically produced chiral compounds. In this system, an immobilized enzyme HPLC reactor based upon α-chymotrypsin (ACHT-IMER) was linked through a switching valve to a chromatographic column containing a crown ether-based chiral stationary phase (CR-CSP). Although ACHT is normally stereospecific for the L-enantiomorphs of aromatic amino acids, L-tryptophan (L-TRP), L-tyrosine and L-phenylalanine, we were able to achieve the hydrolysis of D-TRP methyl and ethyl esters on the ACHT-IMER. The percent of hydrolysis of the D-TRP esters could be manipulated by varying the resident time in the ACHT-IMER. In a series of experiments, D,L-TRP methyl or ethyl esters were injected into the ACHT-IMER, the flow stopped and after resuming the flow, the eluent which contained the hydrolyzed aminoacids, D- and L-TRP, was switched onto the CR-CSP where the enantiomeric composition of the TRP peak was determined. This configuration is a model for other IMER/CSP coupled-column systems which can be used for analytical and preparative applications.
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    Determination of the lipophilicity of some anti-hypoxia drugs: Comparison of TLC and HPLC methods (1993)
    Springer
    Chromatographia 35 (1993), S. 275-280 
    Details
    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Thin-layer chromatography ; Anti-hypoxia drugs ; Lipophilic character and retention
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary The lipophilicity of 14 anti-hypoxia drugs has been determined by reversed phase thin-layer (RPTLC) and reversed phase high performance liquid chromatography (RPHPLC) in eluent systems containing different concentrations of acetonitrile and potassium dihydrogen phosphate. There was significant correlation between lipophilicity and the specific hydrophobic surface area of the drugs in RPTLC, indicating that the drugs behave as an homologous series of compounds. In RPTLC the concentration of buffer has a negligible effect on the retention of the drugs whereas in RPHPLC the buffer concentration influenced the retention. This discrepancy can be explained by the lower sensitivity of RPTLC. There was strong correlation between lipophilicity values determined by both methods, proving that both are suitable for the determination of molecular lipophilicity.
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    Enantiomeric separation of close analogs to naturally occurring 1,4-benzoxazin-3-ones by liquid chromatography using β-cyclodextrin-modified stationary phase (1993)
    Springer
    Chromatographia 35 (1993), S. 302-304 
    Details
    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; 1,4-Benzoxazin-3-ones ; Cyclic hydroxamic acids ; β-Cyclodextrin modified silica gel ; Enantiomer separation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary A liquid chromatographic procedure on a 5 μm LiChroCART column using chemically bonded β-cyclodextrin (ChiraDex) as chiral selector was developed for the enantiomeric separation of four 2H-1,4-benzoxazin-3(4H)-ones which are close analogs of the cyclic hydroxamic acids 2,4-dihydroxy-2H-1,4-benzoxazin-3(4H)-one (DIBOA) and 2,4-dihydroxy-7-methoxy-2H-1,4-benzoxazin-3 (4H)-one (DIMBOA) naturally occurring inGramineae species.
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    A particle size distribution analysis of used HPLC column packing material (1993)
    Springer
    Chromatographia 35 (1993), S. 295-301 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Used column packings ; Particle size distribution ; Scanning electron microscopy ; Column age
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary Particle size distribution analysis and scanning electron microscopy (SEM) were carried out on eight used HPLC columns containing either irregular silica based, spherical silica based or spherical polymer based packing material. Particle size distributions of the used irregular silica based columns were at least bimodat at the outlet ends and either biomodal or log-normal at the inlet ends with regular progressions between the two extremes through the column. A new ODS-3 column showed log-normal size distributions from the inlet to the outlet ends. Spherical silica based column particle size distributions showed distinct shoulders on large central distribution peaks in most column sections with various degrees of shoulder erosion. The spherical resin based column showed a broader inlet particle size distribution progressing to a very narrow outlet distribution. SEMs of both irregular and spherical silica based columns revealed a larger number of undersized particles and debris at the outlet than inlet ends which could have resulted from stationary phase degradation, since this was not seen in the new ODS-3 column. While several SEMs of the spherical silica based columns revealed hollow spheres and twins, the spherical resin based column packing showed stress fractures or wrinkle lines resulting from use or dehydration.
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    Simultaneous isolation and determination of esculin and rutin in natural materials using SPE and HPLC (1993)
    Springer
    Chromatographia 35 (1993), S. 311-316 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Isolation of natural products ; Solid phase extraction ; Chemically bonded phase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary Esculin (ESC) and rutin (RUT) have been simultaneously isolated from pharmaceutical natural materials by solid phase extraction (SPE). Determination of both substances was performed by reversed phase high performance liquid chromatography (RPHPLC) with UV detection. Optimization of the separation conditions showed that simultaneous isolation and determination of rutin and esculin from pharmaceutical material was possible. The recovery obtained was not lower than 95±2%.
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    URL: http://dx.doi.org/10.1007/BF02277516
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    Direct enantiomeric separation of racemic precursors to diltiazem hydrochloride and clentiazem maleate by HPLC on a chiral ovomucoid column (1993)
    Springer
    Chromatographia 35 (1993), S. 305-310 
    Details
    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Enantiomer separation ; Chiral ovomucoid column ; Diltiazem hydrochloride ; Clentiazem maleate
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary A direct HPLC separation method was developed for the determination of the enantiomers of racemic precursors to diltiazem (I) and its 8-chloro derivatives (II). The enantiomers were successfully separated on a chiral ovomucoid column using an aqueous-organic mobile phase (reversed-phase HPLC). The influence of the organic modifier and buffer pH on the retention and enantioselectivity was investigated. The chromatographic conditions chosen for the separation permitted complete resolution of the enantiomers of both the acid (Ib and IIb) and methyl ester precursors (Ia and IIa) within 20 min. The influence of sample load on retention times, theoretical plates numbers, peak heights and peak areas was also investigated. The peak areas showed a good linearity over the concentration range examined, although all the others were influenced significantly by the sample size. An optical antipode of the intermediate to be determined could be detected by the area-percentage method down to ca. 0.1%, together with the determination of its precursor, including its optical purity.
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    URL: http://dx.doi.org/10.1007/BF02277515
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  • 84
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    Sensitive determination of probenecid in urine samples by reversed-phase liquid chromatography and UV-visible detection using solid-phase extraction techniques for sample clean-up (1993)
    Springer
    Chromatographia 35 (1993), S. 317-320 
    Details
    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Probenecid in urine ; Solid-phase extraction
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary This study describes a rapid method for the determination of probenecid in human urine by liquid chromatography with UV detection at 254 nm, after clean-up through a C8 solid-phase extraction column. Liquid chromatography was carried out on a C18-bonded phase using an acetonitrile-acetate buffer (pH=4) gradient elution. Ethacrynic acid was used as internal standard. The system has been applied to the determination of probenecid in the 0.10–100.0 μg/ml concentration range; the limit of detection was 5 ng/mL.
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    URL: http://dx.doi.org/10.1007/BF02277517
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  • 85
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    Effects of organic modifier and ion-pair reagent in liquid chromatography (1993)
    Springer
    Chromatographia 35 (1993), S. 390-394 
    Details
    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Electrostatic model ; Ion-pair reagent ; Organic modifier ; Prediction of k′ values
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary The capacity factors of phenylamine and naphthylamine sulphonic acids in reversed-phase, ion-pair liquid chromatography (RP-IPC) were measured. The combined effects of organic modifier (Cb) and ion-pair reagent concentration (Cp) on retention follow an equation based on the electrostatic model: $$\ln k' = a + b lnC_p + cC_b $$ the experimental value of b correlates well with the value of 0.5 z (z=charge on analyte ion) predicted for mono and divalent ions. The measured value of b is, however, lower than that predicted for trivalent ions, which may be due to the effective number of charges being less than the apparent number. The absolute values of a and c are much larger than those in RP-HPLC in the absence of an ion-pair reagent, quantitative correlation of a and c with retention values in RP-HPLC and solute charges has been observed and a good linear relationship between a and c has been obtained, strongly supporting the validity of the electrostatic retention model. A critical value, R, at which the negative effect of methanol on retention is equal to the positive effect of the ion-pair reagent (TBAI) has been proposed. The critical values obtained are related to the behaviour of the solute, the ion-pair reagent and the stationary and mobile phases, and lies between −0.04 and −0.065 in most cases, which means that the increase by exp times (=2.718×) the ion-pair concentration the original is equivalent to a decrease of 0.04–0.065 in volume fraction of methanol
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    URL: http://dx.doi.org/10.1007/BF02278591
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    Simultaneous determination of two serine proteinases by hydrophobic interaction chromatography (1993)
    Springer
    Chromatographia 35 (1993), S. 381-386 
    Details
    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Bovine trypsin ; Porcine kallikrein ; Bovine pancreatic trypsin inhibitor
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary The possibility of determining more than one enzyme at the same time has been examined. A new approach, based on the measurement of a direct and specific chromatographic signal obtained by hydrophobic interaction chromatography (HIC) of the stable complex formed with the inhibitor aprotinin, is proposed. A basic procedure for the quantitative determination of trypsin and kallikrein, taken as models, is described. The method is precise with a mean coefficient of variation of 3.1% and 3.5% for trypsin and kallikrein, respectively; the limit of determination for both enzymes is 0.17 nmol ml−1 in the original sample.
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    URL: http://dx.doi.org/10.1007/BF02278589
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  • 87
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    Influence of buffer composition on protein adsorption on agarose (Sepharose 6B) (1993)
    Springer
    Chromatographia 35 (1993), S. 387-389 
    Details
    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Buffer composition effect ; Agarose ; Albumin ; Ionic adsorption
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary The adsorption of albumin on a commercial, high quality agarose (Sepharose® 6B) was studied in different buffers of low pH (4.00) and low molarity (9–11 mM). Eluents containing 0.025% albumin were introduced onto small gel columns until saturation and the adsorption capacity of the gel in different buffers calculated. Adsorption was high in solutions of monobasic acids. Although cation-exchange interactions between the protein and the gel matrix were found to be influenced essentially by the sodium ion concentration or ionic strength of the buffer, adsorption in cirate buffers is weak compared to that in formate, acetate or propionate buffers of about the same ionic strength (I=0.01).
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    URL: http://dx.doi.org/10.1007/BF02278590
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  • 88
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    Liquid chromatographic studies for essential fatty acids on a commercial alkyl phenyl bonded silica column (1993)
    Springer
    Chromatographia 35 (1993), S. 399-402 
    Details
    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Polyunsaturated fatty acids ; Direct injection ; Enthalpy of adsorption
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary Retention studies on arachidonic acid 5, 8, 11, 14 all cis eicosatetraenoic acid, C20:4ω6), eicosapentaenoic acid (5, 8, 11, 14, 17 all cis eicosapentaenoic acid, C20∶5ω3) and docosahexaenoic acid (4, 7, 10, 13, 16, 19 all cis docosahexaenoic acid C22∶6ω3) were performed on a commercial μBondapak free fatty acid analysis column. The ternary mobile phase consisting of acetonitrile, water and tetrahydrofuran was used in an isocratic mode with differential refractometry detection. Retention data were mesured at various flow rates with two different, mobile-phase compositions. Capacity factors and enthalpy of adsorption were calculated from the retention data. Finally the retention mechanism is explained.
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    URL: http://dx.doi.org/10.1007/BF02278593
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    A model for polybutadiene coatings on porous silica (1993)
    Springer
    Chromatographia 35 (1993), S. 403-409 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Reversed phase packings ; Polybutadiene coating ; Polymer coatings
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary Non-wetting viscous liquids such as oligobutadiene prefer “active” sites such as pores during the process of physisorption. Thus, polybutadiene (PBD) coatings on porous silica do not result in a homogeneous polymer film but in an inhomogeneous loading where the bulk polymer is mainly sited in the pores of the silica. An increasing polymer loading leads to increasingly filled pores instead of a thicker polymer film. We cannot exclude the possibility that most of the surface is covered at least with a thin polymer film since the chromatographic behaviour is relatively good for polypeptides, which are highly susceptible to the silanol groups of silica.
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    URL: http://dx.doi.org/10.1007/BF02278594
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    Highly sensitive method for the determination of a new anthracycline: Pirarubicin (1993)
    Springer
    Chromatographia 35 (1993), S. 435-438 
    Details
    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Pirarubicin and metabolites ; Anthracyclines
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary A new and highly sensitive HPLC method for the simultaneous determination of pirarubicine (THP-doxorubicin) and its metabolites, adriamycin and adriamycinol, in human plasma, is described. Samples were treated by liquid-liquid extraction, the organic phase removed and the residue dissolved in methanol. Separation was on a Lichrocart Supersher RP 8 column, (250×4 mm) 4 μm, with a mobile phase of acetonitrile/methanol/formate-buffer.
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    URL: http://dx.doi.org/10.1007/BF02278598
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    The mechanism of separation of polythiolpeptides and metallopolythiolpeptides by covalent affinity chromatography (1993)
    Springer
    Chromatographia 35 (1993), S. 439-447 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Affinity chromatography ; Polythiolpeptides and metallopolythiolpeptides ; Mechanism of separation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary Covalent affinity chromatography with thiol-disulphide interchange (CAD-TDI) is a method for the separation of thiolproteins based on the interaction between disulphide bridges immobilised on a insoluble support and thiol groups in the protein undergoing separation. Mercury-thionein, cadmium-thionein and apothionein have been used as low molcular weight thiolproteins rich in cysteine residue. The proposed separation mechanism is more complex than those in the literature and can have anionic, cationic or a free radical character depending on the protein, the ligand and the conditions during separation.
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    URL: http://dx.doi.org/10.1007/BF02278599
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    Determination of phenolic antioxidants in pharmaceutical formulations by liquid chromatography and migration study on HDPE packagings (1993)
    Springer
    Chromatographia 35 (1993), S. 455-458 
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    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; UV and electrochemical detectors ; Phenolic antioxidants ; Migration studies
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary The performance of UV diode-array, spectrometric and electrochemical detectors was compared in the chromatographic analysis of trace amounts of six phenolic antioxidants. Quantitative validation was undertaken; the linearity range was wider using UV detection although the limits of detection were lower with electrochemical detection. UV detection was applied both to identification of an antioxidant in a hydrophilic suspension and to a migration study.
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    URL: http://dx.doi.org/10.1007/BF02278602
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    Quantitative structure chromatography relationships in reversed-phase high performance liquid chromatography: Prediction of retention behaviour using theoretically derived molecular properties (1993)
    Springer
    Chromatographia 37 (1993), S. 241-249 
    Details
    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Pattern recognition ; Principal components ; Multiple linear regressions ; M.O. properties
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary The use of theoretically calculated molecular properties as predictors for retention in reversed-phase HPLC has been explored. HPLC retention times have been measured for a series of 47 substituted aromatic molecules in three solvent mixtures and steric and electronic properties of these compounds have been derived using semi-empirical molecular orbital and empirical theoretical methods. A subset of the experimental data (a training set) was used to derive property-retention time relationships and the remaining data were then used to test the predictive capability of the methods. Good retention time prediction was possible using derived regression equations for individual solvents and after including solvent parameters it was possible to predict retention for all solvents using a single equation. This method showed that the most useful properties were calculated log P and the calculated dipole moment of the solutes, and the calculated solvent polarisability. In addition, 90% of the data were used to train an artificial neural network and the remaining 10% of the data used to test the network; excellent prediction was obtained, the neural network approach being as successful as the regression analysis.
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    URL: http://dx.doi.org/10.1007/BF02278628
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    Composition of 2-(3,5-dibromo-2-pyridylazo) diethylaminophenol chelates by liquid chromatography (1993)
    Springer
    Chromatographia 37 (1993), S. 284-286 
    Details
    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Composition of chelates ; 3,5-diBr-PADAP
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary The composition of 3,5-diBr-PADAP metal chelates was determined by liquid chromatography employing appropriate eluents and non-polar bonded stationary phase. The metal-to-ligand ratios were 1∶2, 1∶2 and 1∶2 for Cu(II), Co(III) and Cr(III) respectively, and the V(V)-to-ligand ratio found to be 1∶1∶1 in V(V)-3,5-diBr-PADAP-H2O2 in the presence of H2O2. The results are in agreement with literature data.
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    URL: http://dx.doi.org/10.1007/BF02278635
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    HPLC determination of chlorine in air and water samples following precolumn derivatization to 4-bromoacetanilide (1993)
    Springer
    Chromatographia 37 (1993), S. 492-496 
    Details
    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Chlorine in air and water ; Precolumn derivation ; 4-Bromoacetanilide
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary Chlorine has been be determined in air and water samples by a rapid and sensitive method entailing precolumn derivatization to 4-bromoacetanilide. A mixed potassium bromide-acetanilide reagent was used as a trapping agent for chlorine in air, and for its derivatization. The 4-bromoacetanilide formed was determined by reversed-phase HPLC on an ODS column, using methanol-water, 65∶35 (v/v) as mobile phase; detection was at 240 nm. A rectilinear calibration graph was obtained for the range 0.1–30 μg mL−1 chlorine; the limit of detection found to be 0.01 μg mL−1. The precolumn derivative has been found to have a shelf-life of at least 21 days; this enables the use of the method for samples transported from the field to the analytical laboratory, or the testing of a variety of conditions for chlorine scrubbing studies without the need for immediate analysis of samples. Humic substances do not cause any interference with the proposed method and the presence of nitrite does not lead to artificially high results and consequent misleading conclusions of the presence of high levels of chlorine.
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    URL: http://dx.doi.org/10.1007/BF02275785
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    Polymers immobilized on silica gels as stationary phases for liquid chromatography (1993)
    Springer
    Chromatographia 37 (1993), S. 549-561 
    Details
    ISSN: 1612-1112
    Keywords: Column liquid chromatography ; Stationary phases ; Polymer-coated silica ; Polymer-filled silica
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Summary A variety of stationary phases for HPLC can be prepared by immobilization of organic polymers on silica gels. Such silica-polymer adsorbents combine the strength of an inorganic matrix with the selectivity and chemical inertness of organic resins. Moreover, effective shielding of residual silanols and the increase of column packing stability can also be achieved by modification of silica gels with organic polymers. There are two main approaches to the preparation of such composite sorbents. The first includes various routes for coating the silica surface with a polymer layer and the second is by filling the silica pores with a polymer network. A survey of polymers immobilized on silica gels and of various experimental procedures which can be used for composite sorbent synthesis is given in this review.
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    URL: http://dx.doi.org/10.1007/BF02275796
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    Differential expression of two genes encoding isoforms of the ATPase involved in sodium efflux in Saccharomyces cerevisiae (1993)
    Springer
    Molecular genetics and genomics 236 (1993), S. 363-368 
    Details
    ISSN: 1617-4623
    Keywords: Saccharomyces cerevisiae ; Sodium efflux ; Lithium efflux ; ATPase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The ENA2 gene encoding a P-type ATPase involved in Na+ and Li+ effluxes in Saccharomyces cerevisiae has been isolated. The putative protein encoded by ENA2 differs only in thirteen amino acids from the protein encoded by ENA1/PMR2. However, ENA2 has a very low level of expression and for this reason did not confer significant Li+ tolerance on a Li+ sensitive strain. ENA1 and ENA2 are the first two units of a tandem array of four highly homologous genes with probably homologous functions.
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    URL: http://dx.doi.org/10.1007/BF00277134
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    Post-transcriptional regulation of IME1 determines initiation of meiosis in Saccharomyces cerevislae (1993)
    Springer
    Molecular genetics and genomics 237 (1993), S. 375-384 
    Details
    ISSN: 1617-4623
    Keywords: Regulation of meiosis ; Saccharomyces cerevisiae ; IME1
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The IME1 gene of Saccharomyces cerevisiae is required for initiation of meiosis. Transcription of IME1 is detected under conditions which are known to induce initiation of meiosis, namely starvation for nitrogen and glucose, and the presence of MATa1 and MATα2 gene products. In this paper we show that IME1 is also subject to translational regulation. Translation of IME1 mRNA is achieved either upon nitrogen starvation, or upon G1 arrest. In the presence of nutrients, constitutively elevated transcription of IME1 is also sufficient for the translation of IME1 RNA. Four different conditions were found to cause expression of Imel protein in vegetative cultures: elevated transcription levels due to the presence of IME1 on a multicopy plasmid; elevated transcription provided by a Gal-IME1 construct; G1 arrest due to α-factor treatment; G1 arrest following mild heat-shock treatment of cdc28 diploids. Using these conditions, we obtained evidence that starvation is required not only for transcription and efficient translation of IME1, but also for either the activation of Ime1 protein or for the induction/activation of another factor that, either alone or in combination with Ime1, induces meiosis.
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    URL: http://dx.doi.org/10.1007/BF00279441
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    Molecular and genetic characterization of SPT4, a gene important for transcription initiation in Saccharomyces cerevisiae (1993)
    Springer
    Molecular genetics and genomics 237 (1993), S. 449-459 
    Details
    ISSN: 1617-4623
    Keywords: Saccharomyces cerevisiae ; Transcription ; spt mutants
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Mutations in the SPT4 gene of Saccharomyces cerevisiae were isolated as suppressors of δ insertion mutations that interfere with adjacent gene transcription. Recent genetic evidence indicates that the SPT4 protein functions with two other proteins, SPT5 and SPT6, in some aspect of transcription initiation. In this work we have characterized the SPT4 gene and we demonstrate that spt4 mutations, like spt5 and spt6 mutations, cause changes in transcription. Using the cloned SPT4 gene, spt4 null mutations were constructed; in contrast to spt5 and spt6 null mutants, which are inviable, spt4 null mutants are viable and have an Spt− phenotype. The DNA sequence of the SPT4 gene predicts a protein product of 102 amino acids that contains four cysteine residues positioned similarly to those of zinc binding proteins. Mutational analysis suggests that at least some of these cysteines are essential for SPT4 function. Genetic mapping showed that SPT4 is a previously unidentified gene that maps to chromosome VII, between ADE6 and CLY8.
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    URL: http://dx.doi.org/10.1007/BF00279450
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    Characterization of the cyrl-2 UGA mutation in Saccharomyces cerevisiae (1993)
    Springer
    Molecular genetics and genomics 237 (1993), S. 463-466 
    Details
    ISSN: 1617-4623
    Keywords: Saccharomyces cerevisiae ; cyrl-2 ; Nonsense mutation ; CAMP
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary cyrl-2 is a temperature-sensitive mutation of the yeast adenylate cyclase structural gene, CYR1. The cyrl-2 mutation has been suggested to be a UGA mutation since a UGA suppressor SUP201 has been isolated as a suppressor of the cyrl-2 mutation. Construction of chimeric genes restricted the region containing the cyrl-2 mutation, and the cyrl-2 UGA mutation was identified at codon 1282, which lies upstream of the region coding for the catalytic domain of adenylate cyclase. Alterations in the region upstream of the cyrl-2 mutation site result in null mutations. The complete open reading frame of the cyrl-2 gene expressed under the control of the GAL1 promoter complemented cyrl-dl in a galactose-dependent manner. These results suggest that at the permissive temperature weak readthrough occurs at the cyrl-2 mutation site to produce low levels of active adenylate cyclase. An endogenous suppressor in yeast cells is assumed to be responsible for this readthrough.
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    URL: http://dx.doi.org/10.1007/BF00279452
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