bluuebunny/crossref_metadata_2025_split huggingface dataset:
Vectors for the inducible overexpression of glutathione S‐transferase fusion proteins in yeast
Authors: Mitchell David A. , Marshall Tricia K. , Deschenes Robert J. 
Published: May 2005

AbstractA rapid and convenient method of protein purification involves creating a fusion protein with glutathione S‐transferase (GST) (Smith and Johnson, Gene 67, 31–40, 1988). In this report, we describe two vectors for the conditional expression of GST fusions in Saccharomyces cerevisiae. The parent plasmid is based on a high‐copy, galactose‐inducible shuttle vector previously described (Baldari et al., EMBO J. 6, 229–243, 1987). We have demonstrated the use of this system by creating fusions between GST and the yeast RAS2 gene. GST‐Ras2 fusion proteins undergo the post‐translational modifications required for Ras2p to become membrane localized. These vectors provide a useful system for the expression an dpurification of eukaryotic proteins requiring post‐translational modification.
Regulation of glycolytic enzymes and the crabtree effect in galactose‐limited continuous cultures of Saccharomyces cerevisiae
Authors: Sierkstra Laurens N. , Nouwen Nico P. , Verbakel John M. A. , Verrips C. Theo 
Published: May 2005

AbstractIn order to determine whether the changes in the activities and mRNA levels of enzymes involved in intermediary carbon metabolism previously observed in glucose‐limited continuous cultures (Sierkstra et al., 1992a) were glucose specific, we have analysed their regulation in a galactose‐limited continuous culture of Saccharomyces cerevisiae. The Vmax of the galactose uptake system was shown to be dilution rate (D) dependent, comparable with the high‐affinity glucose uptake. The maximum uptake was observed at D 0·2 h−1 (0·25 mmol min−1 per g) and the minimum uptake (0·1 mmol min−1 per g) at D 0·05 h−1 and 0·3 h−1. The aerobic fermentation of galactose occurred at D 0·275–0·3 h−1 which is identical to the results obtained in glucose‐limited continuous cultures of this strain. Because galactose is not a repressing carbon source, this demonstrates that the Crabtree effect is not mediated by, or in any way related to glucose repression. Moreover, invertase and hexokinase I mRNA levels (both subject to glucose repression at the transcriptional level) were present when the yeast produced ethanol in galactose‐ and glucose‐limited continuous cultures. In glucose‐limited continuous cultures a decrease in alcohol dehydrogenase (I and II) mRNA levels and activity and phosphoglucomutase activity was observed with increasing dilution rates. In addition, at D 0·3 h−1, when the yeast produced ethanol, glucose‐6‐phosphate dehydrogenase and pyruvate decarboxylase were induced and a decrease in respiration was observed. The fact that the same changes were seen in a galactose‐limited culture and that invertase and hexokinase I mRNA levels were present demonstrates that glucose is not specifically involved in the regulation of these enzymes. The changes observed are therefore caused by the growth rate of the organism, the glycolytic flux (e.g. alcohol dehydrogenase) or the overflow metabolism when the yeast produces ethanol (e.g. pyruvate decarboxylase).
A 12·8 kb segment, on the right arm of chromosome II from Saccharomyces cerevisiae including part of the DUR1, 2 gene, contains five putative new genes
Authors: Bussereau Françoise , Mallet Laurent , Jacquet Michel , Gaillon Laurent 
Published: May 2005

AbstractA 12 820 bp fragment from the right arm of chromosome II of Saccharomyces cerevisiae was sequenced and analysed. This fragment contains six non‐overlapping long open reading frames (ORFs) designated from the centromere‐ to the telomere‐proximal ends as: YBR1441, 1443, 1444, 1445, 1446 and 1448. YBR1441 encodes a polypeptide of 845 amino acids which shares a long consensus domain with products of S. cerevisiae MCM2, MCM3, CDC46 and Schizosaccharomyces pombe cdc21+ genes. These genes are involved in DNA replication. YBR1445 encodes a polypeptide of 404 amino acids which has strong similarity with the S. cerevisiae KRE2/MNT1, YUR1, KTR1 gene products. The KRE2/MNT1 protein is an α‐1,2‐mannosyltransferase. The product of YBR1444, which encodes a protein of 375 amino acids, presents a lipase signature sequence and a peroxisomal targeting signal. YBR1448, whose sequence extends further on the telomere‐proximal end of the fragment, is identical to the 3′ end of the DUR1,2 gene encoding urea amidolyase. The two ORFs, YBR1443 and YBR1446, exhibit no significant similarity with any known gene.
Analysis of the inducer‐responsive CAR1 upstream activation sequence (UASI) and the factors required for its operation
Authors: Kovari Ladislau Z. , Park Heui‐Dong , Kovari Iulia A. , Cooper Terrance G. , Fourie Marlene , van Vuuren Hendrik J. J. 
Published: May 2005

AbstractInduced production of arginase (CAR1) enzyme activity and steady‐state CAR1 mRNA in Saccharomyces cerevisiae requires wild‐type ARG80/ARGRI and ARG81/ARGRII gene products. We demonstrate here that these gene products, along with that of the MCM1 gene, are required for the inducer‐dependent UASI‐A, UASI‐B and UASI‐C elements to function but they are not required for operation of inducer‐independent CAR1 UASC1 or UASC2M. Through the use of single and multiple point mutations, the CAR1 UASI‐B and UASI‐C elements were demonstrated to be at least 23 bp in length. Moreover, simultaneous mutation of both ends of an elements gave stronger phenotypes than mutations at either end. The center of the element was more sensitive to mutation than were the ends.
Sequence and function analysis of a 4·3 kb fragment of Saccharomyces cerevisiae chromosome II including three open reading frames
Authors: Schaaff‐Gerstenschläger Ine , Baur Axel , Boles Eckhard , Zimmermann Friedrich K. 
Published: May 2005

AbstractThe nucleotide sequence of a fragment of 4337 base pairs of Saccharomyces cerevisiae chromosome II has been determined. The sequence contains three open reading frames, one of them being incomplete. Deletion analysis showed that YBR12.31 is essential for yeast growth, while deletion mutants of YBR12.32 and YBR12.33 are viable. YBR12.33 is identical to SMY2, isolated as a suppressor of a myo2 mutant (Lillie, S.H. and Brown, S.S., unpublished, EMBL M90654).
Two distinct yeast proteins are related to the mammalian ribosomal polypeptide L7
Authors: Lalo Dominique , Mariotte Sylvie , Thuriaux Pierre 
Published: May 2005

AbstractThe RLP7 gene of Saccharomyces cerevisiae was cloned, sequenced and localized to the right arm of chromosome XIV, close to the centromere. It encodes a predicted polypeptide (RLP7p) of 322 amino acids, with a calculated molecular mass of 36 kDa and an isoelectric point of 9·6. Putative open reading frames very similar to RLP7 are present in two other yeasts, Kluyveromyces lactis and Candida utilis. The RLP7p gene product has significant sequence similarity to the S. cerevisiae YL8 polypeptide of the large ribosomal subunit (Mizuta et al., 1992), itself homologous to the L7 subunit of mammalian ribosomes. However, RLP7p and YL8 do not functionally replace each other, since an rlp7‐Δ::HIS3 strain is completely inviable. Judging from its predicted mass, isoelectric point and amino acid sequence, RLP7p does not correspond to any ribosomal component biochemically identified so far in S. cerevisiae, and also differs from all known ribosomal proteins by the low codon usage bias of its gene.
DNA sequence analysis of a 17 kb fragment of yeast chromosome XI physically localizes the MRB1 gene and reveals eight new open reading frames, including a homologue of the KIN1/KIN2 and SNF1 protein kinases
Authors: Pallier Chantal , Valens Michèle , Puzos Valérie , Fukuhara Hiroshi , Chéret Geneviève , Sor Frédéric , Bolotin‐Fukuhara Monique 
Published: May 2005

AbstractWe report in this paper the sequence of a part of chromosome XI of Saccharomyces cerevisiae. This 17 kbp nucleotide sequence represents the right half of cosmid pUKG151 and contains nine open reading frames, YKL453, 450, 449, 448, 445, 443, 442, 441 and the 5′ part of YKL440. YKL440 was previously identified as the MBR1 gene and plays a role in mitochondrial biogenesis. YKL443 is a homologue of the yeast serine‐rich protein (SRP1), while YKL453 presents strong homologies with the KIN1/KIN2/SNF1 kinase family. It must be pointed out that the size of this gene is well above average for yeast.
Ribosome synthesis during the growth cycle of Saccharomyces cerevisiae
Authors: Ju Qida , Warner Jonathan R. 
Published: May 2005

AbstractWe have measured the content of ribosomes, the rate of synthesis of ribosomal RNA, and the level of the mRNA for ribosomal proteins as a culture of Saccharomyces cerevisiae passes through the growth cycle. The transcription of both ribosomal RNA and ribosomal protein genes disappears at an unexpectedly early stage in the growth cycle, accompanied by a decline in the total RNA content of the culture by nearly 50% and a decline in the number of ribosomes per cell to less than 25% of the maximum value. During this time the cells continue to grow through more than two doublings, initially at the normal log growth rate, which then decline gradually for several hours. The data suggest that the cell can sense an unfavorable change within the medium and responds by employing regulation of both synthesis and degradation of its ribosomes. We conclude that the cell regulates ribosome synthesis and content according to its estimate of the potential for growth.
XI. Yeast sequencing reports. The complete sequence of an 18,002 bp segment of Saccharomyces cerevisiae chromosome XI contains the HBS1, MRP‐L20 and PRP16 genes, and six new open reading frames
Authors: García‐Cantalejo Jesús , Baladrón Victoriano , Esteban Pedro F. , Santos M. Angeles , Bou Germán , Remacha Miguel A. , Revuelta José L. , Ballesta Juan P. G. , Jiménez Antonio , Del Rey Francisco 
Published: May 2005

AbstractWe report the sequence of an 18,002 bp DNA fragment from the right arm of Saccharomyces cerevisiae chromosome XI. This segment contains nine complete open reading frames (ORFs), YKR401 to YKR409, and part of another ORF, YKR400, covering altogether 87·2% of the entire sequence. One of them, YKR400, encodes an NAD‐dependent 5,10‐methylene‐tetrahydrofolate dehydrogenase. YKR404, YKR405 and YKR406 correspond to the previously characterized HBS1, MRP‐L20 and PRP16 genes, coding for a translation elongation factor, a mitochondrial ribosomal protein and an ATP‐binding protein, respectively. The putative product of YKR407 contains the zinc‐binding region signature of neutral zinc metallopeptidases. The five other ORFs do not show significant homology to any known protein. The sequence data reported here have been assigned EMBL accession number Z27116.
III. Yeast sequencing reports. Corrected sequence for the right telomere of Saccharomyces cerevisiae chromosome III
Authors: Louis Edward J. 
Published: May 2005

AbstractA comparison of the sequences of telomere regions from several yeast chromosomes revealed an apparent cloning artifact for the right end of chromosome III. An integrating vector containing G1–3T telomere sequences was used to clone the right end of chromosome III from a strain related to S288C. The sequence of this clone confirmed that the published sequence was incorrect and demonstrated that the right telomere region of chromosome III is similar to other telomeres.
XIII. Yeast sequencing reports. Cloning and sequencing of the NES24 gene of Saccharomyces cerevisiae
Authors: Yoshida Miho , Shimma Yoh‐Ichi , Uno Isao , Toh‐E Akio 
Published: May 2005

AbstractWe have cloned NES24 using a temperature‐sensitive nes24‐1 mutant as a host and sequenced a 3162 bp XhoI‐EcoRI DNA fragment containing the NES24 gene. Computer analysis revealed that this segment contains a 1806 bp open reading frame which is needed for complementation of the nes24‐1 mutation. We found SUP8 in the region upstream of the NES24 gene, placing the NES24 gene on chromosome XIII. A protein homology search indicated that NES24 encodes a new protein. The disruption of the NES24 gene resulted in temperature‐sensitive growth. The sequence has been deposited in DDBJ/EmBL/GenBank data bases under Accession Number D15052.
Vectors for Cu2+‐inducible production of glutathione S‐transferase‐fusion proteins for single‐step purification from yeast
Authors: Ward Alister C. , Castelli Laura A. , Macreadie Ian G. , Azad Ahmed A. 
Published: May 2005

AbstractWe describe six new yeast episomal vectors which encode glutathione S‐transferase (GST) affinity tags. These allow for the production of GST‐fusion proteins in Saccharomyces cerevisiae under the control of the CUP1 promoter. Affinity chromatography with glutathione‐Sepharose permits convenient purification of the fusion protein from a yeast lysate. The presence of a protease cleavage site facilitates subsequent removal of the GST tag. The expression and single‐step purification of both GST and a functional GST‐metallothionein fusion from yeast are shown as an example of the application of these vectors.
Molecular analysis of the malic enzyme gene (mae2) of Schizosaccharomyces pombe
Authors: Viljoen Marinda , Subden Ron E. , Krizus Aldis , van Vuuren Hendrik J. J. 
Published: May 2005

AbstractSequence analysis of a 4·6‐kb HindIII fragment containing the malic enzyme gene (mae2) of Schizosaccharomyces pombe, revealed the presence of an open reading frame of 1695 nucleotides, coding for a 565 amino acid polypeptide. The mae2 gene is expressed constitutively and encodes a single mRNA transcript of 2·0 kb. The mae2 gene was mapped on chromosome III by chromoblotting. The coding region and inferred amino acid sequence showed significant homology with 12 malic enzyme genes and proteins from widely different origins. Eight highly homologous regions were found in these malic enzymes, suggesting that they contain functionally conserved amino acid sequences that are indispensable for activity of malic enzymes. Two of these regions have previously been reported to be NAD‐ and NADP‐binding sites.
XI. Yeast sequencing reports. The complete sequencing of a 24·6 kb segment of yeast chromosome XI identified the known loci URA1, SAC1 and TRP3, and revealed 6 new open reading frames including homologues to the threonine dehydratases, membrane transporters, hydantoinases and the phospholipase A2‐activating protein
Authors: Tzermia Maria , Horaitis Ourania , Alexandraki Despina 
Published: May 2005

AbstractWe report the entire sequence of a 26·4 kb segment of chromosome XI of Saccharomyces cerevisiae. Identification of the known loci URA1, TRP3 and SAC1 revealed a translocation compared to the genetic map. Additionally, six unknown open reading frames have been identified. One of them is similar to catabolic threonine dehydratases. Another one contains characteristic features of membrane transporters. A third one is homologous in half of its length to the prokaryotic hydantoinase HyuA and in the other half to hydatoinase HyuB. A fourth one is homologous to the mammalian phospholipase A2‐activating protein. A fifth one, finally, is homologous to the hypothetical open reading frame YCR007C of chromosome III. The sequence has been deposited in the EMBL data library under Accession Number X75951.
VIII. Yeast mapping reports. Genetic mapping of STE20 to the left arm of chromosome VIII
Authors: Horecka Joe , Sprague George F. 
Published: May 2005

AbstractSTE20 is a newly‐discovered element of the Saccharomyces cerevisiae pheromone response pathway. We have isolated a recessive ste20 mutation and have used it to map the gene to the left arm of chromosome VIII, establishing the gene order STE20‐CEN8‐GPA1‐ARG4.
A new nuclear suppressor system for a mitochondrial RNA polymerase mutant identifies an unusual zinc‐finger protein and a polyglutamine domain protein in Saccharomyces cerevisiae
Authors: Bröhl Stefanie , Lisowsky Thomas , Riemen Gudula , Michaelis Georg 
Published: May 2005

AbstractA yeast strain with a point mutation in the nuclear gene for the core subunit of mitochondrial RNA polymerase was used to isolate new extragenic suppressors. Spontaneously occurring phenotypical revertants were analysed by crosses with the wild‐type and tetrad dissection. One of the new nuclear suppressor mutants was characterized by temperature‐sensitive growth on non‐fermentable carbon sources. This mutant was transformed with a genomic yeast library. Two independent types of DNA clones were isolated which both complemented the temperature‐sensitive defect. Subcloning and DNA sequencing identified two novel yeast genes which code for proteins with the characteristic features of transcription factors. Both factors exhibit highly structured protein domains consisting of runs and clusters of asparagine and glutamine residues. One of the proteins contains in addition zinc‐finger domains of the C2H2‐type. Therefore the genes are proposed to be named AZF1 (asparagine‐rich zinc‐ffinger protein) and PGD1 (polyglutamine domain protein). Gene disruption of both reading frames has no detectable influence on the vegetative growth on complete glucose or glycerol media, indicating that the genes may act as high copy number suppressors of the mutant defect. Additional transformation experiments showed that AZF1 is also an efficient suppressor for the original defect in the core subunit of mitochondrial RNA polymerase. The DNA sequences for the AZF1 and PGD1 genes were submitted to the EMBL data base (Accession Numbers: Z26253 and Z26254).
II. Yeast sequencing reports. Nucleotide sequence analysis of an 11·7 kb fragment of yeast chromosome II including BEM1, a new gene of the WD‐40 repeat family and a new member of the KRE2/MNT1 family
Authors: Mallet Laurent , Bussereau Françloise , Jacquet Michel 
Published: May 2005

AbstractThis paper reports the DNA sequence and analysis of an 11·7 kb segment localized on the right arm of Saccharomyces cerevisiae chromosome II. This fragment contains one incomplete and five long and non‐overlapping open reading frames (ORFs) designated from centromere to telomere‐proximal side as: YBR1406, 1409, 1410, 1411, 1412 and 1413. YBR1406 corresponds to the 5′ end to PGI1 encoding phosphoglucoisomerase. YBR1410 encodes a polypeptide of 798 amino acids whose C terminus contains five repeats (WD‐40 repeat) similar to those found in the β‐subunits of G proteins and different yeast proteins such as Tup1, Prp4 and Cdc4. The higher similarity score is obtained with dTAFII80, a component of the RNA polymerase II transcriptional complex TFIID. YBR1411 encodes a polypeptide of 464 amino acids which belongs to the family of α‐mannosyltransferases: KRE2/MNT1, KTR1, KTR2, YUR1 and the product of previously sequenced ORF YBR1445. YBR1412 corresponds to BEM1. The two ORFs, YBR1409 and YBR1413, which do not exhibit significant similarity with any known coding sequences, define new genes. The sequence has been deposited in the EMBL Data Library under Accession Number Z21487.
Genetic interrelationship among species of the genus Zygosaccharomyces as revealed by small‐subunit rRNA gene sequences
Authors: James S. A. , Collins M. D. , Roberts I. N. 
Published: May 2005

AbstractThe phylogenetic interrelationship of species of the genus Zygosaccharomyces was examined by 18S rRNA gene sequencing. Comparative analysis of the sequence data revealed the genus to consist of a number of distinct subdivisions. The most prevalent species associated with food spoilage, Z. bailii, Z. bisporus and Z. rouxii, along with Z. mellis were found to form one subdivision. Zygosaccharomyces cidri and Z. fermentati formed a distinct species pair, as did Z. microellipsoides and Z. mrakii. Zygosaccharomyces florentinus formed a separate line displaying no specific relationship with any of the other Zygosaccharomyces species examined. Comparison with nine published ascosporogenous yeast 18S rRNA gene sequences showed that Z. microellipsoides and Z. mrakii were genealogically very close to Torulaspora delbrueckii (both displaying 99·8% 18S rRNA sequence similarity), raising the possibility that these two Zygosaccharomyces species should be moved to the genus Torulaspora.The topologies of trees derived from complete 18S rRNA gene sequences and from individual domains within the gene were compared and the implications of using partial sequence data for inferring phylogenetic relationships discussed.
X. Yeast sequencing reports. Sequence and function analysis of a 9·46 kb fragment of Saccharomyces cerevisiae chromosome X
Authors: Miosga Thomas , Witzel Andrea , Zimmermann Friedrich K. 
Published: May 2005

AbstractIn the framework of the European yeast genome sequencing project, we have determined the nucleotide sequence of the cosmid clone 233 provided by F. Galibert (Rennes Cedex, France). We present here 9464 base pairs of this cosmid located on the left arm of Saccharomyces cerevisiae chromosome X. This sequence contains two new open reading frames and includes the published sequences of the RADH gene (also identified as SRS2/HPR5) and the 3′‐end of the gene BCK1/SLK1/SSP31. Deletion mutants of the two unknown genes J0909 and J0911 are viable. The sequence has been deposited in the EMBL data library under accession number X77087.
A new approach for isolating cell wall mutants in Saccharomyces cerevisiae by screening for hypersensitivity to calcofluor white
Authors: Ram Arthur F. J. , Wolters Aly , Hoopen Rogier Ten , Klis Frans M. 
Published: May 2005

AbstractTo study cell wall assembly, a simple screening method was devised for isolating cell wall mutants. Mutagenized cells were screened for hypersensitivity to Calcofluor White, which interferes with cell wall assembly. The rationale is that Calcofluor White amplifies the effect of cell wall mutations. As a result, the cells stop growing at lower concentrations of Calcofluor White than cells with normal cell wall. In this way, 63 Calcofluor White‐hypersensitive (cwh), monogenic mutants were obtained, ordered into 53 complementation groups.The mannose/glucose ratios of the mutant cell walls varied from 0.15 to 3.95, while wild‐type cell walls contained about equal amounts of mannose and glucose. This indicates that both low‐mannose and low‐glucose cell wall mutants had been obtained. Further characterization showed the presence of three low‐mannose cell wall mutants with a mnn9‐like phenotype, affected, however, in different genes. In addition, four new killer‐resistant (kre) mutants were found, which are presumably affected in the synthesis of β1,6‐glucan. Most low‐glucose cell wall mutants were not killer resistant, indicating that they might be defective in the synthesis of β1,3‐glucan. Eleven cwh mutants were found to be hypersensitive to papulacandin B, which is known to interfere with β1,3‐glucan synthesis, and four cwh mutants were temperature‐sensitive and lysed at the restrictive temperature. Finally, nine cwh mutants were hypersensitive to caffeine, suggesting that these were affected in signal transduction related to cell wall assembly.
II. Yeast sequencing reports. The two genes encoding yeast ribosomal protein S8 reside on different chromosomes, and are closely linked to the hsp70 stress protein genes SSA3 and SSA4
Authors: Logghe Marc , Molemans Francis , Fiers Walter , Contreras Roland 
Published: May 2005

AbstractA 7·4 kb segment of chromosome II was sequenced and analysed. This segment is part of the 25 kb insert of cosmid clone α1004.10 which is located on the left arm of chromosome II. Sequence analysis revealed four open reading frames (ORFs), of which two had been characterized previously (SSA3, AAR2) and one was not identified. The other ORF was precisely 600 bp long and the deduced protein sequence predicted a very basic protein (pI=11·1; molecular weight=22·5 kDa). Evidence was found that the ORF is the S40 ribosomal protein gene (RPG) S8. Consensus splice signals were found in the 5′ leader sequence and also potential RPG‐specific sequences. Chromoblot analysis revealed a second copy of the S8 RPG on chromosome IV or VIII. This copy is also closely linked to an hsp70 protein gene, SSA4. The sequence has been deposited in the EMBL data library under accession number Z26879.