Nucleotide sequence of the ADE 1 gene of the yeast Saccharomyces cerevisiae

The yeast ADE 1 gene has been cloned and sequenced. The primary structure deduced from the nucleotide sequence demonstrated that phosphoribosylaminoimidazole-succinocarboxamide synthetase is a protein with molecular weight of 34 500 D.     

Nucleotide sequence of the Saccharomyces cerevisiae CTT1 gene and deduced amino-acid sequence of yeast catalase T

A 2642-base-pair DNA fragment containing the catalase T (CTT1) structural gene of the yeast Saccharomyces cerevisiae and its flanking regions has been sequenced. The gene codes for a protein of 562 amino acids (relative molecular mass 64,449) and appears to contain no intron. The amino acid sequence of catalase T derived from the DNA sequence shows 40.7% homology (52.2% including conservative replacements) to that of bovine liver catalase. All amino acids previously postulated to participate directly in catalysis by liver catalase and most of the amino acids of the immediate environment of hemin, the prosthetic group of catalase, are conserved in catalase T. The data obtained indicate that the folding of polypeptide chains of the two catalases compared has been conserved within a central region consisting mainly of the beta-barrel domain, which bears the prosthetic group, and a major part of the "wrapping domain". N- and C-terminal regions involved in subunit interactions are less well conserved. It is suggested that their structure is more similar to that of the corresponding regions of Penicillium vitale catalase. However, catalase T lacks the C-terminal flavodoxin-like domain present in this protein.     

Nucleotide sequence of the dihydrofolate reductase gene of Saccharomyces cerevisiae

The nucleotide sequence of a DNA fragment that contained the Saccharomyces cerevisiae gene DFR coding for dihydrofolate reductase (DHFR) was determined. The DHFR was encoded by a 633-bp open reading frame, which specified an Mr24264 protein. The polypeptide was significantly related to the DHFRs of chicken liver and Escherichia coli. The yeast enzyme shared 60 amino acid (aa) residues with the avian enzyme and 51 aa residues with the bacterial enzyme. DHFR was overproduced about 40-fold in S. cerevisiae when the cloned gene was present in the vector YEp24. As isolated from the Saccharomyces library, the DFR gene was not expressed in E. coli. When the gene was present on a 1.8-kb BamHI-SalI fragment subcloned into the E. coli vector, pUC18, weak expression in E. coli was observed.     

Nucleotide sequence of AMS1, the structure gene of vacuolar alpha-mannosidase of Saccharomyces cerevisiae

AMS1, a structure gene of the vacuolar membrane alpha-mannosidase of Saccharomyces cerevisiae, has been characterized and found to encode both constituent polypeptides of the enzyme, a 107 kDa polypeptide and a 73 kDa polypeptide. The nucleotide sequence of AMS1 demonstrates that the gene encodes 1083 amino acids with a molecular weight 124,497. Although the enzyme is considered to exist on the inner surface of the vacuolar membrane, the predicted primary amino acid sequence does not have a hydrophobic stretch suitable for a signal sequence in its N-terminal region.     

Nucleotide sequence and functional analysis of the RAD1 gene of Saccharomyces cerevisiae.

The RAD1 gene of Saccharomyces cerevisiae is involved in excision repair of damaged DNA. The nucleotide sequence of the RAD1 gene presented here shows an open reading frame of 3,300 nucleotides. Two ATG codons occur in the open reading frame at positions +1 and +334, respectively. Since a deletion of about 2.7 kilobases of DNA from the 5' region of the RAD1 gene, which also deletes the +1 ATG and 11 additional codons in the RAD1 open reading frame, partially complements UV sensitivity of a rad1 delta mutant, we examined the role of the +1 ATG and +334 ATG codons in translation initiation of RAD1 protein. Mutation of the +1 ATG codon to ATC affected the complementation ability of the RAD1 gene, whereas mutation of the +334 ATG codon to ATC showed no discernible effect on RAD1 function. These results indicate that translation of RAD1 protein is initiated from the +1 ATG codon. Productive in-frame RAD1-lacZ fusions showed that the RAD1 open reading frame is expressed in yeasts. The RAD1-encoded protein contains 1,100 amino acids with a molecular weight of 126,360.     

Nucleotide sequence of the RAD10 gene of Saccharomyces cerevisiae.

The RAD10 gene is one of several genes in Saccharomyces cerevisiae required for incision of u.v.-irradiated or cross-linked DNA. We have determined the nucleotide sequence of the RAD10 gene and its flanking regions. The RAD10 nucleotide sequence presented here differs significantly from that recently reported. The RAD10 protein predicted from the nucleotide sequence contains 210 amino acids with a calculated mol. wt. of 24 310. The middle portion of the RAD10 protein, which is highly basic and also contains eight of the total of 10 tyrosine residues present in the protein, may be involved in DNA binding by ionic interactions and tyrosine intercalation between the bases of DNA. A genomic deletion of the entire RAD10 gene does not affect viability; however, the rad10 deletion mutant is highly u.v. sensitive.     

Nucleotide sequence of two rasH related-genes isolated from the yeast Saccharomyces cerevisiae.

A complete nucleotide sequence of two ras-related yeast genes (c- rassc -1 and c- rassc -2) isolated from the yeast strain Saccharomyces cerevisiae is reported. They encode predicted polypeptides of 40,000 and 41,000 daltons, respectively. The N-terminal 170 amino acids from both genes show extensive amino acid homology to other ras genes from vertebrates, whereas their C-termini have diverged. These genes should be useful in the elucidation of a normal biological function of ras-related genes in a simple system like yeast.     

Post-secretional modification of exo-1,3-beta-D-glucanases from Saccharomyces cerevisiae.

Exo-1,3-beta-D-glucanase secreted by Saccharomyces cerevisiae undergoes extracellular modifications in its carbohydrate moiety that change the affinity towards the lectin concanavalin A. The transition of negatively reacting enzyme form into positively reacting one depends on temperature. Results from experiments with glucono-delta-lactone and from treatments in vitro with hydrolases suggest a glycosidase-mediated mechanism.     

Nucleotide sequence of the extracellular glucoamylase gene STA1 in the yeast Saccharomyces diastaticus.

The complete nucleotide sequence of the extracellular glucoamylase gene STA1 from the yeast Saccharomyces diastaticus has been determined. A single open reading frame codes for a 778-amino-acid protein which contains 13 potential N-glycosylation sites. In the 5'- and 3'-flanking regions of the gene, there are striking sequence homologies to the corresponding regions of ADH1 for alcohol dehydrogenase and MAT alpha 2 for mating type control in the yeast Saccharomyces cerevisiae. The putative precursor begins with a hydrophobic segment that presumably acts as a signal sequence for secretion. The presumptive signal sequence showed a significant homology to that of Bacillus subtilis alpha-amylase precursor. The next segment, of ca. 320 amino acids, contains a threonine-rich tract in which direct repeat sequences of 35 amino acids exist, and is bordered by a pair of basic amino acid residues (Lys-Lys) which may be a proteolytic processing signal. The carboxy-terminal half of the precursor is a presumptive glucoamylase which contains several peptide segments showing a high degree of homology with alpha-amylases from widely diverse organisms including a procaryote (B. subtilis) and eucaryotes (Aspergillus oryzae and mouse). Analysis of both the nucleotide sequence of the STA1 gene and the amino acid composition of the purified glucoamylase suggested that the putative precursor is processed to yield subunits H and Y of mature enzyme by both trypsin-like and chymotrypsin-like cleavages.     

The Saccharomyces cerevisiae SPR1 gene encodes a sporulation-specific exo-1,3-beta-glucanase which contributes to ascospore thermoresistance.

A number of genes have been shown to be transcribed specifically during sporulation in Saccharomyces cerevisiae, yet their developmental function is unknown. The SPR1 gene is transcribed during only the late stages of sporulation. We have sequenced the SPR1 gene and found that it has extensive DNA and protein sequence homology to the S. cerevisiae EXG1 gene which encodes an exo-1,3-beta-glucanase expressed during vegetative growth (C. R. Vasquez de Aldana, J. Correa, P. San Segundo, A. Bueno, A. R. Nebrada, E. Mendez, and F. del Ray, Gene 97:173-182, 1991). We show that spr1 mutant cells do not hydrolyze p-nitrophenyl-beta-D-glucoside or laminarin in a whole-cell assay for exo-1,3-beta-glucanases. In addition to the absence of this enzymatic activity, spr1 mutant spores exhibit reduced thermoresistance relative to isogenic wild-type spores. These observations are consistent with the notion that SPR1 encodes a sporulation-specific exo-1,3-beta-glucanase.     

Nucleotide sequence of the CLS4 (CDC24) gene of Saccharomyces cerevisiae

The nucleotide sequence of the CLS4 gene controlling Ca2+ regulatory process of bud emergence, which was cloned previously [Ohya et al., J. Bacteriol. 165 (1986) 28-33], was determined. The CLS4 (CDC24) locus encodes a protein consisting of 736 amino acid (aa) residues with an Mr of 83,970. By primer extension mapping, the mRNA start point was located 139 bp upstream from the translation start codon. The predicted CLS4 protein was hydrophilic with two serine + threonine-rich domains in the middle and C-terminal regions. It has two putative Ca2+-binding regions, one being partly homologous to the Ca2+-binding domain of the S-100a protein and the other that of alpha-lactalbumin.