Sequence of a 4·8 kb fragment of Saccharomyces cerevisiae chromosome II including three essential open reading frames

The nucleotide sequence of a fragment of 4867 base pairs of Saccharomyces cerevisiae chromosome II has been determined. The sequence contains three complete open reading frames. In addition to the already known gene RPB5, coding for a subunit shared by all three DNA directed RNA polymerases, two new open reading frames could be identified. YBR12.03 codes for a protein of 183 amino acids with homology to one of the proteins of the Bacillus subtilis riboflavin biosynthesis operon (RibG). Deletion mutants of YBR12.03 can germinate but stop growing after five to seven cell divisions on YPD. Supplementation with high concentrations of riboflavin does promote growth. YBR12.05 codes for a protein of 386 amino acids with homology to STI1, a stress-inducible protein of S. cerevisiae. Deletion mutants of YBR12.05 are not viable.     

Galactose-inducible Expression Systems in Candida maltosa using Promoters of Newly-isolated GAL1 and GAL10 Genes

The GAL1 and GAL10 gene cluster encoding the enzymes of galactose utilization was isolated from an asporogenic yeast, Candida maltosa. The structure of the gene cluster in which both genes were divergently transcribed from the central promoter region resembled those of some other yeasts. The expression of both genes was strongly induced by galactose and repressed by glucose in the medium. Galactose-inducible expression vectors in C. maltosa were constructed on low- and high-copy number plasmids using the promoter regions of both genes. With these vectors and the β-galactosidase gene from Kluyveromyces lactis as a reporter, galactose-inducible expression was confirmed. Homologous overexpression of members of the cytochrome P-450 gene family in C. maltosa was also successful by using a high-copy-number vector under the control of these promoters. © 1997 by John Wiley & Sons, Ltd.     

Sequence and function analysis of a 2·73 kb fragment of Saccharomyces cerevisiae chromosome II

The nucleotide sequence of a fragment of 2728 base pairs of Saccharomyces cerevisiae chromosome II has been determined. The sequence contains two open reading frames, one of them being incomplete. Deletion mutants of YBR11.21 are viable. YBR11.20 is identical to the recessive omnipotent suppressor SUP45 (SUP1).     

Sequence and function analysis of a 4·3 kb fragment of Saccharomyces cerevisiae chromosome II including three open reading frames

The 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).     

Homolog of a-factor receptor gene in Saccharomyces exiguus

Homologs of Saccharomyces cerevisiae STE3, a -factor receptor gene were detected from S. exiguus NFRI 3539 by low stringency Southern hybridization. This strain might have at least two types of homolog. One of these homologs, designated as e-STE3 was cloned. Its nucleotide sequence revealed 60% identity to STE3. The putative protein coding region consisted of 453 amino acid residues. The amino acid sequence identity between STE3 and e-STE3 was 62%, and that of the N-terminal 303 amino acid residues considered to be the pheromone binding domain was 79%. The e-STE3 sequence submitted to the DDBJ/EMBL/GenBank data libraries is available under Accession Number AB003086. © 1998 John Wiley & Sons, Ltd.     

II. Yeast sequencing reports. Sequence analysis of a 31 kb DNA fragment from the right arm of Saccharomyces cerevisiae chromosome II

The nucleotide sequence of a 31 352 bp fragment from chromosome II of Saccharomyces cerevisiae has been determined and analysed. The fragment originates from the right arm of chromosome II, located between the GAL7,10,1 and the PHO3,5 loci, at a distance of about 130 kb from the centromere. The sequence contains a tRNA tandem repeat and 17 open reading frames (ORFs) larger than 100 amino acids. One of them extends into adjacent DNA and is incomplete. The two tRNA genes, coding for a tRNA^asp and a tRNA^arg, and three of the ORFs, had been sequenced previously, i.e. HSP26, SEC18, and UBC4. Four other ORFs showed similarity with yeast genes; amino acid transporter genes, the RAD54, SNF2 and STH1 family, the SPS2 gene and the bromodomain of SPT7, respectively. Two showed homology with sequences from other organisms, i.e. with a Plasmodium falciparum gene encoding a surface antigen and with a gene from Saimirine herpes virus respectively. Three ORFs, YBR0726, YBR0735 and YBR0740 are completely contained in YBR0727, YBR0734 and YBR0739 respectively, and thus probably do not represent real genes. Two ORFs, YBR0727 and YBR0745 most likely contain an intron. The sequences have been deposited in the EMBL data library under Accession Number X76294.     

The nucleotide sequence of a 2·1 kb fragment from chromosome VI of Saccharomyces cerevisiae identifies a tRNAGly gene,part of a delta element and a palindromic sequence

The nucleotide sequence was determined of a 2·1 kb DNA fragment located at approximately 35 kb to the right of the centromere of chromosome VI from Saccharomyces cerevisiae. Analysis revealed the presence of a tRNA^GLy gene, part of a delta element and a remarkable palindromic sequence. The longest open reading frame found encodes a putative protein of 195 amino acids. Although the fragment was isolated by hybridization to a human diacylglycerol kinase cDNA, no evidence was obtained for the presence of a gene encoding diacylglycerol kinase.     

The Genes Encoding the Transcription Factor yTAFII60, the G4p1 Protein and a Putative Glucose Transporter are Contained in a 12·3 kb DNA Fragment on the Left Arm of Saccharomyces cerevisiae Chromosome VII

We report the nucleotide sequence of a DNA fragment of 12 325 base pairs from the left arm of the Saccharomyces cerevisiae chromosome VII. Inspection of the coding capacity revealed 11 open reading frames (ORFs) longer than 100 amino acids. Five ORFs are significantly homologous to known proteins. The region encoding ORF G2985 corresponds (100%) to the gene encoding the yeast TATA binding protein-associated factor TAF_II60. The G3075 ORF is 47·8% identical to the hypothetical yeast protein YB88. G3080 shows 36·7% identity to the eel calmodulin. G3085 shows 94·9% identity with the published sequence of the quadruplex DNA binding protein G4p1. G3090 reveals 46·7% identity with the probable glucose transport protein yBR1625. The DNA sequence has been submitted to the EMBL data library under Accession Number X97644. © 1997 by John Wiley & Sons, Ltd.     

The sequence of a 17 933 bp segment of Saccharomyces cerevisiae chromosome XIV contains the RHO2, TOP2, MKT1 and END3 genes and five new open reading frames

We report the DNA sequence of a 17 933 bp fragment from the left arm of chromosome XIV of Saccharomyces cerevisiae. Analysis of the sequence reveals the presence of ten open reading frames (ORFs) larger than 100 codons. Four of these were previously identified as genes RHO2, TOP2, MKT1 and END3. Additionally, the NH₂ end coding region of PMS1 is found in the 3′ end of the sequence. No significant homology to any known protein has been found for the other five ORFs. The nucleotide sequence has been deposited at EMBL, with Accession Number X89016.     

Candida maltosa NADPH-cytochrome P450 reductase: Cloning of a full-length cDNA,Heterologous expression in Saccharomyces cerevisiae and function of the N-terminal region for membrane anchoring and proliferation of the endoplasmic reticulum

A full-length cDNA for NADPH-cytochrome P450 reductase from Candida maltosa was cloned and sequenced. The derived amino acid sequence showed a high similarity to the reductases from other eukaryotes. Expression in Saccharomyces cerevisiae under control of the GAL10 promoter resulted in an approximately 70-fold increase in NADPH-cytochrome c reductase activity in the microsomal fraction. The functional integrity of the heterologously expressed reductase as an electron transfer component for alkane hydroxylating cytochrome P450 from C. maltosa was shown in a reconstituted system containing both enzymes in a highly purified state. The signal-anchor sequence of the reductase was identified within the N-terminal region of the protein by means of constructing and expressing fusion proteins with the cytosolic form of yeast invertase. The first 33 amino acids turned out to be sufficient for stable membrane insertion, wild-type membrane orientation and retention in the endoplasmic reticulum. As shown by immunoelectron microscopy, the heterologously expressed reductase was integrated into the endoplasmic reticulum of the host organism. It triggered a strong proliferation of the membrane system. This membrane-inducing property of the reductase was transferable to the cytosolic reporter protein with the same N-terminal sequences that confer membrane insertion. The nucleotide sequence of the cDNA of NADPH-cytochrome P450 reductase from C. maltosa is available from the EMBL data library under Accession Number X76226.     

IV. Yeast sequencing reports. New open reading frames,one of which is similar to the nifV gene of Azotobacter vinelandii,Found on a 12·5 kbp fragment of chromosome IV of Saccharomyces cerevisiae

The nucleotide sequence of a 12·5 kbp segment of the left arm of chromosome IV is described. Five open reading frames (ORFs) longer than 100 amino acids were detected, all of which are completely confined to the 12·5 kbp region. Two ORFs (D1271 and D1286) correspond to previously sequenced genes (PPH22 and VMA1 or TFP1, respectively). ORF D1298 shows the characteristics of α-isopropylmalate and homocitrate synthase genes and is similar to the nifV gene of Azotobacter vinelandii. Two more ORFs have no apparent homologue in the data libraries. Conversely, two smaller ORFs of 25 and 85 amino acids encoding the ribosomal protein YL41A and an ATPase inhibitor, respectively, were detected. Although a substantial part of the 12·5 kbp fragment apparently lacks protein-coding characteristics, no other elements, such as tRNA genes or transposons, were found. The nucleotide sequence data reported in this paper will appear in the EMBL, GenBank and DDBJ Nucleotide Sequence Databases under the Accession Number X83276.