摘要:

AbstractAmine oxidase from the yeastHansenula polymorphais a peroxisomal protein. The signal for routing of the protein into peroxisomes has not been identified yet. Expression of a mutant amine oxidase inH. Polymorphahas revealed that the C‐terminal sequence, which possesses an internal SRL tripeptide, is not involved in targeting (Faberet al., unpublished). We have explored heterologous expression of the amine oxidase gene (AMO) inSaccharomyces cerevisiaeto investigate the conservation of peroxisomal targeting pathways between yeasts. Surprisingly, wide‐type amine oxidase is not recognized as a peroxisomal protein byS. cerevisiae. The enzyme, which was fully active and acumulated to levels similar to those found inH. polymorpha, stayed entirely in the cytosol. However, fusing a SKL or a SRL sequence to the C‐terminus forced the protein at least partially into peroxisomes of the heterologous host. These data suggest that the functional targeting sequence of amine oxidase may differ from the C‐terminal peroxisomal targeting signal S/C/A‐K/R/H‐L (Gouldet al., 1989). Contrary to the established tripeptide motif, the amine oxidase targeting signal appears not to be conserved between the different ye

ISSN:0749-503X    

DOI:10.1002/yea.320080402

出版商:John Wiley&Sons, Ltd.    

年代:1992

数据来源: WILEY

摘要:

AbstractACandida maltosachromosomal DNA fragment which confers high frequency transformation ofC. maltosaand autonomous replication of recombinant plasmids was cloned and sequenced. Analysis of the nucleotide sequence of the cloned DNA revealed a sequence homologous forC. maltosaautonomously replicating sequence (ARS) elements. Vector pRJ1 forC. maltosawas constructed, which contained a 1.3 kb ARS sequence, pICEM‐19H and theADE1gene ofC. maltosa. Southern blot analysis suggested that the copy number of pRJ1 inC. maltosawas approximately 20 per genome. The sequence analysis also revealed an open reading frame, encoding a polypeptide with high homology (70%) to the RS15 protein ofBrugia pagangi. This open reading frame has an intron with canonical sites for correct splicing inSaccharomyces cerevisia

ISSN:0749-503X    

DOI:10.1002/yea.320080403

出版商:John Wiley&Sons, Ltd.    

年代:1992

数据来源: WILEY

摘要:

AbstractThe α and β components of the secreted K1killer toxin ofSaccharaomyces cerevisiaeare derived from residues 45–147 and 234–316, respectively, of the 316 residue prepotoxin (ppTox). The β N‐terminus is produced by Kex2 cleavage after Lys Arg233, when β1a(the mature sequence of β‐lactamase)is fused at this site and the fusion is expressed form thePGKpromoter in pDT17, a multicopy plasmid, unexpectedly modest levels of βla secretion resulted. Over‐expression of Kex2 failed to increase βla secretion while akex2‐null mutation reduced secretion by 98%. βla secretion in a Kex+strain was not enhanced by inactivation of the a toxin component or by deletion of most of its central hydrophobic segments. However SP‐βla, produced by deletion of ppTox residues 35–176, expressed 10‐fold higher βla activity and the precursor was not secreted with similar efficiency in akex−2null strain. Fusions of βla to ppTox at Ala34or Ala46also led to efficient secretion in bothKEX2andkex−2‐null strains. Since these βla fusions differ only in segments well downstream of the signal peptide and all had similar transcript levels, the efficiency of βla secretion is apparently determined by the efficiency with efficiency with which these fusions are translocated to the Golgi compartment where Kex2 is active. Efficiency is high for the shorter fusions but is 10% or less for the longer fusions; even this fraction is apparently diverted to the vacuole if not cleaved by Kex2. SP‐βla was athe most efficient construct tested; secreted βla reacahed 4% of total cell protein, modestly exceeding levels produced by fusion to the MFα1‐encoded preproα‐factor, suggesting potential f

ISSN:0749-503X    

DOI:10.1002/yea.320080404

出版商:John Wiley&Sons, Ltd.    

年代:1992

数据来源: WILEY

摘要:

AbstractTwoSaccharomyces cerevisiaegenes previously unknown to be required for DNA synthesis have ben identified by screening a collection of temperature‐sensitive mutants. The effects of mutations inDNA43andDNA52on the rate of S phase DNA synthesis were detected by monitoring DNA synthesis in synchronous populations that were obtained by isopycnic density centrifugation.dna43‐1anddna52‐1cells undergo cell‐cycle arrest at the restrictive temperature (37°C), exhibiting a large‐budded terminal phenotype; the nuclei of arrested cells are located at the neck of the bud have failed to undergo DNA replication. These phenotypes suggest thatDNA43andDNA52are required for entry into or completion of S phase.DNA43andDNA52were cloned by their abilities to suppress the temperature‐sensitive lethal phenotypes ofdna43‐1anddna52‐1cells, respectively. DNA sequence analysis suggested thatDNA43andDNA52encode proteins of 59.6 and 80.6 kDa, respectively. BothDNA43andDNA52are essential for viability and genetic mapping experiments indicate that they represent previously unidentified genes:DNA43is located on chromosome IX, 32 cM distal fromhis5andDNA52is located on chromosome IV,

ISSN:0749-503X    

DOI:10.1002/yea.320080405

出版商:John Wiley&Sons, Ltd.    

年代:1992

数据来源: WILEY

摘要:

AbstractEthanol and CO2production from gluecose by non‐proliferating suspensions of aerobicaly‐grown, glucose‐derepressed wild‐typeSacharomyces cerevisiaeis inhibited by O2; monitoring by mass spectrometry provides a direct method for measurement of the Pasteur effect.Under aerobic conditons, that part of the CO2evolved equivalent to the O2consumed, is produced by respiration: subtraction of this respiratory CO2from the total gives, CO2produced by aerobic glycolysis. Pasteur quotients (anaerobic CO2/aerobic glycolytic CO2) were within the range 1.2 to 3.0. The Pasteur effect was not observed in the presence of carbonyl cyanidm‐chlorophenylhydrazone, an uncoupler of mitochondrial energy metabolism, or in a ρ cytoplasmicpetitemutant. A ‘non‐allosteric’ mutant with an altered regulatory subunit of phosphofructokinase showed no Pasteur effect. Strains bearing a nonsense mutationpfk1in the catalytic subnit of soluble phosphofructokinase (PFKI) also showed no Pasteur effect; the residual fermentative activity of this strain was dependent on PFKII, the particulate phosphofructokinase. A double mutant lacking both PFKI and glucose‐6‐phosphat dehydrogenase showed similar characteristics to those of the singlepfk1mutant; this indicates that the hexose monophosphate shunt is not acting to bypass the phosphofructokinase block. A ‘hyper‐allosteric’ mutant altered in the regulatory subunit encoded by the genePFK2showed characteristics of glucose fermentation and ethanol oxidation very similar to those of wild‐type organisms. These results indicate that either of the two phosphofructokin

ISSN:0749-503X    

DOI:10.1002/yea.320080406

出版商:John Wiley&Sons, Ltd.    

年代:1992

数据来源: WILEY

摘要:

AbstractTheSaccharomyces cerevisiae ADR1gene has recently been demonstrated to control transcription of several genes encoding peroxisomal proteins or proteins necessary for peroxisome formation. Therefore, the effect of two other genes(SNF1 (CAT1, CCR1)andSNF4 (CAT3))known to control derepression of glucose‐repressible genes was studied. Levels of transcripts of genes encoding catalase A, fatty acid β‐oxidation enzymes and of thePAS1gene are reduced insnf1andsnf4mutants of ethanol as well as on oleic acid medium. By immunogold labelling with an antibody directed against peroxisomal thiolase, clusters of peroxisomes were detected in wild‐types cells, whereas smaller single peroxisomes were observed inadr1mutant cells. Results of immunofluorescence experiments are consistent with these observations. No peroxisomes were detected insnf1andsnf4mutants by immunogold labelling as well as by imunofluore

ISSN:0749-503X    

DOI:10.1002/yea.320080407

出版商:John Wiley&Sons, Ltd.    

年代:1992

数据来源: WILEY

ISSN:0749-503X    

DOI:10.1002/yea.320080408

出版商:John Wiley&Sons, Ltd.    

年代:1992

数据来源: WILEY

摘要:

AbstractMSB2was identified previously as a multicopy suppressor of a temerature‐sensitive mutation inCDC24, a gene required for polarity establishment and bud formation inSaccharomyces cerevisiae. The inferredMSB2product contains 1306 amino acids, 42% of which are Ser or Thr. Its Ser+Thr‐richnes and hydrophobicity profile suggest that Msb2p may be an integral membrane protein containing a long, periplasmic, N‐terminal domain and a short, cytoplasmic, C‐terminal domain. Cells that lackMSB2display no obvious mutant phenotypes.MSB2is located between the centromere andKSS1on the right arm of chromosome VII. Although physical mapping suggests thatMSB2andLEU1(on the left arm of chromosome VII) are approximately 40 kb apart, the genetic map distance observed betweenleulandmsb2::URA3marker was only

ISSN:0749-503X    

DOI:10.1002/yea.320080409

出版商:John Wiley&Sons, Ltd.    

年代:1992

数据来源: WILEY

摘要:

AbstractWe have entirely sequaenced an 8.3 kb segment localized on the left arm of chromosome XI ofSaccharomyces cerevisiae. Five new open reading frames have been uncovered. One of them encodes a new mitochondrial carrier protein which is dispensable for growth on glycerol medium. Another could be a new member of the G protein family. A third possesses thePAAKKmotif common to H1 histones.

ISSN:0749-503X    

DOI:10.1002/yea.320080410

出版商:John Wiley&Sons, Ltd.    

年代:1992

数据来源: WILEY

ISSN:0749-503X    

DOI:10.1002/yea.320080401

出版商:John Wiley&Sons, Ltd.    

年代:1992

数据来源: WILEY