摘要:

AbstractThe rDNAs of strains of the cactophilicPichiaspeciesP. amethionina, P. antillensis, P. barkeri, P. cactophila, P. caribaea, P. deserticola, P. heedii, P. kluyveri, P. norvegensis, P. opuntiae, P. pseudocactophila, P. thermotoleransand their varieties and anamorphs were mapped with 15 restriction endonucleases, and compared toP. membranaefaciensandP. salictariaas possible non‐cactophilic relatives. The existence of species complexes among those taxa was confirmed.P. membranaefacienswas a plausible ancestral species, and its closest relative in the cactophilic group wasP. deserticola. These two species appeared to be moderately related toP. heediiand toP. barkeri, but the latter was shown clearly to belong to theP. kluyvericomplex, in spite of a 6 mol% G+C difference in their nuclear DNAs.P. cactophilaandP. pseudocactophilaostensibly emerged fromP. norvegensis, a facultatively cactophilic yeast. TheP. amethionina, P. cactophilaandP. opuntiaespecies complexes appeared independent from one another and from all other species studied.P. salictariadid not appear to be related toP. amethionin

ISSN:0749-503X    

DOI:10.1002/yea.320090402

出版商:John Wiley&Sons, Ltd.    

年代:1993

数据来源: WILEY

摘要:

AbstractAmine oxidase (AMO) is a peroxisomal matrix protein ofHansenula polymorpha, which is induced during growth of the yeast in media containing primary amines as a sole nitrogen source. The deduced amino acid sequence of the protein contains an SRL sequence at nine amino acids from the C‐terminus. In this study, we have examined the possible role of the SRL motif in sorting of AMO to peroxisomes by mutating the corresponding gene sequence. For this purpose, we have developed a DNA construct that is specifically integrated into theAMOlocus of theH. polymorphagenome, placing the mutant gene under the control of the endogenousAMOpromoter and eliminating expression of the wild‐type gene. Analysis of a stable transformant, containing the desired gene configuration, showed that mutation of the C‐terminal sequence neither interfered with correct targeting of the protein into the peroxisome nor displayed significant effects on its activity. From this, it was concluded that the SRL‐containing C‐terminus is not essential for peroxisomal targeting of AMO inH. p

ISSN:0749-503X    

DOI:10.1002/yea.320090403

出版商:John Wiley&Sons, Ltd.    

年代:1993

数据来源: WILEY

摘要:

AbstractExpression of the vacuolar carboxypeptidase S (CPS1) gene inSaccharomyces cerevisiaeis regulated by the availability of nutrients. Enzyme production is sensitive to nitrogen catabolite repression; i.e. the presence of ammonium ions maintains expression of the gene at a low level. Transfer of ammonium–glucose pre‐grown cells to a medium deprived of nitrogen causes a drastic increase inCPS1RNA level provided that a readily usable carbon source, such as glucose or fructose, is available to the cells. Derepression of the gene by nitrogen limitation is cycloheximide‐insensitive. Neither glycerol, ethanol, acetate nor galactose support derepression ofCPS1expression under nitrogen starvation conditions. Non‐metabolizable sugar analogs (2‐deoxyglucose, 6‐methyl‐glucose or glucosamine) do not allow derepression ofCPS1, showing that the process is energy‐dependent. Production of carboxypeptidase yscS also increases several‐fold when ammonium‐pregrown cells are transferred to media containing glucose and a non‐readily metabolizable nitrogen source such as proline, leucine, valine or leucyl‐glycine. Analysis ofCPS1expression inRAS2+(high cAMP) andras2mutant (low cAMP) strains and in cells grown at low temperature (23°C) and in heat‐shocked cells (38°C) shows that steady‐state levels ofCPS1mRNA are not controlled by a low

ISSN:0749-503X    

DOI:10.1002/yea.320090404

出版商:John Wiley&Sons, Ltd.    

年代:1993

数据来源: WILEY

摘要:

AbstractWe cloned theSaccharomyces kluyveri HIS3homolog, k‐HIS3, and made a partial deletion of the gene. The k‐HIS3gene complemented aHIS3deletion inS. cerevisiae. The DNA sequences of the open reading frames (ORFs) of theHIS3homologs are 70% identical at the DNA level and 83% identical at the deduced amino acid level. The ORF upstream of the k‐HIS3gene is related to thePET56gene ofS. cerevisiaefound upstream of theHIS3gene ofS. cerevisiae. The ORF downstream from the k‐HIS3gene is not related to theDED1gene found downstream of theHIS3gene inS. cer

ISSN:0749-503X    

DOI:10.1002/yea.320090405

出版商:John Wiley&Sons, Ltd.    

年代:1993

数据来源: WILEY

摘要:

AbstractWe have cloned, sequenced and physically mapped theCYS3gene ofSaccharomyces cerevisiae. This gene can complement thecys3–1allele, and disruptions at this locus lead to cysteine auxotrophy. The predictedCYS3product is closely related (46% identical) to the rat cystathionine γ‐lyase (Ericksonet al., 1990), but differs in lacking cysteine residues. These results provide further evidence that the S288C strain of yeast resembles mammals in synthesizing cysteine solely via atrans‐sulfuration pathway. TheCYS3product was found to have strong homology to three other enzymes involved in cysteine metabolism: theEscherichia coli metBandmetCproducts and theS. cerevisiaeMET25 gene product. Thetrans‐sulfuration enzymes appears to form a diverged family and carry out related functions from bacteria to

ISSN:0749-503X    

DOI:10.1002/yea.320090406

出版商:John Wiley&Sons, Ltd.    

年代:1993

数据来源: WILEY

摘要:

AbstractYeast flocculation involves binding of surface lectins to carbohydrate receptors on neighbouring cell walls. Brewing strains ofSaccharomyces cerevisiaenormally become flocculent in the stationary phase of growth. This paper presents evidence that lectins are synthesized in exponential phase, inserted into the cell wall, and activated later at the time of flocculation onset.Cycloheximide failed to prevent flocculation unless it was added in early growth; with later additions progressively larger degrees of flocculation occurred. Flocculation onset was delayed by cycloheximide but was otherwise cycloheximide insensitive. Preflocculent cells could be artificially activated to full flocculation by heat. Artificial activation of samples from growing yeast cultures confirmed the progressive synthesis of lectins throughout exponential growth. Pronase E treatment of whole cells prior to heating prevented any activation of flocculation.It was concluded that lectins were synthesized continuously from an early stage of growth and rapidly inserted into the cell wall (accessible by pronase E), where they remained inactive for up to 14 h, before being activated at flocculation onset by an as‐yet unknown mechanism. It was found that lectin synthesis and activation occurred in all brewing strains teste

ISSN:0749-503X    

DOI:10.1002/yea.320090407

出版商:John Wiley&Sons, Ltd.    

年代:1993

数据来源: WILEY

摘要:

AbstractWe have constructed two secretion vectors forSchizosaccharomyces pombeusing an SV40 promoter and the secretion signals of the pGKL killer toxin complex derived fromKluyveromyces lactis. Although indigenous secretory glycoproteins tend to accumulate in the periplasmic space ofS. pombe, we have succeeded in the secretion of mouse α‐amylase into the culture medium. The efficiency of secretion, processing pattern, stability and culture conditions for mouse α‐amylase were studied inS. pombe. The 128 kDa killer secretion signal was more effective in directing secretion of mouse α‐amylase than the 28 kDa killer secretion signal. We detected a chymostatin‐sensitive protease activity in the culture medium ofS. pombe, which digests mouse α‐amylase secreted into the culture medium. The addition of 5 μg/ml chymostatin was shown to protect mouse α‐amylases from

ISSN:0749-503X    

DOI:10.1002/yea.320090408

出版商:John Wiley&Sons, Ltd.    

年代:1993

数据来源: WILEY

摘要:

AbstractPurification ofSaccharomyces cerevisiaecystathionine γ‐lyase (γ‐CTLase) was hampered by the presence of a protein migrating very close to it in various types of column chromatography. The enzyme and the contaminant were nevertheless separated by polyacrylamide gel electrophoresis. N‐terminal amino acid sequence analysis indicated that they are coded for byCYS3(CYI1) andMET17(MET25), respectively, leading to the conclusion thatCYS3is the structural gene for γ‐CTLase and that the contaminant isO‐acetylserine/O‐acetylhomoserine sulfhydrylase (OAS/OAH SHLase). Based on these findings, we purified γ‐CTLase by the following strategy: (1) extraction of OAS/OAH SHLase from aCYS3‐disrupted strain; (2) preparation of antiserum against it; (3) identification of a strain devoid of the OAS/OAH SHLase protein using this antiserum; and (4) extraction of γ‐CTLase from this strain. Purified γ‐CTLase had cystathionine γ‐synthase (γ‐CTSase) activity ifO‐succinylhomoserine, but notO‐acetylhomoserine, was used as substrate. From this notion we discuss the evolutional relationship betweenS. cerevisiae

ISSN:0749-503X    

DOI:10.1002/yea.320090409

出版商:John Wiley&Sons, Ltd.    

年代:1993

数据来源: WILEY

摘要:

AbstractThe sexual adhesion protein ofSaccharomyces cerevisiaeMATα cells, α‐agglutinin, could not be extracted from the cell wall with hot sodium dodecyl sulfate (SDS), but became soluble after digestion of the cell with laminarinase. This indicates that it is intimately associated with cell wall glucan. A fusion protein was constructed consisting of the signal sequence of yeast invertase, guar α‐galactosidase, and the C‐terminal half of the α‐agglutinin. Most of the fusion protein was incorporated in the cell wall. A small amount could be extracted with SDS, but most of it could only be extracted with laminarinase. On the other hand, cells containing a construct consisting of the signal sequence of invertase and α‐galactosidase released most of the α‐galactosidase into the medium and all cell wall‐associated α‐galactosidase was released by SDS. Labelling with antibodies showed that the α‐galactosidase part of the fusion protein was exposed on the surface of the cell wall. The results demonstrate that the C‐terminal half of the α‐agglutinin contains the information needed to incorporat

ISSN:0749-503X    

DOI:10.1002/yea.320090410

出版商:John Wiley&Sons, Ltd.    

年代:1993

数据来源: WILEY

摘要:

AbstractThe cytoplasmically inherited M double‐stranded (ds) RNA genome segment of killer virus ofSaccharomyces cerevisiaeis heat‐curable in some yeast strains but not in others. Temperature sensitivity is conferred on both M1and M2dsRNA satellite virus segments by the L‐A‐HN allele of the killer helper virus genome, but not by the L‐A‐H allele. Both diploidy and mating type heterozygosity of the host cell are also correlated with increased virus

ISSN:0749-503X    

DOI:10.1002/yea.320090411

出版商:John Wiley&Sons, Ltd.    

年代:1993

数据来源: WILEY