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PubMed2002

Adherence of yeast and filamentous forms of Candida albicans to cultured enterocytes.

OBJECTIVE: Systemic candidiasis is a major cause of complicating infections in intensive care units. Morbidity and mortality are high, even in those who receive appropriate antifungal therapy. Because the intestinal tract is considered a major portal of entry for systemic candidiasis, experiments were designed to clarify the ability of yeast and filamentous forms, as well as the INT1 gene product, to influence adherence of Candida albicans to the intestinal epithelium. DESIGN: Controlled. SETTING: University teaching hospital research laboratory. SUBJECTS: Mature Caco-2 and HT-29 cultured enterocytes. INTERVENTIONS: C. albicans INT1 mutant strains, defective in filament production, were used to observe the ultrastructural surface interactions of C. albicans with cultured intestinal epithelial cells, namely Caco-2 and HT-29 cells. These mutant strains also were used to quantify the effect of the INT1 gene product on C. albicans adherence (yeast and filamentous forms) to cultured enterocytes. Ultrastructural surface interactions of C. albicans with cultured enterocytes were observed with high resolution scanning electron microscopy. C. albicans adherence to cultured enterocytes was quantified by using a colorimetric enzyme-linked immunosorbent assay. MEASUREMENTS AND MAIN RESULTS: Both yeast and filamentous forms of C. albicans appeared tightly adherent to the apical surface of cultured enterocytes, and INT1 appeared to have little, if any, effect on these ultrastructural surface interactions. The distal ends of C. albicans filaments appeared to mediate adherence to enterocyte apical microvilli, and thigmotropism (contact guidance) appeared to play a role in C. albicans adherence. The absence of functional INT1 was associated with decreased adherence of C. albicans yeast forms to cultured enterocytes. CONCLUSIONS: Although functional INT1 appeared to facilitate adherence of C. albicans yeast forms to cultured enterocytes, the role of INT1 in adherence of filamentous forms was unclear, and both yeast and filamentous forms could adhere to, and perhaps invade, the apical surface of cultured enterocytes.

Caco-2 Cells↗
PubMed2003

Can yeast isolation in peritoneal fluid be predicted in intensive care unit patients with peritonitis?

OBJECTIVE: To generate and validate a predictive score of yeast isolation based on independent risk factors of yeast isolation in intensive care unit patients with peritonitis. DESIGN: Retrospective cohort study to determine independent risk factors of yeast isolation, generation of the score, and validation in a prospective cohort of patients with peritonitis. SETTING: Tertiary-care, university-affiliated hospital. PATIENTS: Two hundred twenty-one patients with peritonitis hospitalized in a surgical intensive care unit between 1994 and 1999 for the retrospective cohort and 57 patients in the prospective cohort (2000). MEASUREMENTS AND MAIN RESULTS: Four independent risk factors of yeast isolation in peritoneal fluid (similar odds ratio) were found in the retrospective cohort: female gender, upper gastrointestinal tract origin of peritonitis, intraoperative cardiovascular failure, and previous antimicrobial therapy at least 48 hrs before the onset of peritonitis. A score based on the number of risk factors was constructed (grade A = zero or one risk factor, grade B = at least two risk factors, grade C = at least three risk factors, and grade D = four risk factors), and validated in the prospective cohort. For a grade C score, sensitivity was 84%, specificity was 50%, positive and negative predictive values were 67% and 72%, respectively, and overall accuracy was 71%. CONCLUSIONS: Four independent risk factors of yeast isolation in the peritoneal fluid were identified in critically ill surgical patients with peritonitis. The presence of at least three of these factors (grade C score) was associated with a high rate of yeast detection. This approach could be helpful to initiate early antifungal therapy in this patient population.

Aged↗
PubMed2001

Analysis of the constitution of the beer yeast genome by PCR, sequencing and subtelomeric sequence hybridization.

The lager brewing yeasts, Saccharomyces pastorianus (synonym Saccharomyces carlsbergensis), are allopolyploid, containing parts of two divergent genomes. Saccharomyces cerevisiae contributed to the formation of these hybrids, although the identity of the other species is still unclear. The presence of alleles specific to S. cerevisiae and S. pastorianus was tested for by PCR/RFLP in brewing yeasts of various origins and in members of the Saccharomyces sensu stricto complex. S. cerevisiae-type alleles of two genes, HIS4 and YCL008c, were identified in another brewing yeast, S. pastorianus CBS 1503 (Saccharomyces monacensis), thought to be the source of the other contributor to the lager hybrid. This is consistent with the hybridization of S. cerevisiae subtelomeric sequences X and Y' to the electrophoretic karyotype of this strain. S. pastorianus CBS 1503 (S. monacensis) is therefore probably not an ancestor of S. pastorianus, but a related hybrid. Saccharomyces bayanus, also thought to be one of the contributors to the lager yeast hybrid, is a heterogeneous taxon containing at least two subgroups, one close to the type strain, CBS 380T, the other close to CBS 395 (Saccharomyces uvarum). The partial sequences of several genes (HIS4, MET10, URA3) were shown to be identical or very similar (over 99%) in S. pastorianus CBS 1513 (S. carlsbergensis), S. bayanus CBS 380T and its close derivatives, showing that S. pastorianus and S. bayanus have a common ancestor. A distinction between two subgroups within S. bayanus was made on the basis of sequence analysis: the subgroup represented by S. bayanus CBS 395 (S. uvarum) has 6-8% sequence divergence within the genes HIS4, MET10 and MET2 from S. bayanus CBS 380T, indicating that the two S. bayanus subgroups diverged recently. The detection of specific alleles by PCR/RFLP and hybridization with S. cerevisiae subtelomeric sequences X and Y' to electrophoretic karyotypes of brewing yeasts and related species confirmed our findings and revealed substantial heterogeneity in the genome constitution of Czech brewing yeasts used in production.

Alleles↗
PubMed1994

Synthesis of heat-shock proteins in mycelia and yeast forms of Paracoccidioides brasiliensis.

The induction of heat-shock proteins has been postulated to play a role not only in thermo-adaptation, but also in phase transition of the dimorphic fungi. In this study, we used yeast and mycelial forms of the thermally dimorphic fungus Paracoccidioides brasiliensis to evaluate the effect of temperature on the induction of the heat-shock response. We also evaluated protein synthesis by P. brasiliensis caused by exposure to low pH and H2O2. Analysis of protein synthesis by SDS-PAGE disclosed that P. brasiliensis mycelia increased synthesis of all major constitutive proteins when stressed at 37 degrees C and increased synthesis of three non-constitutive proteins of 134, 82 and 28 kDa at 40 degrees C. Yeasts incubated at 40 degrees C showed decreased synthesis of five constitutive proteins (136, 98, 62, 57 and 54 kDa) and the appearance of three new proteins (134, 82 and 28 kDa). There was a decrease in the synthesis of all major constitutive proteins except for three proteins of 141, 136 and 16 kDa when yeast cells were incubated at 25 degrees C. When stressed by low pH and H2O2, P. brasiliensis yeast increased synthesis of one (134 kDa) and five (134, 104, 82, 52 and 40 kDa) non-constitutive proteins, respectively. P. brasiliensis mycelia and yeast forms disclosed the same profile of protein synthesis when stressed at temperatures that trigger phase transition (37 degrees C for mycelia; 25 degrees C for yeast).(ABSTRACT TRUNCATED AT 250 WORDS)

Autoradiography↗
PubMed1992

Yeast site-specific ribonucleoprotein endoribonuclease MRP contains an RNA component homologous to mammalian RNase MRP RNA and essential for cell viability.

RNase MRP is a site-specific ribonucleoprotein endoribonuclease that cleaves RNA sequence complementary to mammalian mitochondrial origins of replication in a manner consistent with a role in primer RNA metabolism. The same activity in the yeast Saccharomyces cerevisiae has recently been identified; it cleaves an RNA substrate complementary to a yeast mitochondrial origin of replication at an exact site of linkage of RNA to DNA. We have purified this yeast enzyme further and detect a single, novel RNA of 340 nucleotides associated with the enzymatic activity. The single-copy nuclear gene for this RNA was sequenced and mapped to the right arm of chromosome XIV. The identity of the clone, as encoding the RNA copurifying with enzymatic activity, was confirmed by a match to the directly determined sequence of the RNA. The gene sequence also identified a 340-nucleotide RNA in total yeast RNA and in purified RNase MRP enzyme preparations. Inspection of the sequence of the yeast RNA revealed homologies to the RNA component of mouse RNase MRP, 49% overall with specific regions of much greater similarity. The flanking regions of the gene showed characteristics of an RNA polymerase II transcription unit, including a TATAAA box and a 7/8 match to the yeast cell cycle box UAS. The RNase MRP RNA gene was deleted by insertional replacement and found to be essential for cellular viability, indicating a critical nuclear role for RNase MRP.

Animals↗
PubMed1994

Conserved and nonconserved functions of the yeast and human TATA-binding proteins.

Although the TATA-binding protein (TBP) is highly conserved throughout the eukaryotic kingdom, human TBP cannot functionally replace yeast TBP for cell viability. To investigate the basis of this species specificity, we examine the in vivo transcriptional activity of human TBP at different classes of yeast promoters. Consistent with previous results, analysis of yeast/human hybrid TBPs indicates that growth defects are not correlated with the ability to promote TATA-dependent polymerase II (Pol II) transcription or to respond to acidic activator proteins. Human TBP partially complements the growth defects of a yeast TBP mutant with altered TATA element-binding specificity, suggesting that it carries out sufficient Pol II function to support viability. However, human TBP does not complement the defects of yeast TBP mutants that are specifically defective in transcription by RNA polymerase III. Three independently isolated derivatives of human TBP that permit yeast cell growth replace arginine 231 with lysine; the corresponding amino acid in yeast TBP (lysine 133) has been implicated in RNA polymerase III transcription. Transcriptional analysis indicates that human TBP functions poorly at promoters recognized by RNA polymerases I and III and at RNA Pol II promoters lacking a conventional TATA element. These observations suggest that species specificity of TBP primarily reflects evolutionarily diverged interactions with TBP-associated factors (TAFs) that are necessary for recruitment to promoters lacking TATA elements.

Amino Acid Sequence↗
PubMed2002

Subcellular localization of the yeast proteome.

Protein localization data are a valuable information resource helpful in elucidating eukaryotic protein function. Here, we report the first proteome-scale analysis of protein localization within any eukaryote. Using directed topoisomerase I-mediated cloning strategies and genome-wide transposon mutagenesis, we have epitope-tagged 60% of the Saccharomyces cerevisiae proteome. By high-throughput immunolocalization of tagged gene products, we have determined the subcellular localization of 2744 yeast proteins. Extrapolating these data through a computational algorithm employing Bayesian formalism, we define the yeast localizome (the subcellular distribution of all 6100 yeast proteins). We estimate the yeast proteome to encompass approximately 5100 soluble proteins and >1000 transmembrane proteins. Our results indicate that 47% of yeast proteins are cytoplasmic, 13% mitochondrial, 13% exocytic (including proteins of the endoplasmic reticulum and secretory vesicles), and 27% nuclear/nucleolar. A subset of nuclear proteins was further analyzed by immunolocalization using surface-spread preparations of meiotic chromosomes. Of these proteins, 38% were found associated with chromosomal DNA. As determined from phenotypic analyses of nuclear proteins, 34% are essential for spore viability--a percentage nearly twice as great as that observed for the proteome as a whole. In total, this study presents experimentally derived localization data for 955 proteins of previously unknown function: nearly half of all functionally uncharacterized proteins in yeast. To facilitate access to these data, we provide a searchable database featuring 2900 fluorescent micrographs at http://ygac.med.yale.edu.

Algorithms↗
PubMed2003

Correlations between gene expression and gene conservation in fission yeast.

Genes can be expressed at a wide range of levels, and they show different degrees of cross-species conservation. We compared gene expression levels to gene conservation by integrating microarray data from fission yeast (Schizosaccharomyces pombe) with lists of "core" genes (present in worm and budding and fission yeasts), "yeast-specific" genes (present in budding and fission yeasts, but not in worm), and "pombe-specific" genes (present in fission yeast only). Whereas a disproportionate number of core genes are highly expressed in vegetatively growing cells, many pombe-specific genes are expressed at lower levels. This bias is less pronounced in cells undergoing sexual development, when many pombe-specific genes become highly expressed. This implies that organism-specific proteins are more likely to function during specialized processes such as cellular differentiation. Accordingly, pombe-specific genes were overrepresented among genes induced during sexual development; they were particularly enriched in a group of genes induced during meiotic prophase, when homologous chromosomes pair and recombine. This raises the possibility that organism-specific genes with functions in meiotic prophase favor speciation by preventing fruitful meiosis between closely related organisms. Finally, the set of genes induced late during sexual differentiation, at the time of spore formation, was enriched in yeast-specific genes, indicating that these genes play specialized roles in ascospore development.

Conserved Sequence↗
PubMed1984

Formation of inverted dimer plasmids after transformation of yeast with linearized plasmid DNA.

The formation of an inverted dimer plasmid on transformation with linear molecules is formally analogous to the fusion of the daughters of a broken chromosome at their broken ends. In the latter case, this leads to the formation of a dicentric chromosome, which could break at anaphase. Hence the process is cyclic. Similarly, when our linear molecules are modified by the addition of a cloned yeast centromere, dicentric inverted dimers are not obtained. Instead, we obtain monocentric plasmids with partial duplication and deletion that apparently derive from a process of fusion, bridge-breakage, and fusion. This is not surprising, since it is known that dicentric plasmids undergo breakage in yeast (Mann and Davis 1983). However, any apparent similarity of this process to that which occurs with a broken chromosome in maize must be tempered by the special nature of the transformation process. Most significantly, inverted dimers are rare when sonicated carrier DNA is not present during the transformation. This requirement is not understood, but it is a condition that may not be met in a yeast cell harboring a broken chromosome. It is possible that carrier DNA induces a repair process that results in fusion. On the other hand, a property of the transformation process that results in an inhibition of fusion may be overcome by the presence of carrier DNA. Most inverted dimers are apparently formed from an interaction between two input linear molecules. We cannot rule out the possibility that a minor fraction derive from a single molecule. Thus, the fusion of two input molecules is a much more efficient process than a replicative process that could occur with single linear molecule. For a similar fusion process to occur with a broken yeast chromosome, replication would be required. We do not know if a broken yeast chromosome can replicate. Evidence consistent with the presence of a breakage-fusion-bridge process in yeast has been obtained through the formation of dicentric chromosomes via meiotic recombination (Haber et al. 1984). Spores from these meioses sometimes give rise to a clone that is mixed for markers of the chromosome that could have been dicentric. A process of fusion-bridge-breakage could account for the formation of some of these mixed clones. However, the dicentric chromosomes apparently often survive meiotic disjunction and break in the spore's first mitotic anaphase or possibly in a later generation. Thus, the interpretation of the origin of these mixed clones is uncertain. Some aspects of the fusion process are especially intriguing.(ABSTRACT TRUNCATED AT 400 WORDS)

Chromosome Mapping↗
PubMed1993

Wheat (Triticum aestivum L.) [gamma]-Gliadin Accumulates in Dense Protein Bodies within the Endoplasmic Reticulum of Yeast.

Following their sequestration into the endoplasmic reticulum (ER), wheat storage proteins may either be retained and packaged into protein bodies within this organelle or transported via the Golgi to vacuoles. We attempted to study the processes of transport and packaging of wheat storage proteins using the heterologous expression system of yeast. A wild-type wheat [gamma]-gliadin, expressed in the yeast cells, accumulated mostly within the ER and was deposited in protein bodies with similar density to natural protein bodies from wheat endosperm. This suggested that wheat storage proteins contain sufficient information to initiate the formation of protein bodies in the ER of a heterologous system. Only a small amount of the [gamma]-gliadin was transported to the yeast vacuoles. When a deletion mutant of the [gamma]-gliadin, lacking the entire N-terminal repetitive region, was expressed in the yeast cells, the mutant was unable to initiate the formation of protein bodies within the ER and was completely transported to the yeast vacuole. This strongly indicated that the information for packaging into dense protein bodies within the ER resides in the N-terminal repetitive region of the [gamma]-gliadin. The advantage of using yeast to identify the signals and mechanisms controlling the transport of wheat storage proteins and their deposition in protein bodies is discussed.

Journal Article↗
PubMed2004

The role of pre-existing aggregates in Hsp104-dependent polyglutamine aggregate formation and epigenetic change of yeast prions.

Amyloid-like protein aggregates have been implicated in various diseases and in the protein-based inheritance of yeast prions. The molecular chaperone Hsp104 has been shown to be necessary for the aggregate formation of polyglutamine in yeast, and for the maintenance of several yeast prion phenotypes through the formation of self-propagating aggregates. In this paper, we show that the polyglutamine aggregates that are formed independently of Hsp104, are required for Hsp104 to efficiently produce more aggregates. Similarly, in the yeast prion [PSI+] system, Hsp104-dependent epigenetic changes to the [PSI+] prion phenotype require the presence of prion aggregates in the normal [psi-] state. We also show that the co-localization of different prion aggregates suggests that cross-seeding by different yeast prions increases the probability of Hsp104-dependent epigenetic change. These findings highlight the role of pre-existing aggregates in chaperone-dependent establishment of the epigenetic trait in yeast prions, and possibly in the pathology of several neurodegenerative diseases.

Epigenesis, Genetic↗
PubMed1995

HIV disease and Malassezia yeasts: a quantitative study of patients presenting with seborrhoeic dermatitis.

Seborrhoeic dermatitis (SD) is a disease that affects 1-3% of the general population, 3-5% of young adults and 20-83% of patients with AIDS. Malassezia yeasts have been associated with the pathogenesis of this condition. The association between the Malassezia yeasts and HIV-related SD is still a controversial subject. The objective of our study was to investigate the role of Malassezia yeasts in the pathogenesis of SD in the HIV population comparing the number of yeasts' cells with the severity of the disease and degree of immunosuppression. We used two quantitative counting methods: direct counting with Sellotape-stripped skin and recovery of the yeasts in culture using contact plates. This investigation has demonstrated that there is a trend between numbers of yeasts present on lesional skin, severity of SD and CD4-positive T lymphocytes count in HIV-positive patients. No quantitative differences were observed between HIV-related and non-HIV related SD.

Adult↗
PubMed1993

Gastrointestinal stability of baker's yeast allergens: an in vitro study.

An in vitro model was established to study the stability of baker's yeast (Saccharomyces cerevisiae) allergens in conditions simulating the gastrointestinal tract. The protocol consisted of 2 hr incubation under gastric conditions (pH 1.2, +37 degrees C and gastric enzymes) and 2 hr incubation under duodenal conditions (pH 6.8, +37 degrees C and duodenal enzymes). These were studied together and separately, as well as under pure acidic conditions without gastric enzymes. The yeast extracts contained equal amounts of allergen and were analyzed by IgE-immunoblotting. The acidic conditions had partly an enhancing and slightly degrading effect on the yeast allergens, whereas the gastric enzymes destroyed several allergens, including the important intermediate allergens of 31 and 45 kD. After treatment under both gastric and duodenal conditions most of the yeast allergens were destroyed, except mannan and a 10 kD protein component. The findings suggest that the allergen exposure caused by baker's yeast takes place mainly on the mucosal surfaces orally and oesophageally and through viable baker's yeast organisms that manage to pass the stomach and duodenum and possibly lead to intestinal growth of the organism. Patients with IgE production against the 10 kD allergen and mannan are, however, moderately exposed to allergens consisting of soluble antigenic material only.

Allergens↗
PubMed2006

Classical and molecular analyses to characterize commercial dry yeasts used in wine fermentations.

AIMS: The aim of the work was to apply PCR-temperature gradient gel electrophoresis (PCR-TGGE) and restriction enzyme analysis (RE) assays to identify commercially available starters of Saccharomyces cerevisiae sensu stricto complex. METHODS AND RESULTS: To characterize an analysed pool of 62 active dry yeasts of different brands used in wine fermentation practices, classical microbiological tests were also performed as well as evaluation of contamination with lactic acid bacteria and non-Saccharomyces yeasts. PCR-TGGE and RE were used in order to provide fast and reliable methods to identify and differentiate enological yeasts. Proposed molecular methods enabled to identify particular strains within 36 h after colony isolation and directly from dry yeast suspension. CONCLUSIONS: The methods are highly recommended to obtain reliable results on yeast strain differentiation in a significantly shorter time if compared to classical fermentation tests. SIGNIFICANCE AND IMPACT OF THE STUDY: The obtaining of yeast strain differentiation in a short time and without plating is a good tool for a rapid discrimination among enological strains used as starters in enological practices.

Colony Count, Microbial↗
PubMed1992

Isolation and characterization of sporulation-specific promoters in the yeast Saccharomyces cerevisiae.

A library of random yeast genomic DNA:lacZ fusions has been constructed using an episomal yeast-Escherichia coli shuttle vector (pCS1). Plasmid pCS1 requires insertion of a promoter and an in frame ATG codon upstream of its resident truncated lacZ gene to regulate expression in yeast. Yeast genomic DNA fragments of 4-6 kb were generated by partial digestion with Sau3A and ligated into the unique BamHI site of plasmid pCS1 to generate a library of 5 x 10(4) individual E. coli transformants. This library was screened to identify promoter-lacZ fusions that were expressed uniquely during sporulation. Of 342 yeast transformants that exhibited beta-galactosidase activity, two were found to express the lacZ gene in a sporulation-specific manner. This paper presents the characterization of two genomic yeast DNA fragments containing promoters that control lacZ expression during the sporulation process. Expression from the promoter present in plasmid pJC18 occurred from 11-21 hours into the sporulation process, while the promoter in plasmid pJC217 was active from 4-14 hours. Staining of nuclear DNA to correlate nuclear morphology with timing of gene expression showed when each of these promoters was active in terms of the morphological stages of sporulation.

Blotting, Southern↗
PubMed1998

The relationship between salivary histatin levels and oral yeast carriage.

Candida species are common commensal inhabitants of the oral cavity. Human saliva contains antifungal proteins called histatins. We tested the hypothesis that oral yeast status is related to salivary histatin levels. Thirty subjects were divided into two groups based on the presence (n = 15) or absence (n = 15) of yeast on oral mucosa surfaces. Unstimulated and stimulated submandibular and sublingual and parotid saliva was collected from each subject. Salivary flow rates were measured and histatin concentrations were determined in the stimulated saliva samples. The yeast colony positive group showed lower median unstimulated parotid saliva flow rates as well as lower median concentrations of total histatins in submandibular and sublingual saliva. There was a negative correlation between yeast colony-forming units and unstimulated parotid saliva flow rates and between yeast colony-forming units and submandibular and sublingual saliva histatin concentration and secretion. The results suggest that oral yeast status may be influenced by unstimulated parotid saliva flow rates and by submandibular and sublingual histatin concentration and secretion.

Aged↗
PubMed1976

Lysis of yeast cell walls. Lytic beta-(1 leads to 6)-glucanase from Bacillus circulans WL-12.

When grown in a mineral medium with yeast cell walls or yeast glucan as the sole carbon source, Bacillus circulans WL-12 produces wall-lytic enzymes in addition to non-lytic beta-(1 leads to 3) and beta-(1 leads to 6)-glucananases. The lytic enzymes were isolated from the culture liquid by adsorption on insoluble yeast glucan in batch operation. After digestion of the glucan, the mixture of enzymes was chromatographed on hydroxylapatite on which the lytic activity could be resolved into one lytic beta-(1 leads to 6)glucanase and two lytic beta-(1 leads to 3)-glucanase was further purified by chromatography over diethylamino-ehtyl-agarose and carboxymethyl cellulose. Its specific activity on pustulan was 6.2 units per mg of protein. The enzyme moved as a single protein with a molecular weight of 54000 during sodium dodecylsulphate electrophoresis in slab gels. Hydrolysis of pustulan went thorugh a series of oligosaccharides, leading to a mixture of gentiotriose, gentiobiose and glucose. The enzyme also produced small amounts of gentiobiose from laminarin and pachyman and on this basis its lytic activity on yeast cell walls,was attribut beta-(1 leads to 3)-linked oligosaccharides were not detected. The lytic beta-(1 leads to 6)-glucanase has an optimum pH of 6.0. Pustulan hydrolysis followed Michaelis-Menten kinetics. A Km of 0.29 mg pustulan per ml and a V of 9.1 micro-equivalents of glucose released/min per mg of enzyme were calculated. The enzyme has no metal ion requirement. The lytic beta-(1 leads to 6)-glucanase differs in essence from the non-lytic beta-(1 leads to 6)-glucanase of the same organism by its positive action on yeast cell walls and yeast glucan and its much lower specific activity on soluble pustulan.

Bacillus↗
PubMed1976

Lysis of yeast cell walls. Lytic beta-(1 leads to 3)-glucanases from Bacillus circulans WL-12.

Bacillus circulans WL-12 when grown in a mineral medium with yeast cell walls or yeast glucan as the soli carbon source, produced five beta-glucanases. Two beta-(1 leads to 3)-glucanases (I and II), which are lytic to yeast cell walls, were isolated from the culture liquid by batch adsorption on yeast glucan, and separated by chromatography on hydroxylapatite. Lytic beta-(1 leads to 3)-glucanase I was further purified by carboxymethylcellulose chromatography. The specific activity of lytic beta-(1 leads to 3)-glucanase I on laminarin was 4.1 U per mg of protein. The enzyme moved as a single protein with a molecular weight of 40000 during sodium dodecylsulfate electrophoresis in slab gels. It was specific for the beta-(1 leads to 3)-glucosidic bond but the enzyme did not hydrolyze laminaribiose. Hydrolysis of laminarin went through a series of oligosaccharides, and laminaribiose and glucose accumulated till the end of the reaction. A small amount of gentibiose was also produced from laminarin. Products from yeast cell walls and yeast glucan included laminaripentaose, laminaritriose, laminaribiose, glucose and gentiobiose, but no laminaritetraose was detected. This glucanase has an optimum pH of 5.5.

Bacillus↗