Purification of endo-(1→3)-β-D-glucanases lysing yeast cell walls from Rhizoctonia solani

T, Usui; Totani, Kiichiro; Totsuka, Atsushi; Oguchi, Masao

doi:10.1016/0304-4165(85)90126-6

Cited by 16 publications

(11 citation statements)

References 23 publications

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“…min-' . mg protein -') (Reichelt and Fleet, 1981;Sanchez et al, 1982;Notario, 1982;Usui et al, 1985;Tangarone et al, 1989;de la Cruz et al, 1995). The increase of V,,,, with the size of the substrate, indicates that the number of 1,3-/?-glucosidic bonds/molecule of substrate influence directly the activity of the 74-kDa endo-p-glucanase, suggesting a putative repetitive attack where the enzyme cleaves several times the glucan chain before dissociation (Robyt and French, 1970;Thoma, 1976).…”

Section: Discussionmentioning

confidence: 99%

Purification and Characterization of an Endo‐1,3‐β‐Glucanase from Aspergillus fumigatus

Fontaine

Hartland

Beauvais

et al. 1997

European Journal of Biochemistry

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An endo-I ,3-p-glucanase was purified from a cell wall autolysate of Aspcvgillus ,fumigatus. This pglucanase activity was associated with a glycosylated 74-kDa protein. Using a sensitive colorimetric assay and a high-performance anion-exchange chromatography with a pulsed electrochemical detector for product analysis, it was shown that the endoglucanase hydrolysed exclusively linear 1,3-P-glucan chains, had an optimum pH of 7.0 and an optimum temperature of 60°C. A substrate kinetic study gave a K,, value of 0.3 mg/ml for soluble (laminarin and laminari-oligosaccharides) and 1 .18 mg/ml for insoluble (curdlan) 1,3-P-glucan. Laminar-oligosaccharide degradation, analysed by HPLC, showed that the endoglucanase bind to the subtrate at several positions and suggested that the active site of the enzyme recognized five glucose units linked by a 1,3-1 bond. The association of the present endo-1,3-P-glucanase with the cell wall of A. fumigatus suggests a putative role for this enzyme during cell-wall morphogenesis.Keywords: endo-I ,3-P-glucanase ; cell wall : Aspergillus furnigutus; filamentous fungus.1 ,3-P-Glucanases are classified as exo-l,3-/$glucanase (1,3-P-glucan glucohydrolase) and endo-1,3-P-glucanase (1,3-1-glucan glucanohydrolase). Endoglucanase activities cleave inside a glucan chain in a more or less random fashion: while the exoglucanase activities release glucose residues from the non-reducing end. Many yeast and filamentous fungi produce both exocellular and cell-wall-associated exo-p-glucanases and endo-8-glucanases (Notario, 1982;Hien and Fleet, 1983 ;Copa-Patino, et al. 1989;Stahmann et al., 1993). 1,3-p-Glucan is a predominant polysaccharide of the fungal cell wall, and is thought to be responsible for the shape and rigidity of the wall (Cabib et al., 1982;Fleet, 1991). It has been suggested that 1,3-p-glucanase activities are essential to the continuous rearrangement of the wall 1,3-P-glucans during fungal growth (Cabib et al., 1988;Wessels, 1988). In the filamentous fungus Coprinus cinereus, endo-p-glucanases seem to play an essential role in stipe elongation (Kamada et al., 1985). In Sacchuronzyces cerevisiae, the production of exo-P-glucanases is growth-associated and cellcycle regulated (del Rey et al., 1979;Larriba et al., 1984;Jiang et al., 1995), suggesting that their activities are required at very specific stages during morphogenesis. For example, bud initiation, cell expansion, cell conjugation and sporulation are characterised by a higher activity of particular 1,3-~-glucanases (del Rey et al., 1982; Cenamor et al., 1987). However, gene disruption of yeast exo-p-glucanase activities does not modify the cellular growth (Larriba et al., 1995).Correspondence to T. Fontaine, lnstitut Pasteur, Laboratoire des Aspergillus, 25 rue du Docteur E. Roux, F-75724 Paris cedex 15, FranceAbbreviafions. Abz(OH)ONHNHZ, pamino-hydroxybenzoic acid hydrazide; Con-A, concanavalin A lcctin; dp, degree of polymerisation; Gn, oligosaccharide containing n 1,3-P-linked glucose residues ; G,,r, reduced oligosacchari...

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Section: Discussionmentioning

confidence: 99%

Purification and Characterization of an Endo‐1,3‐β‐Glucanase from Aspergillus fumigatus

Fontaine

Hartland

Beauvais

et al. 1997

European Journal of Biochemistry

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“…Their involvement in fungal cell wall formation during hyphal tip growth, as tightly balanced antagonists of glucan synthase, is controversial (Bartnicki-Garcia, 1968Wessels, 1988). 1,3-p-Glucanases have been studied in order to use them for various technical purposes such as preparing protoplasts (Kitamoto et al, 1987), analysing fungal cell wall structure (Usui et al, 1985), clarifying slimy must (Dubourdieu et al, 1985), and in the determination or degradation of barley pglucan, which causes problems if it accumulates during the brewing process (Bamforth, 1980). In most cases, a system of 1,3-P-glucanases rather than a single enzyme was found.…”

Section: Introductionmentioning

confidence: 99%

Purification and characterization of four extracellular 1,3- -glucanases of Botrytis cinerea

Stahmann

Schimz

Sahm

1993

Journal of General Microbiology

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~ ~~~The filamentous fungus Botvytis cinerea was found to produce four 1,3+glucanases (Glu I, II, 111, IV), These enzymes were visualized by activity-staining after separation by native polyacrylamide gel electrophoresis (PAGE). During growth on glucose as the single carbon source, only Glu III was detectable in the culture supernatant of B. cinerea. After glucose was exhausted from the medium, the extracellular (1,3)( 1,6)-/%~-glucan (cinerean) capsule of the fungus was degraded. In this phase the other three enzymes became detectable and the amount of all four enzymes increased. The enzyme with the greatest activity was Glu II, which was purified to homogeneity by SDS-PAGE. Its M, was 75000 and its isoelectric point (PI) was 5.2. Glycosylation of Glu I1 was shown by the periodic acid/Schiff reaction after SDS-PAGE. Glu I1 cleaved cinerean and laminarin. Both substrates were degraded in an exo-manner as shown by product characterization, and by studying the viscosity decrease in comparison with the liberation of reducing groups. The concentration of substrate that gave halfmaximal velocity (Se5) for Glu II was 580 pg ml-' for cinerean and 152 pg ml-' for laminarin. For Glu 111, also purified to homogeneity by SDS-PAGE, an M, of 84 000 and a PI of 3-6 were determined. Glu III cleaved laminarin (So.5 119 pg ml-') but not cinerean. Glu I and Glu IV were purified by activity-stained native PAGE. Both enzymes cleaved cinerean and laminarin in an exo-manner. Glu I focused at PI 4.9; its Se5 was 275 pg ml-' with cinerean and 138 pg ml-' with laminarin. The PI of Glu IV was 3.4; its So.5 was 171 pg ml-' for cinerean and 27 pg ml-' for lamina rin.

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“…The presence of 1,3-P-glucanases has been well documented in a variety of fungi and bacteria (Doi et al, 1973;Manners & Wilson, 1973;Martin et al, 1980;Tanaka & Phaff, 1965;Usui et al, 1985). B. circulans WL-12 was selected as the source of the genes encoding 1,3-P-glucanase for these studies for the following M .…”

Section: Introductionmentioning

confidence: 99%

“…Many micro-organisms, including B. circulans, which exhibit 1,3-P-glucanase activity, produce multiple enzyme species (Doi et al, 1973;Kobayashi et al, 1974a;Sietsma & Wessels, 1979;Skujins et al, 1965 ;Usui et al, 1985;Yamamoto et al, 1983). The isolation of clones which produce unique 1,3-P-glucanases would enable both the circumvention of the problems associated with purifying individual 1,3-P-glucanases from a complex mixture of similar activities and the opportunity to introduce these genes directly into plants for in vivo analysis of their efficacies.…”

Section: Introductionmentioning

confidence: 99%

Cloning of two genes from Bacillus circulans WL-12 which encode 1,3- -glucanase activity

Fiske

Tobey-Fincher²,

Fuchs³

1990

Journal of General Microbiology

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Two genes encoding distinct 1,3-P-glucanases have been cloned from Bacillus circulms and expressed in Escherichiia cofi. A cosmid library of B. circulans WL-12 DNA was constructed in the broad-host-range cosmid pLAFR1 and screened in E. coli for clones which exhibited 1,3-P-glucanase activity. Two 1,3-P-glucanase-positive clones were identified which contained genes encoding two independent 1,3-P-glucanases as shown by biochemical, physical and molecular analyses. The cosmids, designated pMON5401(27.1 kb insert) and pMON5402 (24.7 kb insert), encoded 68 kDa and 40 kDa 1,3-P-glucanases, respectively. Both 1,3-P-glucanases were purified from their respective E. coli strains, characterized biochemically, and were shown to exhibit lytic activity against purified yeast cell wall preparations.

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Purification of endo-(1→3)-β-D-glucanases lysing yeast cell walls from Rhizoctonia solani

Cited by 16 publications

References 23 publications

Purification and Characterization of an Endo‐1,3‐β‐Glucanase from Aspergillus fumigatus

Purification and Characterization of an Endo‐1,3‐β‐Glucanase from Aspergillus fumigatus

Purification and characterization of four extracellular 1,3- -glucanases of Botrytis cinerea

Cloning of two genes from Bacillus circulans WL-12 which encode 1,3- -glucanase activity

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