Purification and Characterization of β-1,6-Glucanase of Streptomyces rochei Application in the Study of Yeast Cell Wall Proteins

Abstract

  A β-1,6-glucanase was purified to apparent homogeneity from a commercial yeast digestive enzyme prepared from Streptomyces rochei by a series of column chromatographies. The molecular mass of the purified enzyme was 60 kDa by SDS-PAGE. The purified enzyme had an optimum pH range from 4.0 to 6.0 and was stable in the same pH range. The enzyme was stable under 50°C but lost almost all activity at 60°C. The enzyme was specific to β-1,6-glucan and had little activity towards β-1, 3-glucan and β-1, 4-glucan. When the β-1,6-glucan was hydrolyzed with the purified enzyme for 5 h, the reaction products contained 20% glucose, 36% gentiobiose, and 44% other oligosaccharides, suggesting that the enzyme is an endo-type glucanase. When the purified enzyme was used for the digestion of the cell wall of Saccharomyces cerevisiae, cell-wall proteins covalently bound to the cell-wall glucan were recovered as soluble forms, suggesting that this enzyme is useful for analysis of yeast-cell wall proteins.