Purification and Enzymatic Properties of a-Galactosidase from Pycnoporus cinnabarinus

抄録

α-Galactosidase (EC 3.2. 1.22) from Pycnoporus cinnabarinus was purified by precipitation with ammoniumsulfate, column chromatographies on DEAE-SephadexA-50, Sephadex G-100, CMS ephadex C-50, and SP-Sephadex C-50, and isoelectric focusing. The purified enzyme was homogeneous on polyacrylamide disc gel electrophoresis, and the molecular weight was estimated to be about 210, 000 by gel filtration on Sephacryl S-200 and about 52, 000 by sodium dodecyl sulfate gel electrophoresis. The enzyme exhibited the optimum pH at 5.0 and was stable between pH 3 and 9. The optimumtemperature of the enzyme was 75°C. The enzymewas thermostable at pH 5.0 and completely lost its activity after heating at 90°C or at pH 3.5. The Michaelis constants were 0.31 mM for p-nitrophenyl α-D-galactoside, 0.80mM for melibiose, 2.16mM for raffinose, and 1.15mM for stachyose. The α-galactosidase activity was strongly inhibited by Ag⁺, Hg⁺⁺, p-chloromercuribenzoate, galactose, and melibiose. The enzyme also seemed to catalyse a glycosyltransfer reaction.