Beta-D-glycosidase activities of Humicola grisea: biochemical and kinetic characterization of a multifunctional enzyme.
A beta-D-glycosidase activity was purified from mycelium of Humicola grisea var. thermoidea grown on avicel as the main carbon source. The purified enzyme was a glycoprotein and migrated as a single polypeptide band on polyacrylamide gel electrophoresis under native or denaturing conditions. The apparent molecular weight of the enzyme was estimated to be 55 kDa by gel filtration and SDS-PAGE. The enzyme was active against o-nitrophenyl beta-D-galactoside; p-nitrophenyl beta-D-glucoside, p-nitrophenyl beta-D-fucoside, lactose and cellobiose, PNP fucoside (synthetic substrate) and cellobiose (natural substrate) being the best utilized. A comparison of the properties of beta-D-galactosidase, beta-D-glucosidase and beta-D-fucosidase showed that three activities exhibited similar pH and temperature optima and the same thermostability. The hydrolysis rate of substrate mixtures suggests that the enzyme possesses a common catalytic site for all the substrates assayed.
Peralta
R M
RM
Departamento de Biologia, Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, Brasil.
Terenzi
H F
HF
Jorge
J A
JA
D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D006026:Glycoside Hydrolases / Q000032:analysis / Q000302:isolation & purification / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007700:Kinetics; D003904:Mitosporic Fungi / Q000201:enzymology*; D008970:Molecular Weight; D011489:Protein Denaturation; D013379:Substrate Specificity; D013696:Temperature; D005644:alpha-L-Fucosidase / Q000378:metabolism; D001616:beta-Galactosidase / Q000378:metabolism; D001617:beta-Glucosidase / Q000378:metabolism
D006026:Glycoside Hydrolases; D001617:beta-Glucosidase; D001616:beta-Galactosidase; D005644:alpha-L-Fucosidase