Expression of a glucose-tolerant beta-glucosidase from Humicola grisea var. thermoidea in Saccharomyces cerevisiae.

A beta-glucosidase gene (bgl4) from Humicola grisea var thermoidea was successfully expressed in Saccharomyces cerevisiae. The recombinant protein (BGL4(Sc)) was initially detected associated with yeast cells and later in the culture medium. BGL4(Sc) showed optimal pH and temperature of 6.0 and 40 degrees C, respectively, and an apparent molecular mass of 57 kDa. The enzyme showed activity against cellobiose and synthetic substrates, and was inhibited more than 80% by Fe2+, Cu2+, Zn2+, and Al3+. Using p-nitrophenyl-beta-D-glucopyranoside (pNPG) as substrate, BGL4(Sc) presented a V(max) of 6.72 micromol min(-1) mg total protein(-1) and a K (m) of 0.16 mM under optimal conditions. Most important, BGL4(Sc) is resistant to inhibition by glucose and the calculated K (i) value for this sugar is 70 mM. This feature prompts BLG4(Sc) as an ideal enzyme to be used in the saccharification process of lignocellulosic materials for ethanol production.