Enhanced biochemical characteristics of β-glucosidase via adsorption and cross-linked enzyme aggregate for rapid cellobiose hydrolysis.

Abstract

With proper design, immobilization can be useful tool to improve the stability of enzymes, and in certain cases even their activity, selectivity, productivity and economic viability. An immobilized β-glucosidase (BGL, EC 3.2.1.21) through matrix adsorption and cross-linked enzyme aggregate (ad-CLEA) technology is presented in this work. After adsorption and precipitation, BGL was immobilized to poly(glycidyl methacrylate-co-ethylenedimethacrylate) (PGMA/EDMA) microparticles using glutaraldehyde as the cross-linker. Immobilized BGL exhibits lower apparent Km but much higher Vmax than that of the soluble enzyme, suggesting greater enzyme-substrate affinity and rapid velocity. Besides, ad-CLEA-BGL presents better thermostability retaining activity nearly 70% for 3h and approximately 50% for 5h at 70°C, high operational reusability remaining more than 90% activity after nine uses and excellent storage stability holding about 95% activity after 45days. Furthermore, the cellobiose is completely hydrolyzed within 1h with ad-CLEA-BGL, which is significantly more efficient than soluble enzyme (about 4h). Therefore, BGL was successfully immobilized on PGMA/EDMA microparticles with an ad-CLEA technology and the immobilization greatly enhances the biochemical characteristics. This work indicates promising application for ad-CLEA-BGL in utilizing agricultural remnants, bio-converting cellobiose to fermentable reducing sugar and ethanol production.