Overexpression of GH3 β-Glucosidase from Aspergillus niger NL-1 in Pichia pastorls by Constructing Multi-Copy Gene and Using FM21 Medium

摘要

The β-glucosidase encoded by bgl1 of Aspergillus niger NL-1 is a particularly attractive candidate for some industrial purposes. In present study, we aimed to enhance the copy number of the bgl1 gene and optimize fermentation conditions using FM21 medium to increase the recombinant β-glucosidase production and decrease the costs in production of the enzyme. We constructed multi-copy bgl1 gene clones using high Zeocin concentration screening. The clone which can resist 3000 βg/mL Zeocin showed 2.7-fold higher β-glucosidase production than that of the parent strain. The optimal fermentation conditions of production for the β-glucosidase in shake flask cultivation using FM21 medium were determined as initial pH 6.0, agitation speed 240 rpm, adding concentration 1.0% (v/v) of methanol every 24 h, histidine 0.1% and PTM1 0.4% (v/v). The β-glucosidase activity was up to 114.1 U/mL on the optimal conditions after 216 h induction. Thus the report provides an industrial means to produce the β-glucosidase in P. pastoris.