Characterization of a recombinant thermostable β-glucosidase from Putranjiva roxburghii expressed in Saccharomyces cerevisiae and its use for efficient biomass conversion

摘要

A beta-glucosidase gene from Putranjiva roxburghii (PRGH1) was heterologously expressed in Saccharomyces cerevisiae to enable growth on cellobiose. The recombinant enzyme was secreted to the culture medium, purified and biochemically characterized. The enzyme is a glycoprotein with a molecular weight of similar to 68 kDa and exhibited enzymatic activity with beta-linked aryl substrates like pNP-Fuc, pNP-G1c, pNP-Ga1 and pNP-Cel with catalytic efficiency in that order. Significant enzyme activity was observed for cellobiose, however the enzyme activity was decreased with increase in chain length of glycan substrates. Using cellobiose as substrate, the enzyme showed optimal activity at pH 5.0 and 65 degrees C. The enzyme was thermostable up to 75 degrees C for 60 min. The enzyme showed significant resistance towards both glucose and ethanol induced inhibition. The recombinant S. cerevisiae strain showed advantages in cell growth, glucose and bio-ethanol production over the native strain with cellobiose as sole carbon source. In simultaneous saccharification and fermentation (SSF) experiments, the recombinant strain was used for bio-ethanol production from two different cellulosic biomass sources. At the end of the SSF, we obtained 9.47 g L-1 and 14.32 g L-1 of bio-ethanol by using carboxymethyl cellulose and pre-treated rice straw respectively. This is first report where a beta-glucosidase gene from plant origin has been expressed in S. cerevisiae and used in SSF.