Expression of A Xylanase from Thermobifide fusca in Pichia pastoris And Its Characterization Analysis

Abstract:

A codon-optimized mature peptide gene encoding a thermostable xylanase from Thermobifide fusca has been cloned into the expression plasmid pPIC9K and named as pPIC9K-xyl11M. The pPIC9K-xyl11M was linearized with SalⅠ and integrated into the genome of Pichia pastoris GS115 by electroporation. The recombinant P. pastoris GS115/xyl11M was screened by G418 and then was induced with methanol to express reXyl11M. The reXyl11M activity expressed by P. pastoris transformant reached 105.3 U/mL. The molecular weight of reXyl11M was estimated to be 34 000 by SDS-PAGE. The reXyl11M displayed the highest activity at 70 ℃ and pH 6.0. It was stable at a temperature range of 30~75 ℃，and at a pH range of 6.5~7.5. Co2+，Ba2+，Cu2+，Li+ had a strong enzyme activation effect，Fe3+ had obvious inhibitory effect，its activity was not significantly affected by other metal ions and EDTA. This revealed that reXyl11M was successfully expressed in P. pastoris and had good enzymatic characterizations.