Separation and Purification of an β-1,3-glucanase from Trichoderma Strain

Objective:The purification methods of β-1,3-glucanase from Trichoderma strain LE02 were investigated.Methods: Crude enzyme was precipitated by ammonium sulfate,ethanol and acetone respectively,and then purified by DEAE-Sepharose CL-6B anion-exchange chromatography.Molecular weight of the purified β-1,3-glucanase was determined by SDS-PAGE.Results: Two-step salting-out using ammonium sulfate was better than ethanol or acetone precipitation.Crude enzyme was salted-out by ammonium sulfate,followed by dialysed,concentrated with polyethylene 6 000,and further purified by DEAE-Sepharose CL-6B anion-exchange chromatography.The purified activity recovery of β-1,3-glucanase was 78.71%,and specific activity increased from 12.839U/mg to 689.932 U/mg.And the purified β-1,3-glucanase was a single band on both Native-PAGE and SDS-PAGE,with molecular weight of 80.137kDa.Conclusions: β-1,3-Glucanase of electrophoresis grade was obtained by ammonium sulfate two-step salting-out and anion-exchange chromatography,and its activity recovery was high.