A method of purification, identification and characterization of beta-glucosidase from Trichoderma koningii AS3.2774

作者：Lin, Yuanshan; Chen, Guiguang; Ling, Min; Liang, Zhiqun^*

来源：Journal of Microbiological Methods, 2010, 83(1): 74-81.

DOI：10.1016/j.mimet.2010.07.019

• beta-glucosidase
• Electroelution
• Native gradient-polyacrylamide gel
• electrophoresis
• Trichoderma koningii

In this study, we used native gradient-polyacrylamide gel electrophoresis and electroelution (NGGEE) to purify enzymatic proteins from Trichoderma koningii AS3.2774. With this method, we purified eight enzymatic proteins and classified them to the cellulase system by comparing secretions of T. koningii in inductive medium and in repressive medium. It resulted in 24-fold beta-glucosidase (BG) purification with a recovery rate of 5.5%, and a specific activity of 994.6 IU mg(-1) protein. The final yield of BG reached 8 mu g under purifying procedure of NGGEE We also identified BG using the enzyme assay with thin-layer chromatography and MALDI-TOFMS. This BG had one subunit with a molecular mass of 69.1 kDa as determined by sodium dodecylsulfate-polyacrylamide gel electrophoresis. The hydrolytic activity of the BG had an optimal pH of 5.0, an optimal temperature of 50 degrees C. an isoelectric point of 5.68 and a K-m for p-nitrophenyl-beta-D-glucopyranoside of 2.67 mM. Taken together, we show that NGGEE is a reliable method through which mu g grade of active proteins can be purified.

• 出版日期2010-10
• 单位广西大学; 湖南农业大学