纤维素酶
毕赤酵母
酵母
纤维素
酿酒酵母
细胞壁
化学
热稳定性
酶分析
生物化学
酶
核化学
基因
重组DNA
有机化学
作者
Baosheng Shi,Ke Xia,Hua Yu,Jing Xie,Yingmin Jia,Guo Runfang
出处
期刊:Journal of Microbiology and Biotechnology
[Journal of Microbiology and Biotechnology]
日期:2015-11-28
卷期号:25 (11): 1856-1862
被引量:6
标识
DOI:10.4014/jmb.1503.03029
摘要
In order to improve the stability of endoglucanase under thermal and acidic conditions, the endoglucanase gene was fused to the N-terminus of the Saccharomyces cerevisiae pir gene, encoding the cell wall protein PIR. The fusion gene was transformed into Pichia pastoris GS115 for expression. A resulting strain with high expression and high activity was identified by examining resistance to Geneticin 418, Congo red staining, and quantitative analysis of enzyme activity. SDS-PAGE analysis revealed that the endoglucanase was successfully displayed on the yeast cell surface. The displayed endoglucanase (DEG) showed maximum activity towards sodium carboxyl methyl cellulose at approximately 275 IU/g cell dry weight. DEG exhibited greater than 60% residual activity in the pH range 2.5-8.5, higher than free endoglucanase (FEG), which had 40% residual activity at the same pH range. The highest tolerated temperature for DEG was 70°C, much higher than that of FEG, which was approximately 50°C. Moreover, DEG showed 91.1% activity at 65°C for 120 min, while FEG only kept 77.8% residual activity over the same period. The half-life of DEG was 270 min at 65°C, compared with only 150 min for FEG. DEG could be used repeatedly at least three times. These results suggest that the DEG has broad applications as a yeast whole-cell biocatalyst, due to its novel properties of high catalytic efficiency, acid-thermal stabilities, and reusability.
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