Expression of an Endo-β-1,4-glucanase Gene from Orpinomyces PC-2 in Pichia pastoris

International Journal of Molecular Sciences

Xin Jin¹  Nan Meng¹ 

[1] Department of Chemical Engineering and Bioengineering, Zhejiang University, Hangzhou 310027, China;

关键词: endo-β-1;    4-glucanase;    Pichia pastoris;    heterologous expression;    neutral cellulase;    induction medium;   

DOI  :  10.3390/ijms12053366

来源: mdpi

【 摘 要 】

The endo-β-1,4-glucanase gene celE from the anaerobic fungus Orpinomyces PC-2 was placed under the control of an alcohol oxidase promoter (AOX1) in the plasmid pPIC9K, and integrated into the genome of a methylotrophic yeast P. pastoris GS115 by electroporation. The strain with highest endo-β-1,4-glucanase activity was selected and designed as P. pastoris egE, and cultivated in shaking flasks. The culture supernatant was assayed by SDS-polyacrylamide gel electrophoresis and showed a single band at about 52 kDa. Furthermore, the recombinant P. pastoris egE was proved to possess the ability to utilize sodium carboxymethyl cellulose as a carbon source. The recombinant endoglucanase produced by P. pastoris showed maximum activity at pH 6.0 and temperature 45 °C, indicating it was a mesophilic neutral endo-β-1,4-glucanase, suitable for denim biofinishing/washing. Further research was carried out in suitable fermentation medium in shaking flasks. The most favorable methanol addition concentration was discussed and given as 1.0%. After methanol induction for 96 h, the endo-β-1,4-glucanase activity reached 72.5 IU mL⁻¹. This is the first report on expression and characterization of endo-β-1,4-glucanase from Orpinomyces in P. pastoris. The endo-β-1,4-glucanase secreted by recombinant P. pastoris represents an attractive potential for both academic research and textile industry application.

【 授权许可】

CC BY   
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