High Expression of Rhizomucor miehei Lipase in Pichia pastoris by Fusion to CBD

Abstract

Abstract: Rhizomucor miehei lipase（RML）, one of the important enzymes applied in industries, has a broad prospect of application. CBD, which is capable of tightly binding cellulose, has been found in diverse hydrolytic enzymes（cellulose, xylanase）. The presence of a CBD is shown to increase the effective concentration of enzyme on substrates, thereby assisting the enhancement of hydrolytic enzyme activity. Here the CBD from Trichoderma harzianum endoglucanase II（THEGII）was connected to the N-terminal of RML through an endogenous linker peptide. Then the fusion genes were cloned into the secreted expression vector pPICZαA to construct recombinant plasmid pPICZαA-CBD-RML. The plasmid was linearized and transformed into Pichia pastoris GS115 to obtain Pichia pastoris recombinant strain. Compared with that of the mature RML without CBD-fusion, secretion of CBD-RML by this fusion construct was about 17% enhanced in addition to similar temperature activity and stability between RML and CBD-RML. Therefore, the CBD from THEGII can be used as a secretion enhancer for secretory production of heterologous protein in Pichia pastoris.

Key words: Pichia pastoris, RML, CBD, Secreted expression

Xue Chongchong, Zhang Junhui, Lin Ying, Han Shuangyan . High Expression of Rhizomucor miehei Lipase in Pichia pastoris by Fusion to CBD[J]. Biotechnology Bulletin, 2013, 0(2): 118-123.

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