Synthesis and high-expression of chitin deacetylase from Colletotrichum lindemuthianum in Pichia pastoris GS115
首发时间:2012-05-23
Abstract:A gene, ClCDA, encoding chitin deacetylase from Colletotrichum lindemuthianum, was optimized according to the codon usage bias of Pichia pastoris and synthesized in vitro by overlap extension PCR. It was secretorily expressed in P. pastoris GS115 using constitutive expression vector pHMB905A. The expression level reached the highest with 110 mg/l culture supernatant after 72 h methanol induction, which comprises of 77.27 U/mg chitin deacetylase activities. SDS-PAGE, mass spectrometry and deglycosylation assays demonstrated that part of recombinant protein was glycosylated with an apparent molecular mass of 33 kDa. The amino acid sequences of recombinant proteins were confirmed by mass spectrometry.
keywords: Chitin deacetylase Colletotrichum lindemuthianum high-expression Pichia pastoris Synthesis
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Synthesis and high-expression of chitin deacetylase from Colletotrichum lindemuthianum in Pichia pastoris GS115
摘要:A gene, ClCDA, encoding chitin deacetylase from Colletotrichum lindemuthianum, was optimized according to the codon usage bias of Pichia pastoris and synthesized in vitro by overlap extension PCR. It was secretorily expressed in P. pastoris GS115 using constitutive expression vector pHMB905A. The expression level reached the highest with 110 mg/l culture supernatant after 72 h methanol induction, which comprises of 77.27 U/mg chitin deacetylase activities. SDS-PAGE, mass spectrometry and deglycosylation assays demonstrated that part of recombinant protein was glycosylated with an apparent molecular mass of 33 kDa. The amino acid sequences of recombinant proteins were confirmed by mass spectrometry.
关键词: Chitin deacetylase; Colletotrichum lindemuthianum; high-expression; Pichia pastoris; Synthesis
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Synthesis and high-expression of chitin deacetylase from Colletotrichum lindemuthianum in Pichia pastoris GS115
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