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Synthesis and high-expression of chitin deacetylase from Colletotrichum lindemuthianum in Pichia pastoris GS115
Kang Lixin 1,Chen Xiaomei 2,Zhai Chao 3 #,Ma Lixin 3 *
1.College of Life Sciences, Hubei University, WuHan 430062
2.State Key Laboratory of Genetic Engineering, School of Life Sciences, Fudan University, Shanghai, 200433
3.College of Life Sciences, Hubei University, Wuhan , 430062
*Correspondence author
#Submitted by
Subject:
Funding: The Research Fund for the Doctoral Program of Higher Educatio(No.No.20104208120005), the National Sciences Foundation of China (No.No.31100057 )
Opened online:23 May 2012
Accepted by: none
Citation: Kang Lixin,Chen Xiaomei,Zhai Chao.Synthesis and high-expression of chitin deacetylase from Colletotrichum lindemuthianum in Pichia pastoris GS115[OL]. [23 May 2012] http://en.paper.edu.cn/en_releasepaper/content/4478455
 
 
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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