title: Expression, activation and characterization of porcine trypsin in Pichia pastoris GS115.
abstract: Trypsin is a typical member of serine protease families, specifically cleaving the carboxyl group of peptides at the basic amino acids arginine and lysine. The gene fragment of porcine trypsin with its propeptide coding sequence was optimized and synthesized according to the codon usage bias of Pichia pastoris. The optimized sequence was integrated into the genome of P. pastoris GS115 using the vector pHBM905A. The yield of the recombinant protein was 0.48mg/ml with a maximum activity of 19.2U/ml after 96-h induction in a 5-l fermenter. An optimum activity for the recombinant trypsin was observed at 35°C and pH 8.5. This is the first time to express the porcine trypsinogen with P. pastoris expression system. This report also found that the propeptide was cleaved from the recombinant protein and the enzymogen was transferred into trypsin at the later phase of the fed-batch cultivation. In particular, the activation process can be initiated by changing pH.
authors.affiliation: Hubei Collaborative Innovation Center for Green Transformation of Bio-resources, Hubei Key Laboratory of Industrial Biotechnology, College of Life Sciences, Hubei University, Wuhan, People's Republic of China.
authors.forename: Min
authors.initials: M
authors.lastname: Shu
authors.affiliation: Hubei Collaborative Innovation Center for Green Transformation of Bio-resources, Hubei Key Laboratory of Industrial Biotechnology, College of Life Sciences, Hubei University, Wuhan, People's Republic of China.
authors.forename: Wei
authors.initials: W
authors.lastname: Shen
authors.affiliation: Hubei Collaborative Innovation Center for Green Transformation of Bio-resources, Hubei Key Laboratory of Industrial Biotechnology, College of Life Sciences, Hubei University, Wuhan, People's Republic of China.
authors.forename: Xiaojuan
authors.initials: X
authors.lastname: Wang
authors.affiliation: Hubei Collaborative Innovation Center for Green Transformation of Bio-resources, Hubei Key Laboratory of Industrial Biotechnology, College of Life Sciences, Hubei University, Wuhan, People's Republic of China.
authors.forename: Fei
authors.initials: F
authors.lastname: Wang
authors.affiliation: Hubei Collaborative Innovation Center for Green Transformation of Bio-resources, Hubei Key Laboratory of Industrial Biotechnology, College of Life Sciences, Hubei University, Wuhan, People's Republic of China. Electronic address: malixing@hubu.edu.cn.
authors.forename: Lixin
authors.initials: L
authors.lastname: Ma
authors.affiliation: Hubei Collaborative Innovation Center for Green Transformation of Bio-resources, Hubei Key Laboratory of Industrial Biotechnology, College of Life Sciences, Hubei University, Wuhan, People's Republic of China. Electronic address: chaozhai@hotmail.com.
authors.forename: Chao
authors.initials: C
authors.lastname: Zhai
mesh_terms: D000818:Animals; D005285:Fermentation; D006863:Hydrogen-Ion Concentration; D010843:Pichia / Q000235:genetics*; D011994:Recombinant Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D013552:Swine; D013696:Temperature; D014357:Trypsin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
chemical_list: D011994:Recombinant Proteins; D014357:Trypsin
keywords: Autocatalysis; High-density fermentation; High-level expression; Pichia pastoris; Porcine trypsin