title: High-level expression of human arginase I in Pichia pastoris and its immobilization on chitosan to produce L-ornithine.
abstract: BACKGROUND
  L-ornithine (L-Orn), is an intermediate metabolite in the urea cycle that plays a significant role in humans. L-Orn can be obtained from the catalysis of L-arginine (L-Arg) by arginase. The Pichia pastoris expression system offers the possibility of generating a large amount of recombinant protein. The immobilized enzyme technology can overcome the difficulties in recovery, recycling and long-term stability that result from the use of free enzyme.
  
  
  METHODS
  The recombinant human arginase I (ARG I) was obtained using an optimized method with the Pichia pastoris GS115 as the host strain. Chitosan paticles were cross-linked with glutaraldehyde and rinsed exhaustively. Then the expressed ARG I was immobilized on the crosslinked chitosan particles, and the enzymatic properties of both the free and immobilized enzymes were evaluated. At last, the immobilized ARG I was employed to catalyze L-Arg to L-Orn.
  
  
  RESULTS
  The results indicated that these two states both exhibited optimal activity under the same condition of pH10 at 40 °C. However, the immobilized ARG I exhibited the remarkable thermal and long-term stability as well as broad adaptability to pH, suggesting its potential for wide application in future industry. After a careful analysis of its catalytic conditions, immobilized ARG I was employed to catalyze the conversion of L-Arg to L-Orn under optimal condition of 1 % glutaraldehyde, 1 mM Mn(2+), 40 °C, pH10 and an L-arginine (L-Arg) concentration of 200 g/L, achieving a highly converted content of 149.g/L L-Orn.
  
  
  CONCLUSIONS
  In this work, ARG Ι was abundantly expressed, and an efficient, facile and repeatable method was developed to synthesize high-quality L-Orn. This method not only solved the problem of obtaining a large amount of arginase, but also provided a promising alternative for the future industrial production of L-Orn.
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, 430062, People's Republic of China. snowprincess1989@163.com.
authors.forename: Xue
authors.initials: X
authors.lastname: Zhang
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, 430062, People's Republic of China. 664617377@qq.com.
authors.forename: Jin
authors.initials: J
authors.lastname: Liu
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, 430062, People's Republic of China. kukuyu2012@hotmail.com.
authors.forename: Xianhong
authors.initials: X
authors.lastname: Yu
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, 430062, People's Republic of China. 448244821@qq.com.
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, 430062, People's Republic of China. li-yi@hotmail.com.
authors.forename: Li
authors.initials: L
authors.lastname: Yi
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, 430062, People's Republic of China. 598392746@qq.com.
authors.forename: Zhezhe
authors.initials: Z
authors.lastname: Li
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, 430062, People's Republic of China. 514944228@qq.com.
authors.forename: Yunyun
authors.initials: Y
authors.lastname: Liu
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, 430062, People's Republic of China. malixing@hubu.edu.cn.
authors.forename: Lixin
authors.initials: L
authors.lastname: Ma
mesh_terms: D001119:Arginase / Q000096:biosynthesis* / Q000235:genetics; D001120:Arginine / Q000378:metabolism; D048271:Chitosan / Q000737:chemistry; D004800:Enzymes, Immobilized / Q000737:chemistry / Q000235:genetics; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D009952:Ornithine / Q000096:biosynthesis*; D010843:Pichia / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000235:genetics
chemical_list: D004800:Enzymes, Immobilized; D011994:Recombinant Proteins; D048271:Chitosan; D001120:Arginine; D009952:Ornithine; C574619:ARG1 protein, human; D001119:Arginase