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AMB Express. 2012; 2: 14.
Published online Feb 29, 2012. doi:  10.1186/2191-0855-2-14
PMCID: PMC3306196
High level expression of bikunin in Pichia pastoris by fusion of human serum albumin
Xing-Hua Gou,1 Yu-Ying Liu,2 Qi-Lei Chen,1 Jian-Jun Tang,2 Da-Yu Liu,1 Liang Zou,1 Xiao-Yong Wu,1 and Wei Wangcorresponding author1,3
1Faculty of Bioindustry, Chengdu University, Waidong Shilingzhen, Chengdu, 610106, China
2Genekey Biotech. (Chengdu) Co., Ltd. Chengdu, 610041, China
3Meat Proceeding Key Lab. of Sichuan Province, 610106, China
corresponding authorCorresponding author.
Xing-Hua Gou: xinghuagou/at/cdu.edu.cn; Yu-Ying Liu: superliuyy/at/yahoo.com.cn; Qi-Lei Chen: cranberry1997/at/sina.com; Jian-Jun Tang: tangjianjun2006/at/126.com.cn; Da-Yu Liu: liudy1014/at/163.com; Liang Zou: zouliang/at/cdu.edu.cn; Xiao-Yong Wu: wuxiaoyong/at/cdu.edu.cn; Wei Wang: wangwei/at/cdu.edu.cn
Received December 21, 2011; Accepted February 29, 2012.
Abstract
Bikunin is a proteoglycan exhibiting broad-spectrum inhibitory activity against serine proteases and could potentially suppress tumor cell invasion and metastasis. Here, we have successfully expressed recombinant human bikunin (rh-bikunin) in Pichia pastoris and also established the purification procedure. Different fusion genes of h-UTI and domain I, domain I and domain II, domain I, domain II and domain III of human serum albumin (HSA) were inserted into expression vector pPICZαA. After expressed in shake flask, rh-bikunin was produced in an 30-L fermenter and purified by affinity chromatography and cation exchange chromatography. The final expression levels were 200 mg/L and we got totally 1.08 g (3650 IU/mg) of active purified rh-bikunin (purity is 98%) from 20 L of fermentation broth. The rh-bikunin consists of unique form with molecular masses of 25 kDa, and has the same N-terminals sequence as human native bikunin. This study provided a new method for high level expression of active rh-bikunin by using HSA as fusion parter.
Keywords: Human bikunin, Fusion expression, Human serum albumin, Pichia pastoris
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