Molecular cloning, sequencing, expression of Chinese sturgeon cystatin in yeast Pichia pastoris and its proteinase inhibitory activity
文献类型： 外文期刊
作者： Bai, Junjie 1 ; Ma, Dongmei 2 ; Lao, Haihua 2 ; Jian, Qing 3 ; Ye, Xing; Luo, Jianren; Xong, Xiaoyong; Li, Yingh 1 ;
作者机构： 1.CAFS, Pearl River Fisheries Res Inst, Key Lab Trop & Subtrop Fish Breeding & Cultivat, Guangzhou 510380, Guangdong, Peoples R China
2.CAFS, Pearl River Fisheries Res Inst, Key Lab Trop & Subtrop Fish Breeding & Cultivat, Guangzhou 510380, Guangdong, Peoples R China; Zhajiang Ocean Univ, Zhanjiang 520425, Peoples R China; Shanghai Fisheries Univ, Coll Fisheries, Shanghai 200090, Peoples R China; Jinan Univ, Coll Life Sci & Technol, Dept Biotechnol, Guangzhou 510632, Guangdong, Peoples R China
3.CAFS, Pearl River Fisheries Res Inst, Key Lab Trop & Subtrop Fish Breeding & Cultivat, Guangzhou 510380, Guangdong, Peoples R China; Zhajiang Ocean Univ, Zhanjiang
期刊名称：JOURNAL OF BIOTECHNOLOGY （ 影响因子：3.307； 五年影响因子：3.778 ）
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收录情况： SCI
摘要： Cystatin, a superfamily of cysteine proteinase inhibitor of cathepsins and other cysteine proteinases, is widely distributed in animal tissues and body fluids. Although considerable attention has been given to mammalian and avian cystatins, little is known about cystatins from other vertebrates. In this study, a cDNA coding for Chinese sturgeon (Acipenser sinensis) cystatin was isolated and characterized. The corresponding mature cystatin peptide cDNA is 336 nucleotides long and encodes a protein of 112 amino acids. Sequence comparison showed that the cloned cystatin was a homolog of the mammalian Family II cystatin. The cystatin cDNA of Chinese sturgeon was subcloned into yeast expression vector pPICZalphaA and transformed into Pichia pastoris GS115 strain. After methanol induction, SDS-PAGE analysis of the culture supernatant indicated that the yield of recombinant cystatin was about 215 mg/l medium supernatant in shaking-flask fermentation medium, accounting for 73.6% of the total supernatant secreted proteins. Our data also showed that the recombinant cystatin is active in inhibiting the protease activity of papain and cathepsin B. Heat stability of the recombinant cystatin was also measured.
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