Prokaryotic expression and bioactivity analysis of insulin-like growth factor-I from Cynoglossus semilaevis Güther

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Prokaryotic expression and bioactivity analysis of insulin-like growth factor-I from Cynoglossus semilaevis Güther
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Affiliation:1. Key Laboratory of Sustainable Development of Marine Fisheries, Ministry of Agriculture, Qingdao Key Laboratory for Marine Fish Breeding and Biotechnology, Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Qingdao 266071, Chi
Clc Number:S965; Q959
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Güther to clone the mature peptide domain of the gene. The mature peptide consisted of 70 amino acids, including four functional domains B-C-A-D. The mature peptide fragment was subcloned into the prokaryotic expression vector pET-28a to construct a recombinant plasmid that was transformed into BL21 (DE3). Induction of the plasmid by IPTG yielded a special fusion polypeptide containing 6 at the N-terminus. SDS-PAGE analysis revealed that the IGF-I polypeptide had a molecular weight of 11.4 kD. The recombinant IGF-I protein proximately accounted for 58.5% of the whole bacterial protein 3 h post induction with IPTG. Thus,. Western-blotting analysis suggested the fusion polypeptide exhibited antigenicity to 6 antibodies. The IPTG-induced bacterial precipitate was denaturalized using 6 mol/L guanidine HCl, purified using Ni-NTA affinity chromatography, and annealed by gradient dialysis in urea, thereby yielding the purified protein. The recombinant IGF-I protein promoted the proliferation of breast cancer MDA231 cells, suggesting it is biologically active. Our results provide a basis and suggested methods for regulating the growth of farmedGüther.

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刘芝亮,徐永江,柳学周,史宝,王妍妍. 半滑舌鳎类胰岛素生长因子-Ⅰ的原核表达及活性分析[J]. Jounal of Fishery Sciences of China, 2013,[volume_no](4):706-712

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Online: July 17,2013
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