title: A novel expression vector for the improved solubility of recombinant scorpion venom in Escherichia coli.
abstract: Recombinant scorpion anti-excitation peptide (rANEP) has previously been expressed using the pET32a system and purified via affinity chromatography. However, rANEP is expressed in BL21(DE3) cells as an inclusion body, and the affinity tag can not be removed. To overcome this problem, we used a variety of protein, DsbA, MBP, TrxA, intein, and affinity tags in fusion and co-expression to achieve soluble and functional rANEP without any affinity tag. In the pCIT-ANEP expression vector, the highest soluble expression level was approximately 90% of the total cellular proteins in E. coli, and the rANEP was cleaved by the intein protein and subsequently purified to obtain rANEP, which had the same activity as the natural ANEP. The purity of rANEP obtained using this method was over 95%, with a quantity of 5.1 mg from of purified rANEP from 1 L of culture. This method could expand the application of the soluble expression of disulfide-rich peptides in E. coli.
authors.affiliation: School of Life Science and Biopharmaceutics, Shenyang Pharmaceutical University, 103 Wenhua Road, Shenyang, Liaoning Province 110016, China.
authors.forename: Tianqing
authors.initials: T
authors.lastname: Han
authors.affiliation: School of Life Science and Biopharmaceutics, Shenyang Pharmaceutical University, 103 Wenhua Road, Shenyang, Liaoning Province 110016, China.
authors.forename: Hongyan
authors.initials: H
authors.lastname: Ming
authors.affiliation: School of Life Science and Biopharmaceutics, Shenyang Pharmaceutical University, 103 Wenhua Road, Shenyang, Liaoning Province 110016, China.
authors.forename: Lili
authors.initials: L
authors.lastname: Deng
authors.affiliation: School of Life Science and Biopharmaceutics, Shenyang Pharmaceutical University, 103 Wenhua Road, Shenyang, Liaoning Province 110016, China.
authors.forename: Hejie
authors.initials: H
authors.lastname: Zhu
authors.affiliation: School of Life Science and Biopharmaceutics, Shenyang Pharmaceutical University, 103 Wenhua Road, Shenyang, Liaoning Province 110016, China.
authors.forename: Zeyu
authors.initials: Z
authors.lastname: Liu
authors.affiliation: School of Life Science and Biopharmaceutics, Shenyang Pharmaceutical University, 103 Wenhua Road, Shenyang, Liaoning Province 110016, China.
authors.forename: Jinghai
authors.initials: J
authors.lastname: Zhang
authors.affiliation: School of Life Science and Biopharmaceutics, Shenyang Pharmaceutical University, 103 Wenhua Road, Shenyang, Liaoning Province 110016, China. Electronic address: songyongbo@syphu.edu.cn.
authors.forename: Yongbo
authors.initials: Y
authors.lastname: Song
mesh_terms: D000345:Affinity Labels; D004926:Escherichia coli / Q000737:chemistry* / Q000235:genetics / Q000502:physiology*; D024861:Genetic Enhancement; D005822:Genetic Vectors / Q000235:genetics*; D015202:Protein Engineering / Q000379:methods*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry; D012604:Scorpion Venoms / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics*; D012995:Solubility
chemical_list: D000345:Affinity Labels; D011994:Recombinant Proteins; D012604:Scorpion Venoms
keywords: ANEP; Analgesic activity; Buthus martensii Karsch; Intein; trxA