title: Significant expression of a Chinese scorpion peptide, BmK1, in Escherichia coli through promoter engineering and gene dosage strategy.
abstract: Heterologous expression is an efficient alternative to conventional extraction to produce a specific Buthus martensii Karsch (BmK) peptide. In this work, BmK1 was successfully expressed in Escherichia coli after genetic codon optimization, but BmK1 content was <6% of total cellular protein. To improve BmK1 expression, a trc promoter library with a wide relative strength was constructed, and three promoters, PpJF136 (0.55), PpJF325 (1.29), and PpJF288 (2.31), were selected to control BmK1 expression. A higher BmK1 expression (>13.9% of total protein) was obtained using a stronger promoter, PpJF325 . Furthermore, a maximum BmK1 content (>21.7% of total protein) was obtained by combining promoter PpJF325 and three copies of the BmK1 gene. The yield of the purified BmK1 achieved 196.74 mg L(-1) in E. coli BL21(DE3) pJF431, which was improved 2.09-fold compared with the control. This was the highest reported production of scorpion peptides in E. coli.
authors.affiliation: Key Laboratory of Synthetic Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, People's Republic of China.|State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, People's Republic of China.
authors.forename: Jianfeng
authors.initials: J
authors.lastname: Wang
authors.affiliation: Key Laboratory of Synthetic Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, People's Republic of China.
authors.forename: Zhiqiang
authors.initials: Z
authors.lastname: Xiong
authors.affiliation: Key Laboratory of Synthetic Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, People's Republic of China.|State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, People's Republic of China.
authors.forename: Yingying
authors.initials: Y
authors.lastname: Yang
authors.affiliation: Key Laboratory of Synthetic Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, People's Republic of China.
authors.forename: Na
authors.initials: N
authors.lastname: Zhao
authors.affiliation: Key Laboratory of Synthetic Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, People's Republic of China.
authors.forename: Yong
authors.initials: Y
authors.lastname: Wang
mesh_terms: D000818:Animals; D004926:Escherichia coli / Q000235:genetics*; D018628:Gene Dosage / Q000235:genetics*; D005818:Genetic Engineering; D011401:Promoter Regions, Genetic / Q000235:genetics*; D012604:Scorpion Venoms / Q000235:genetics*; D012605:Scorpions / Q000235:genetics*
chemical_list: D012604:Scorpion Venoms; C477804:antitumor-analgesic peptide, Buthus martensii
keywords: BmK1; copy number; heterologous expression; promoter engineering; scorpion venom