硫氧还蛋白介导抗菌肽CM4在大肠杆菌中多基因融合表达。
TrxA mediating fusion expression of antimicrobial peptide CM4 from multiple joined genes in Escherichia coli.
作者信息
Zhou Liangfan, Zhao Zhihui, Li Baocun, Cai Yufeng, Zhang Shuangquan
机构信息
Jiangsu Province Key Laboratory for Molecular and Medical Biotechnology, Life Sciences College, Nanjing Normal University, Nanjing 210046, China.
出版信息
Protein Expr Purif. 2009 Apr;64(2):225-30. doi: 10.1016/j.pep.2008.11.006. Epub 2008 Dec 3.
Antimicrobial peptide CM4, a small cationic linear alpha-helical peptide that consists of 35 amino acids, was isolated from Bombyx mori. To improve the expression level of CM4 in Escherichia coli, tandem repeats of CM4 gene were constructed and expressed as fusion proteins (TrxA-nCM4, n=1, 2, 3,...,8) by constructing the vectors of pET32-nCM4 (n=1, 2, 3,...,8). Comparison among the expression levels of soluble fusion protein TrxA-nCM4 (n=1, 2, 3,...,8) suggested that BL21 (DE3)/pET32-3CM4 was an ideal recombinant strain for CM4 production. Under the selected conditions of cultivation and isopropylthiogalactoside (IPTG) induction, the expression level of CM4 was as high as 68mg/l with about 21% of fusion protein in soluble form, which was the highest yield of CM4 reported so far.
抗菌肽CM4是一种由35个氨基酸组成的小型阳离子线性α-螺旋肽，从家蚕中分离得到。为提高CM4在大肠杆菌中的表达水平，构建了CM4基因的串联重复序列，并通过构建pET32-nCM4（n = 1, 2, 3, ..., 8）载体将其表达为融合蛋白（TrxA-nCM4，n = 1, 2, 3, ..., 8）。可溶性融合蛋白TrxA-nCM4（n = 1, 2, 3, ..., 8）表达水平的比较表明，BL21 (DE3)/pET32-3CM4是生产CM4的理想重组菌株。在选定的培养条件和异丙基硫代半乳糖苷（IPTG）诱导下，CM4的表达水平高达68mg/l，约21%的融合蛋白为可溶形式，这是迄今为止报道的CM4的最高产量。
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