Suppression of the toxicity of Bac7 (1-35), a bovine peptide antibiotic, and its production in E. coli.
Suppression of the toxicity of Bac7 (1-35), a bovine peptide antibiotic, and its production in E. coli.
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DOI：
10.1186/s13568-016-0190-3
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发表时间：
2016-03
期刊：
影响因子：
3.7
通讯作者：
Inouye M
中科院分区：
文献类型：
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作者：
Ishida Y;Inouye M
Bac7(1-35)是一种富含Arg和Pro的多肽抗生素，在牛细胞中生产，以保护它们免受微生物感染。它已被证明能抑制大肠杆菌中的蛋白质合成，导致细胞死亡。由于其毒性，目前还没有开发出在大肠杆菌中生产Bac7的经济有效的方法来潜在的临床应用。在此，我们找到了一种抑制Bac7(1-35)在大肠杆菌中的毒性的方法，以建立其高效表达系统。在该系统中，Bac7(1-35)与粘球菌主要的孢子壳蛋白S的C端融合，该蛋白是一种含有Xa因子裂解位点的连接子。所得到的His6-PrS2-Bac7(1-35)(PrS2由两个串联的S蛋白的N端半结构域组成)在低温下通过单蛋白生产系统得到了良好的表达，并通过镍柱一步纯化。在低温条件下，融合蛋白S及其在SPP系统中的表达似乎抑制了Bac7(1-35)的毒性。纯化的His6-PrS2-Bac7(1-35)和经因子Xa处理的His6-PrS2-Bac7(1-35)均被证明是一种有效的无细胞蛋白质合成抑制因子。
Bac7 (1–35) is an Arg- and Pro-rich peptide antibiotic, produced in bovine cells to protect them from microbial infection. It has been demonstrated to inhibit the protein synthesis in E. coli, leading to cell death. Because of its toxicity, no cost effective methods have been developed for Bac7 production in Escherichia coli for its potential clinical use. Here, we found a method to suppress Bac7 (1–35) toxicity in E. coli to establish its high expression system, in which Bac7 (1–35) was fused to the C-terminal end of protein S, a major spore-coat protein from Myxococcus xanthus, using a linker containing a Factor Xa cleavage site. The resulting His6-PrS2-Bac7 (1–35) (PrS2 is consisted of two N-terminal half domains of protein S connected in tandem) was well expressed using the Single-Protein Production (SPP) system at low temperature and subsequently purified in a single step by using a Ni column. The combination of protein S fusion and its expression in the SPP system at low temperature appeared to suppress Bac7 (1–35) toxicity. Both the purified His6-PrS2-Bac7 (1–35) and His6-PrS2-Bac7 (1–35) treated by Factor Xa were proven to be a potent inhibitor for cell-free protein synthesis.
DOI：
10.1007/s10969-009-9072-0
发表时间：
2009-12
期刊：
Journal of structural and functional genomics
影响因子：
--
作者：
Mao, Lili;Tang, Yuefeng;Vaiphei, S Thangminlal;Shimazu, Tsutomu;Kim, Sung-Gun;Mani, Rajeswari;Fakhoury, Elias;White, Eileen;Montelione, Gaetano T;Inouye, Masayori
通讯作者：
Inouye, Masayori