Suppression of the toxicity of Bac7 (1–35), a bovine peptide antibiotic, and its production in E. coli

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 His(6)-PrS(2)-Bac7 (1–35) (PrS(2) 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 His(6)-PrS(2)-Bac7 (1–35) and His(6)-PrS(2)-Bac7 (1–35) treated by Factor Xa were proven to be a potent inhibitor for cell-free protein synthesis.