Use of E. coli for the Production of a Single Protein

Abstract

E. coli has been widely used for recombinant protein production. Here, we introduce a novel expression method in E. coli, the Single Protein Production (SPP) system, in which E. coli is converted into a bioreactor producing only the target protein. In the SPP system, all E. coli cellular mRNAs are eliminated by the induction of MazF, an ACA-specific mRNA interferase, which results in complete cell growth arrest. However, the mRNA for a target protein was engineered to be devoid of ACA sequences, thus resistant to MazF cleavage. Therefore, the SPP system is unique and ideal for expression of toxic proteins and incorporation of toxic amino acid analogues. We have also demonstrated that the SPP system is a cost-effective protein production method for NMR structural studies because cell culture can be highly condensed without affecting protein yields.