Escherichia coli strains with precise domain deletions in the ribonuclease RNase E can achieve greatly enhanced levels of membrane protein production

Abstract

AbstractEscherichia coli is one of the most widely utilized hosts for production of recombinant membrane proteins (MPs). Bacterial MP production, however, is usually accompanied by severe toxicity and low‐level volumetric accumulation. In previous work, we had discovered that co‐expression of RraA, an inhibitor of the RNA‐degrading activity of RNase E, can efficiently suppress the cytotoxicity associated with the MP overexpression process and, simultaneously, enhance significantly the cellular accumulation of membrane‐incorporated recombinant MPs in bacteria. Based on this, we constructed the specialized MP‐producing E. coli strain SuptoxR, which can achieve dramatically enhanced volumetric yields of well‐folded recombinant MPs. Ιn the present work, we have investigated whether domain deletions in the E. coli RNase E, which exhibit reduced ribonucleolytic activity, can result in suppressed MP‐induced toxicity and enhanced recombinant MP production, in a manner resembling the conditions of rraA overexpression in E. coli SuptoxR. We have found that some strains encoding specific RNase E truncation variants can achieve significantly enhanced levels of recombinant MP production. Among these, we have found a single RNase E variant strain, which can efficiently suppress MP‐induced toxicity and achieve greatly enhanced levels of recombinant MP production for proteins of both prokaryotic and eukaryotic origin. Based on its properties, and in analogy to the original SuptoxR strain, we have termed this strain SuptoxRNE22. E. coli SuptoxRNE22 can perform better than commercially available bacterial strains, which are frequently utilized for recombinant MP production. We anticipate that SuptoxRNE22 will become a widely utilized host for recombinant MP production in bacteria.