The E. coli lite strains: new expression strains for the production of toxic membrane proteins

The production of recombinant biopharmaceuticals is a billion-dollar industry, with numerous high-value proteins expressed in the bacterium Escherichia coli. Many recombinant protein production (RPP) systems enable the synthesis and purification of large amounts of soluble recombinant protein. However, such systems are often too strong to allow the expression of difficult-to-fold proteins, such as membrane proteins, with high-level RPP resulting in the production of insoluble aggregates (i.e., inclusion bodies), target protein degradation and even cell death. To aid the production of problematic proteins, we have engineered the E. coli lite strains in which the plasmid copy number regulator pcnB has been disrupted to allow expression levels of protein targets to be immediately lowered. Using the E. coli lite strains, we express a variety of proteins including green fluorescent protein (GFP), human growth hormone (hGH) and three toxic membrane proteins (BamA_ENm, LetB_EPEC and YiaM). We show that the E. coli lite strains are able to substantially lower the expression of target proteins and alleviate the cellular toxicity associated with producing membrane-localised proteins. Additionally, we show that the E. coli lite strains are extremely versatile, functioning with a variety of RPP systems, including lac-, T7 RNA polymerase- and paraBAD-based systems, and operate under different expression and growth regimes. Thus, the E. coli lite strains can be used to quickly lower expression levels of complicated target proteins to improve both protein production and cell growth.