In vivo cleavage of solubility tags as a tool to enhance the levels of soluble recombinant proteins in Escherichia coli

Recombinant proteins are generally fused with solubility enhancer tags to improve the folding and solubility of the target protein of interest. However, the fusion protein strategy usually requires expensive proteases to perform in vitro proteolysis and additional chromatographic steps to obtain tag‐free recombinant proteins. Expression systems based on intracellular processing of solubility tags in Escherichia coli, through co‐expression of a site‐specific protease, simplify the recombinant protein purification process, and promote the screening of molecules that fail to remain soluble after tag removal. High yields of soluble target proteins have already been achieved using these protease co‐expression systems. Herein, we review approaches for controlled intracellular processing systems tailored to produce soluble untagged proteins in E. coli. We discuss the different genetic systems available for intracellular processing of recombinant proteins regarding system design features, advantages, and limitations of the various strategies. Recombinant protein production in different Escherichia coli strains is a cornerstone of modern biotechnology. Controlled intracellular processing (CIP) of recombinant passenger proteins that are fused to solubility enhancer proteins has been proposed as an alternative to obtaining satisfactory yields of soluble recombinant proteins without the need for significant post‐processing steps. In this manuscript, we review different genetic systems currently available for controlled intracellular processing of fusion recombinant proteins, regarding system design features, significant advantages, and limitations of the various strategies.