Improvement in the Orthogonal Protein Degradation in Escherichia coli by Truncated mf-ssrA Tag

Abstract

SsrA peptide tag from Mycoplasma florum has been developed as a versatile biotechnology tool to control orthogonal degradation of tagged proteins in Escherichia coli. Here, using the systematic deletion mutants of mf-ssrA tag, we demonstrated that the residues in two separate regions have different functions in mf-Lon-mediated specific orthogonal target protein degradation in E. coli. The deletion of multiple residues, up to six amino acids, did not fatally abolish its specific degradation activity, instead of being able to improve the stability of the tagged protein in the presence of endogenous proteases before mf-Lon expression in E. coli. Except for previously identified essential residues, the region adjacent to the C-terminal of the mf-ssrA tag was involved in mf-Lon and endogenous protease-mediated degradation. Moreover, the deletion of specific residues made the mf-ssrA tag more effective and compact. The mf-ssrA tag can be implemented in synthetic biology and bioengineering for development of synthetic circuits.

Keywords

mf-ssrA / Protein degradation / Escherichia coli / L region / R region

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