Properties of Lasso Peptide Degradation via ATP-Dependent Protease, ClpXP

Abstract

One of the key limitations for small, peptide drugs is their susceptibility to degradation in the body. Lasso peptides offer a novel structure that exhibits protease resistant properties. This structure could be a key element in the development of stable, potent peptide therapeutics. This research aims to determine the way in which these lasso structures enable protease resistance. Through the addition of the 11 aa ssrA degradation tag to the lasso structure, we hoped to analyze the mechanism in which ClpXP is limited in degrading these peptides. The lasso—sfGFP fusion constructs containing C-terminal ssrA degradation tags were found to be susceptible to in vivo truncations during the 20-hour lasso maturation period. The absence of these truncations in the lasso—A1 construct appears to show a relationship between cell stress and in vivo truncations. The expression of the constructs led to viable substrates for analysis. The resulting reactions failed to show an active ClpXP complex and displayed no degradation of substrates.