Figure 3.

The contribution of amino acid residues in multiple regions of the MP-tmRNA tag to degradation by Lon. (A) Reporter substrates carrying single aspartic acid substitution in key signaling motifs in MP-tmRNA tag were subjected to proteolysis by MP-Lon₆ (100 nM) and analyzed as described in Fig. 2. Each experiment was repeated at least three times and the graph represents mean +/− standard deviation of three independent repeats. (B) The role of the C-terminal four residues of MP-tmRNA tag in degradation by MP-Lon. λ-cI-N-ssrA_MP reporter variants, with the C-terminal residues of the MP-tmRNA tag altered to either –DADA or –YDFD, were analyzed in the in vitro proteolysis assay by MP-Lon₆ (100 nM), as described in Fig. 2. (C) The effect of single aspartic acid substitutions in key signaling motifs of MP-tmRNA tag on degradation of the λ-cI-N-ssrA_MP reporter by EC-Lon. Reporter substrates carrying single aspartic acid substitution in key signaling motifs in MP-tmRNA tag were subjected to proteolysis by EC-Lon₆ (400 nM) and analyzed as described in Fig. 2. Each experiment was repeated at least three times and the graph represents mean +/− standard deviation of three independent repeats.