Regulation of the Escherichia coli HipBA Toxin-Antitoxin System by Proteolysis
Figure 3
Lon degradation of HipB in vitro.
0.6 µM His6-Lon and 0.48 µM His6-HipB were incubated in reaction buffer at 37°C (50 mM Tris-HCl (pH 8.0), 4 mM ATP, 7.5 mM MgCl2) for indicated times with or without the component specified and subjected to SDS-PAGE and silver staining followed by analysis (at least 3 independent experiments were used to calculate HipB turn over). (A) In vitro degradation of His6-HipB by His6-Lon. (B) ATP or MgCl2 were omitted in the assay. Closed squares, no ATP; open squares no MgCl2. (C) Addition of an oligodeoxynucleotide encompassing the 21 bp hip operator (closed squares) or control oligo (open squares) and (D) addition of His6-HipA (closed squares) or control protein (lysoszyme) (open squares) to the degradation assay.