Fig. 4.

Levels of β-galactosidase expressed from a σ^E-dependent lacZ reporter were used to assess the basal and OMP-stimulated RseA-cleavage activities of DegS and L3-loop mutants in vivo. Cytochrome b₅₆₂ with a YYF C-terminal tail (cyt-YYF) or no tail (cyt) was expressed from an arabinose-inducible promoter. Black and gray bars represent β-gal levels in the absence and presence of 0.2% arabinose, respectively. (A) Control experiments showed that arabinose-mediated expression of cyt-YYF but not cyt stimulate wild-type DegS activity. Expression of cyt-YYF did not stimulate catalytically inactive DegS^S201A. (B) β-gal levels with or without arabinose induction of cyt-YYF for DegS variants with different amino acids at residues 182–184. All variants in this panel support arabinose-induced activity at least fourfold higher than the basal activity of wild-type DegS (NPT). (C) DegS variants from the residue 182–184 library with arabinose-induced activity less than fourfold higher than the basal activity of wild-type DegS (NPT). Values in all panels are averages ± SEM (n = 2–8). DegS variants in B and C are designated by the one-letter code for the amino acids at residues 182, 183, and 184.