FIG. 6.

(A) Testing YfgL-His₆ variants with a D223A, D227A, or D229A alteration for defective YaeT associations. Cells expressing wild-type (WT) YfgL-His₆ (YfgL_6His), YfgL(D223A)-His₆, YfgL(D227A)-His₆, or YfgL(D229A)-His₆ were incubated in the presence or absence of the cross-linker DSP prior to lysis under nondenaturing conditions. His-tagged YfgL complexes were precipitated by Penta-His antibody (α-His) and analyzed by Western blotting. YfgL-His₆, YaeT, and SurA were probed using HisProbe-HRP, anti-YaeT antibody, and anti-SurA antibody, respectively. (B) Residue S226 of the YfgL region at E221 to D229 comes in close contact with YaeT. Cells expressing wild-type YfgL-His₆ or YfgL(S226C)-His₆ were incubated with or without the cysteine-directed cross-linker SPDP before they were lysed under denaturing conditions. YfgL-His₆ and conjugated proteins were purified with an Ni-NTA spin column and subjected to Western analysis. HisProbe-HRP and YaeT antibody were used to probe YfgL-His₆ and YaeT.