Membrane Fusion Proteins of Type I Secretion System and Tripartite Efflux Pumps Share a Binding Motif for TolC in Gram-Negative Bacteria
Figure 3
Interaction between TolC and HlyD in vivo.
The in vivo interaction between HlyD and TolC was detected using a chemical cross-linking agent (DSP). E. coli BW25113ΔacrAB ΔtolC210::Tn10 cultures that coexpressed HlyA, HlyB, HlyC, hexa-His-tagged wild-type TolC, and c-Myc-tagged wild-type HlyD (WT) or one of the HlyD mutants (R186A, L190A, T197A and T197Y) are shown. All cultures were treated with (+) or without (−) DSP. Affinity-purified TolC and cross-linked proteins (HlyA and HlyD) were separated by SDS-PAGE and immunoblotted using monoclonal antibodies to His-tag and c-Myc, and polyclonal antibodies to HlyA.