Fig. 6.

ExbD cysteine substitutions share common interfaces between homodimeric and heterodimeric interactions. TCA-precipitated samples of strains expressing chromosomally encoded wild-type ExbD and TonB (W3110) or a ΔexbD ΔtolQR ΔtonB strain (KP1509) coexpressing plasmid-encoded ExbD Cys substitutions and TonB(C18G, A150C) near native levels were resolved on nonreducing or reducing 13% and 11% SDS-polyacrylamide gels and immunoblotted with ExbD-specific polyclonal antibodies or TonB-specific monoclonal antibodies. Reduced and nonreduced samples came from the same culture. The positions of the monomeric proteins or disulfide-cross-linked complexes are indicated on the left. L93C, T94C, D107C, D111C, and D120C ExbD-TonB heterodimers were detected on longer exposures (data not shown). The positions of nonreducing molecular mass standards are indicated between the immunoblots. The strain designations across the top apply to both immunoblots below. All samples were processed on the same day.