FIGURE 5.

Stalled intermediates interact with BamA and BamD during OM translocation. TOP10 cells expressing empty vector (EV), wild-type Pet, PetSB, Pet48aa, Pet1HA, and Pet1HA-FLAG were harvested, spheroplasted, and lysed. Proteins were co-purified with anti-Pet passenger domain antibody, separated on a gradient 4–12% Bis-Tris gel in MES buffer, and analyzed by Western immunoblotting using anti-Pet passenger, anti-BamA (A), or anti-BamD (B) antibodies. A whole cell lysate (WCL) was included as an additional control for BamD. Generally, only the three high molecular weight stalled intermediates (Pet48aa, Pet1HA, and Pet1HA-FLAG) co-purify with the BAM proteins. M, molecular mass markers.