FIG. 5.

RhuR-BlaM is localized to the inner membrane of 4169rifΔrhuR. Fe-stressed cultures of 4169rifΔrhuR(pDJM41, pRK415) (vector) and 4169rifΔrhuR(pDJM41, pAEK21) (RhuR-BlaM) were used to inoculate BHI broth supplemented with 100 μM EDDHA and 1 μM hemin. At stationary phase, cells were collected for isolation of total membranes, outer membranes, and cytoplasmic membranes. Ten micrograms of total protein from each membrane preparation was separated by SDS-PAGE using SDS-12% polyacrylamide gels. Western immunoblotting using a rabbit polyclonal anti-BlaM antiserum was used to detect immunoreactive proteins. The positions of the molecular mass markers (in kilodaltons) are indicated. Arrows indicate the position of the 37-kDa RhuR-BlaM and its major 25-kDa degradation product.