HDX-MS performed on BtuB in E. coli outer membranes delineates the luminal domain’s allostery and unfolding upon B12 and TonB binding

Abstract

Significance TonB - dependent transporters such as BtuB are found in the outer membranes of gram-negative bacteria. They import scarce nutrients essential for growth, such as B12, the substrate of BtuB. Many transport steps remain enigmatic. Recent studies have emphasized force-mediated unfolding or the breakage of the “Ionic Lock,” a moiety far from the B12 binding site. A strong dependence on the membrane environment has been noted. Accordingly, we measured hydrogen exchange on BtuB still embedded in native outer membranes and found that B12 binding is sufficient to break the Ionic Lock. The amino terminus then extends into the periplasm to bind TonB, but we find no evidence of pore formation, which likely requires energy transduction from the inner membrane by TonB.