The bacterial cell envelope is critical for maintaining cell structure and providing a barrier against environmental stressors, necessitating diverse strategies for repairing and defending these layers from external threats. In Firmicutes, one widespread strategy is to use Bce modules - membrane protein complexes that unite a peptide-detoxifying ABC transporter with a stress response coordinating two-component system. These modules provide specific, front-line defense for a wide variety of antimicrobial peptides and small molecule antibiotics as well as coordinate responses for heat, acid, and oxidative stress. Because of these abilities, Bce modules play important roles in virulence and the development of antibiotic resistance in a variety of pathogens. Despite their importance, Bce modules are still poorly understood, with scattered functional data in only a small number of species.In this work, I investigated Bce module function from a structural biology angle, using cryo-electron microscopy to capture snapshots of the B. subtilis BceABRS bacitracin resistance system. Chapter 2 describes acquisition of structures of the BceAB transporter in multiple conformations, which revealed unique features of this unusual ABC transporter that may underlie removal of antimicrobial peptides from the cell membrane. This work was followed up by studies of the full BceAB-S transporter-histidine kinase complex (Chapter 3) that revealed how the two halves of the membrane-bound portion of Bce module interact and regulate each other’s function. While many questions remain about Bce module function, the structures and biochemical assays in these studies provide a framework with which to interpret earlier mutational studies and reveal new mechanistic details of Bce-mediate antimicrobial peptide resistance. Finally, Chapters 1 and 4 are adapted from a review that provides a much-needed synthesis of functional data on Bce modules across all the species in which they have been studied. Understanding these multifunctional membrane complexes will enhance our understanding of bacterial stress sensing and may point toward new therapeutic targets for highly resistant pathogens.
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