Relatively little is known about the cell division machinery, or divisome, of Mycobacteria tuberculosis (Mtb), the causative agent of tuberculosis (TB). There have been no new TB drug classes developed since a multidrug 6-month treatment course was introduced in the 1970s. New therapies are desperately needed for TB as the occurrence of multi-drug and extensive drug-resistant TB has increased rapidly in the past decade. The goal of this project is to advance knowledge of the Mtb divisome as potential drug targets and towards this end achieve an understanding of how the divisome proteins interact with each other and hence, how they are recruited to the divisome. Four transmembrane proteins are targeted for structural characterizations, both individually and as complexes.
The Specific Aims are: (1) characterize nascent and binding-induced structures of intrinsically disordered regions in the protein targets; (2) determine full-length structures of these proteins i lipid bilayers; (3) determine structures of complexes formed between these membrane proteins and with other divisome proteins and peptidoglycan precursors or fragments. Combining expertise in microbiology and structural biology with membrane protein and computational biophysics, the research will lead to critical knowledge on the activities and interactions of central layers in the divisome. An array of tools will be used, including bacterial two-hybrid assays, solid state and solution NMR, and restrained molecular dynamics. To meet the challenge of characterizing the proteins and protein complexes in a native-like, lipid bilayer environment, new structural methodologies, including a novel type of structural restraints from solid state NMR and a novel use of solid state NMR for conformational characterization of intrinsically disordered regions in membrane proteins, will be developed. This interdisciplinary team, with significant and unique experiences of the individuals and through prior joint publications, is ideally positioned to accomplish the proposed studies. These structural characterizations of the Mtb divisome and their functional implications will generate novel therapeutic opportunities for TB. The technologies developed here will represent the frontier of membrane protein structure biology. 1
Mycobacterium tuberculosis is the causative agent of TB that results in 1.2 million deaths per year worldwide as there have been no new TB drug classes developed since the 1970s and the occurrence of multi-drug and extensive drug-resistant TB has increased rapidly in the past decade. Uniquely, M. tuberculosis can enter a latent stage, where the bacterium greatly reduces the cell division rate and consequently there is interest not only in preventing the bacterium from replicating, but also interest in inducing cell division so that antibiotics can kill the bacterium. Through structural characterizations of four key proteins n the cell division apparatus, the divisome, as well as their interactions, the proposed research wil lay the foundation for developing new targets for TB therapies.
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