This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Membrane bound serine/threonine protein kinases (STPKs) are essential components of regulatory pathways in the pathogenic bacterium Mycobacterium tuberculosis, the causative agent of tuberculosis. Nine single-pass, transmembrane kinases are present in TB and these paralogous proteins share significant sequence homology (30-60%), indicating conservation of structure and mechanism. Structural analysis of these proteins is discovering important facts related to global regulation of kinase activity and possible mechanisms for the design of discriminating inhibitors. Our previous crystallographic models of kinase domains of PknE and PknB have implicated homodimerization as a mechanism for allosteric regulation of kinase activity. Structural studies of the enzymatic domains have provided clues towards the mechanism of dimerization, but models comprising a range of ligand-bound and catalytic states are necessary for complete understanding of dimerization and the catalytic cycle. Furthermore, the structure of the extracellular domain of PknE is required to demonstrate the function of this domain in dimerization and kinase regulation. I have crystallized the PknE kinase domain with bound nucleotide and the PknE sensor domain. I request beam time at SSRL to determine these structures which have the potential to reveal for the first time both the intracellular and extracellular domains of a bacterial receptor STPK.
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