Transition metal ions function as required cofactors for myriad metalloenzymes that catalyze a large diversity of biological reactions. These same metal ions, however, are cytotoxic in excess. As a result, all cells dedicate considerable regulatory machinery to maintaining the intracellular concentrations and types of metal ions, i.e., homeostasis, in order to quickly adapt to a changing microenvironment. Emerging evidence suggests that microbial metal ion homeostasis and resistance to toxicity defines a critical component in human host-bacterial pathogen interactions during infection. This proposal seeks continuing support for a project whose long-term goal is to understand the molecular mechanisms of this adaptive response, including how metal ions are sensed by metalloregulatory or """"""""metal sensor"""""""" proteins, and trafficked in the cell, and how perturbations in metal metabolism impact bacterial pathogenesis and virulence. For the next project obacterium tuberculosis and Streptococcus pneumoniae.
Our specific aims are to: 1) Elucidate the structural and thermodynamic basis of Zn-regulation by S. aureus CzrA, a model ArsR-family repressor of the czr (chromosomal zinc-regulated) operon;2) Employ protein ligation strategies to site-specifically incorporate amino acid analogs to directly test our allosteric coupling model for CzrA;3) Use a panel of CzrA mutants to critically evaluate the functional relationship between KZn, the Zn binding affinity of a metal sensor, and the ability to sense zinc in the cell;4) Elucidate the structural determinants of DNA binding and Cu-mediated allosteric regulation of DNA binding by the novel Cu-sensor M. tuberculosis CsoR, the repressor of the Cu-effluxing copper-sensitive operon (cso);and 5) Structurally characterize S. pneumoniae AdcR, a postulated novel Zn-sensor that regulates the expression of a zinc uptake system (adc operon) required for adhesion competence and virulence in pneumohost-bacterial pathogen interactions during infection. The research outlined in this proposal seeks continuing support of a project designed to understand the structure and mechanism of the intracellular regulators of this process, termed """"""""metal sensor"""""""" or metalloregulatory proteins, that control the expression of genes linked to this adaptive response. Completion of these studies may ultimately lead to novel strategies for the targeted development of antimicrobial agents that exploit or induce misregulation of metal homeostasis.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Research Project (R01)
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Macromolecular Structure and Function A Study Section (MSFA)
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Anderson, Vernon
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Indiana University Bloomington
Schools of Arts and Sciences
United States
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Glauninger, Hendrik; Zhang, Yifan; Higgins, Khadine A et al. (2018) Metal-dependent allosteric activation and inhibition on the same molecular scaffold: the copper sensor CopY from Streptococcus pneumoniae. Chem Sci 9:105-118
Capdevila, Daiana A; Braymer, Joseph J; Edmonds, Katherine A et al. (2017) Entropy redistribution controls allostery in a metalloregulatory protein. Proc Natl Acad Sci U S A 114:4424-4429
Martin, Julia E; Lisher, John P; Winkler, Malcolm E et al. (2017) Perturbation of manganese metabolism disrupts cell division in Streptococcus pneumoniae. Mol Microbiol 104:334-348
Martin, Julia E; Edmonds, Katherine A; Bruce, Kevin E et al. (2017) The zinc efflux activator SczA protects Streptococcus pneumoniae serotype 2 D39 from intracellular zinc toxicity. Mol Microbiol 104:636-651
Lisher, John P; Tsui, Ho-Ching Tiffany; Ramos-Montañez, Smirla et al. (2017) Biological and Chemical Adaptation to Endogenous Hydrogen Peroxide Production in Streptococcus pneumoniae D39. mSphere 2:
Chumbler, Nicole M; Rutherford, Stacey A; Zhang, Zhifen et al. (2016) Crystal structure of Clostridium difficile toxin A. Nat Microbiol 1:15002
Martin, Julia E; Giedroc, David P (2016) Functional Determinants of Metal Ion Transport and Selectivity in Paralogous Cation Diffusion Facilitator Transporters CzcD and MntE in Streptococcus pneumoniae. J Bacteriol 198:1066-76
Capdevila, Daiana A; Wang, Jiefei; Giedroc, David P (2016) Bacterial Strategies to Maintain Zinc Metallostasis at the Host-Pathogen Interface. J Biol Chem 291:20858-20868
Fu, Yue; Bruce, Kevin E; Rued, Britta et al. (2016) 1H, 13C, 15N resonance assignments of the extracellular loop 1 domain (ECL1) of Streptococcus pneumoniae D39 FtsX, an essential cell division protein. Biomol NMR Assign 10:89-92
Fu, Yue; Bruce, Kevin E; Wu, Hongwei et al. (2016) The S2 Cu(i) site in CupA from Streptococcus pneumoniae is required for cellular copper resistance. Metallomics 8:61-70

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