Transition metal ions play highly specialized roles in biological catalysis and all cells encode regulatory machinery that controls their intracellular availability so that mis-metallation and metal toxicity is avoided. In bacteria, this process is transcriptionally regulated by a panel of "metal sensor" proteins in response to changes in metal bioavailability. The proposed experiments strongly integrate structural and cell biological approaches to elucidate determinants of metal homeostasis and cross-talk in the respiratory pathogen Streptococcus pneumoniae, focusing on zinc, manganese and copper, three metals whose concentrations change dramatically during the course of invasive disease. In the next project period, our specific aims are to 1) Obtain a structural and physicochemical characterization of the novel zinc uptake regulator S. pneumoniae AdcR (adhesin competence repressor). Here we propose NMR, structural and thermodynamic studies of wild-type and mutant AdcRs in an effort to understand allosteric activation of DNA operator binding;2) Perform a direct test of the thermodynamic "set-point" model for zinc homeostasis in S. pneumoniae, which posits that "free" zinc is buffered in a concentration range dictated by the zinc affinities of the uptake (AdcR) and efflux (SczA, streptococcal czcD activator) regulators. Since intracellular metallation status is dictated by free or chelatable zinc, not total zinc, we propose to measure and manipulate free Zn in the cytoplasm using a FRET-based imaging assay;3) Obtain structural and mechanistic insights into copper resistance and trafficking in S. pneumoniae, a process characterized by features not previously observed;and 4) Carry out structural studies of the CuI sensor CsoR and CsoR-related proteins in Mycobacterium tuberculosis and other bacilli. Small angle X-ray scattering, NMR and crystallographic studies are proposed. Since an emerging aspect of nutritional immunity to infection by bacterial human pathogens is the host control of metal availability, these studies lay the foundation for our long-term goal of disrupting metal availability or inducing metal toxicity by developing new antimicrobial agents that selectively target key components of transition metal homeostasis systems.

Public Health Relevance

Bacterial pathogens require metal ions to perform processes related to both normal physiology and life- threatening invasive disease. As such, host control of metal ion availability is a critical component of the host-pathogen interface. Our studies of this process in the important respiratory pathogen Streptococcus pneumoniae may lead to the identification of new antibiotic targets that play key roles in this process.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Research Project (R01)
Project #
Application #
Study Section
Macromolecular Structure and Function A Study Section (MSFA)
Program Officer
Anderson, Vernon
Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
Indiana University Bloomington
Schools of Arts and Sciences
United States
Zip Code
Higgins, Khadine A; Giedroc, David (2014) Insights into Protein Allostery in the CsoR/RcnR Family of Transcriptional Repressors. Chem Lett 43:20-25
Reeves, B D; Joshi, N; Campanello, G C et al. (2014) Conversion of S-phenylsulfonylcysteine residues to mixed disulfides at pH 4.0: utility in protein thiol blocking and in protein-S-nitrosothiol detection. Org Biomol Chem 12:7942-56
Fu, Yue; Chang, Feng-Ming James; Giedroc, David P (2014) Copper transport and trafficking at the host-bacterial pathogen interface. Acc Chem Res 47:3605-13
Guerra, Alfredo J; Giedroc, David P (2014) Backbone and sterospecific methyl side chain resonance assignments of the homodimeric zinc sensor AdcR (32 kDa) in the apo- and Zn(II)-bound states. Biomol NMR Assign 8:11-4
Braymer, Joseph J; Giedroc, David P (2014) Recent developments in copper and zinc homeostasis in bacterial pathogens. Curr Opin Chem Biol 19:59-66
Hughes, H Velocity; Lisher, John P; Hardy, Gail G et al. (2013) Co-ordinate synthesis and protein localization in a bacterial organelle by the action of a penicillin-binding-protein. Mol Microbiol 90:1162-77
Fu, Yue; Tsui, Ho-Ching Tiffany; Bruce, Kevin E et al. (2013) A new structural paradigm in copper resistance in Streptococcus pneumoniae. Nat Chem Biol 9:177-83
Chakravorty, Dhruva K; Parker, Trent M; Guerra, Alfredo J et al. (2013) Energetics of zinc-mediated interactions in the allosteric pathways of metal sensor proteins. J Am Chem Soc 135:30-3
Lisher, John P; Higgins, Khadine A; Maroney, Michael J et al. (2013) Physical characterization of the manganese-sensing pneumococcal surface antigen repressor from Streptococcus pneumoniae. Biochemistry 52:7689-701
Chakravorty, Dhruva K; Wang, Bing; Lee, Chul Won et al. (2013) Solution NMR refinement of a metal ion bound protein using metal ion inclusive restrained molecular dynamics methods. J Biomol NMR 56:125-37

Showing the most recent 10 out of 24 publications