ZINC COORDINATION AND REGULATION OF ZINC METALLOPROTEINS

Summary

Principal Investigator: David P Giedroc
Affiliation: Indiana University
Country: USA
Abstract: This proposal requests continued support for ongoing studies of the structure, bioinorganic chemistry and allosteric regulation by zinc of selected zinc metalloregulatory transcription factors derived from mammalian and prokaryotic organisms which play roles in zinc homeostasis. Human metal-responsive element (MRE)-binding transcription factor-1 (MTF-1) possesses six Cys2-His2 zinc finger domains and activates the expression of metallothionein genes in a zinc-dependent manner. Previous studies led the PI to hypothesize that the N-terminal four fingers play a structural role in mediating high affinity binding to the MRE while the C-terminal finger domains perform a zinc sensor and/or transducer role. The following experiments on MTF-1 are proposed: 1) Define the solution structure of a C-terminal zinc finger fragment (finger domains 4-6) of MTF-1 using multi-dimensional NMR spectroscopy. These studies will be complemented by equilibrium and kinetic metal binding studies to delineate the unusual metal binding properties of these metalloregulatory domains; and 2) Test the metalloregulatory model of MTF-1 function in vivo by determining the metal-dependent transcriptional activation efficacy of broken-finger mutants of intact MTF-1 by transfection experiments in mammalian and yeast cells. Comparative studies of three evolutionarily related prokaryotic zinc metalloregulatory repressors, Synechococcus SmtB, Synechocystis ZiaR, S. aureus CzrA and a cadmium repressor, S. aureus pI258 CadC are also proposed. These studies will provide a critical test of the metalloregulatory hypothesis which holds that the biological specificity and mechanism of regulation is encoded in the bioinorganic chemistry (metal coordination chemistry, specificity, affinity and stoichiometry) and the thermodynamic linkage relationships between metal binding, DNA binding and protein self-association of these metal switch proteins. Results of the proposed studies will provide new molecular insights into the relationships between structure, coordination chemistry and regulation by metal ions in metalloregulatory transcription factors, which, over the longer term, will permit their systematic re-engineering as templates for metal-site redesign and metal sensors with novel properties, tailored to biomedical and bioremediation uses.
Funding Period: 1991-04-01 - 2004-03-31
more information: NIH RePORT

Top Publications

  1. pmc Crystal structure of the zinc-dependent MarR family transcriptional regulator AdcR in the Zn(II)-bound state
    Alfredo J Guerra
    Department of Chemistry, Indiana University, Bloomington, Indiana 47405, USA
    J Am Chem Soc 133:19614-7. 2011
  2. doi Illuminating allostery in metal sensing transcriptional regulators
    Nicholas E Grossoehme
    Winthrop University, Rock Hill, SC, USA
    Methods Mol Biol 875:165-92. 2012
  3. pmc Backbone resonance assignments of the homotetrameric (48 kD) copper sensor CsoR from Geobacillus thermodenitrificans in the apo- and Cu(I)-bound states: insights into copper-mediated allostery
    H Jerome Coyne
    Department of Chemistry, Indiana University, 212 S Hawthorne Drive, Bloomington, IN 47405 7102, USA
    Biomol NMR Assign 7:279-83. 2013
  4. pmc Backbone and sterospecific methyl side chain resonance assignments of the homodimeric zinc sensor AdcR (32 kDa) in the apo- and Zn(II)-bound states
    Alfredo J Guerra
    Department of Chemistry, Indiana University, Bloomington, IN, 47405 7102, USA
    Biomol NMR Assign 8:11-4. 2014
  5. pmc Energetics of zinc-mediated interactions in the allosteric pathways of metal sensor proteins
    Dhruva K Chakravorty
    Department of Chemistry and Quantum Theory Project, University of Florida, Gainesville, Florida 32611 8435, United States
    J Am Chem Soc 135:30-3. 2013
  6. pmc Allosteric inhibition of a zinc-sensing transcriptional repressor: insights into the arsenic repressor (ArsR) family
    Gregory C Campanello
    Department of Chemistry, Indiana University, Bloomington, IN 47405 7102, USA
    J Mol Biol 425:1143-57. 2013
  7. pmc A new structural paradigm in copper resistance in Streptococcus pneumoniae
    Yue Fu
    Department of Chemistry, Indiana University, Bloomington, Indiana, USA
    Nat Chem Biol 9:177-83. 2013
  8. pmc Solution NMR refinement of a metal ion bound protein using metal ion inclusive restrained molecular dynamics methods
    Dhruva K Chakravorty
    Department of Chemistry and the Quantum Theory Project, University of Florida, 2238 New Physics Building, P O Box 118435, Gainesville, FL 32611 8435, USA
    J Biomol NMR 56:125-37. 2013
  9. pmc Physical Characterization of the Manganese-Sensing Pneumococcal Surface Antigen Repressor from Streptococcus pneumoniae
    John P Lisher
    Department of Chemistry and Interdisciplinary Graduate Program in Biochemistry, Indiana University, Bloomington, Indiana 47405 7102, United States
    Biochemistry 52:7689-701. 2013
  10. pmc Co-ordinate synthesis and protein localization in a bacterial organelle by the action of a penicillin-binding-protein
    H Velocity Hughes
    Department of Biology, Indiana University, Bloomington, IN, 47405, USA
    Mol Microbiol 90:1162-77. 2013

Scientific Experts

  • David P Giedroc
  • Alfredo J Guerra
  • John P Lisher
  • Zhen Ma
  • Dhruva K Chakravorty
  • Nicholas E Grossoehme
  • Hermes Reyes-Caballero
  • Robert A Scott
  • Kenneth M Merz
  • Gregory C Campanello
  • Charles E Dann
  • Chul Won Lee
  • Malcolm E Winkler
  • Krystyna M Kazmierczak
  • Faith E Jacobsen
  • Darin M Cowart
  • Joseph J Braymer
  • H Velocity Hughes
  • Khadine A Higgins
  • Yue Fu
  • Michael J Maroney
  • H Jerome Coyne
  • Bing Wang
  • Feng Ming James Chang
  • Richard D Dimarchi
  • Nicholas Grossoehme
  • Randy J Arnold
  • Brian P Ward
  • Frederik Sommer
  • Yun Wang
  • Xiaohua Chen
  • Meiying Zheng
  • Tong Liu
  • Yves V Brun
  • Ho Ching Tiffany Tsui
  • Kevin E Bruce
  • Trent M Parker
  • Yuzhen Ye
  • C David Sherrill
  • Gail G Hardy
  • David T Kysela
  • Lok To Sham
  • Thomas E Kehl-Fie
  • Matthew A Lauber
  • William E Running
  • Keith W Adams
  • Eric P Skaar
  • James P Reilly
  • Darin Cowart
  • Uma Mahendra Kumar Koppolu
  • Sabeeha S Merchant
  • Jennifer S Cavet
  • Janette Kropat
  • John Kendall
  • Davin Malasarn
  • David R Cooper
  • Minmin Yu
  • Limei Zhang
  • Marcin Cieslik
  • Urszula Derewenda
  • Ian A Wilson
  • Scott A Lesley
  • Nickolas E Grossoehme
  • Li Wei Hung
  • Zygmunt S Derewenda
  • Graham N George
  • Lars Hemmingsen
  • Jacob Shokes

Detail Information

Publications29

  1. pmc Crystal structure of the zinc-dependent MarR family transcriptional regulator AdcR in the Zn(II)-bound state
    Alfredo J Guerra
    Department of Chemistry, Indiana University, Bloomington, Indiana 47405, USA
    J Am Chem Soc 133:19614-7. 2011
    ..This co-repression mechanism is unprecedented in MarR transcriptional regulators...
  2. doi Illuminating allostery in metal sensing transcriptional regulators
    Nicholas E Grossoehme
    Winthrop University, Rock Hill, SC, USA
    Methods Mol Biol 875:165-92. 2012
    ....
  3. pmc Backbone resonance assignments of the homotetrameric (48 kD) copper sensor CsoR from Geobacillus thermodenitrificans in the apo- and Cu(I)-bound states: insights into copper-mediated allostery
    H Jerome Coyne
    Department of Chemistry, Indiana University, 212 S Hawthorne Drive, Bloomington, IN 47405 7102, USA
    Biomol NMR Assign 7:279-83. 2013
    ..These data provide the first spectroscopic evidence that Cu(I)-binding induces an interruption in the long α2 helix of CsoR. ..
  4. pmc Backbone and sterospecific methyl side chain resonance assignments of the homodimeric zinc sensor AdcR (32 kDa) in the apo- and Zn(II)-bound states
    Alfredo J Guerra
    Department of Chemistry, Indiana University, Bloomington, IN, 47405 7102, USA
    Biomol NMR Assign 8:11-4. 2014
    ..Two-dimensional NMR studies reveal large changes in the spectrum upon Zn(II) binding. Near complete backbone and stereospecific methyl group resonance assignments for apo- and Zn(II)-AdcR are presented here. ..
  5. pmc Energetics of zinc-mediated interactions in the allosteric pathways of metal sensor proteins
    Dhruva K Chakravorty
    Department of Chemistry and Quantum Theory Project, University of Florida, Gainesville, Florida 32611 8435, United States
    J Am Chem Soc 135:30-3. 2013
    ..These findings suggest a conserved functional role of metal-mediated second coordination shell hydrogen bonds at allosterically responsive sites in zinc-sensing transcription regulators...
  6. pmc Allosteric inhibition of a zinc-sensing transcriptional repressor: insights into the arsenic repressor (ArsR) family
    Gregory C Campanello
    Department of Chemistry, Indiana University, Bloomington, IN 47405 7102, USA
    J Mol Biol 425:1143-57. 2013
    ....
  7. pmc A new structural paradigm in copper resistance in Streptococcus pneumoniae
    Yue Fu
    Department of Chemistry, Indiana University, Bloomington, Indiana, USA
    Nat Chem Biol 9:177-83. 2013
    ..However, copper binding by CopA(MBD) is not essential for cellular copper resistance, consistent with a primary role of CupA in cytoplasmic Cu(I) sequestration and/or direct delivery to the transmembrane site of CopA for cellular efflux...
  8. pmc Solution NMR refinement of a metal ion bound protein using metal ion inclusive restrained molecular dynamics methods
    Dhruva K Chakravorty
    Department of Chemistry and the Quantum Theory Project, University of Florida, 2238 New Physics Building, P O Box 118435, Gainesville, FL 32611 8435, USA
    J Biomol NMR 56:125-37. 2013
    ..This methodology has potentially broad applicability in the structure determination of metal ion bound proteins, protein folding and metal template protein-design studies...
  9. pmc Physical Characterization of the Manganese-Sensing Pneumococcal Surface Antigen Repressor from Streptococcus pneumoniae
    John P Lisher
    Department of Chemistry and Interdisciplinary Graduate Program in Biochemistry, Indiana University, Bloomington, Indiana 47405 7102, United States
    Biochemistry 52:7689-701. 2013
    ..Mutational studies reveal that Asp7 in site 2 is a critical ligand for Mn(II)-dependent allosteric activation of DNA binding. These findings are discussed in the context of other well-studied DtxR/MntR Mn(II)/Fe(II) metallorepressors. ..
  10. pmc Co-ordinate synthesis and protein localization in a bacterial organelle by the action of a penicillin-binding-protein
    H Velocity Hughes
    Department of Biology, Indiana University, Bloomington, IN, 47405, USA
    Mol Microbiol 90:1162-77. 2013
    ....
  11. pmc Manganese acquisition and homeostasis at the host-pathogen interface
    John P Lisher
    Graduate Program in Biochemistry, Indiana University Bloomington, IN, USA
    Front Cell Infect Microbiol 3:91. 2013
    ....
  12. pmc Recent developments in copper and zinc homeostasis in bacterial pathogens
    Joseph J Braymer
    Department of Chemistry, Indiana University, Bloomington, IN 47405 7102, USA
    Curr Opin Chem Biol 19:59-66. 2014
    ..In this review, we summarize recent findings on how bacterial pathogens mount a metal-specific response to derail host efforts to win the 'fight over metals.' ..
  13. pmc Ratiometric pulse-chase amidination mass spectrometry as a probe of biomolecular complex formation
    Feng Ming James Chang
    Department of Chemistry, Indiana University, 800 East Kirkwood Avenue, Bloomington, Indiana 47405 7102, USA
    Anal Chem 83:9092-9. 2011
    ..New insights into operator DNA binding and the Cu(I)-mediated structural transition in the tetrameric copper sensor CsoR are also obtained...
  14. pmc Elucidation of the functional metal binding profile of a Cd(II)/Pb(II) sensor CmtR(Sc) from Streptomyces coelicolor
    Yun Wang
    Department of Biochemistry and Biophysics, Texas A and M University, College Station, Texas 77843 2128, USA
    Biochemistry 49:6617-26. 2010
    ..The implications of these findings on the ability to predict metal specificity and function from metal-site signatures in the primary structure of ArsR family proteins are discussed...
  15. pmc The CRR1 nutritional copper sensor in Chlamydomonas contains two distinct metal-responsive domains
    Frederik Sommer
    Department of Chemistry and Biochemistry, University of California, Los Angeles, California 90095 1569, USA
    Plant Cell 22:4098-113. 2010
    ..Strains carrying the crr1-ΔCys allele upregulate ZRT genes and hyperaccumulate Zn(II), suggesting that the effect of nickel ions may be revealing a role for the C-terminal domain of CRR1 in zinc homeostasis in Chlamydomonas...
  16. pmc The metalloregulatory zinc site in Streptococcus pneumoniae AdcR, a zinc-activated MarR family repressor
    Hermes Reyes-Caballero
    Department of Chemistry, Indiana University, Bloomington, IN 47405 7102, USA
    J Mol Biol 403:197-216. 2010
    ..These studies implicate His42, His108 and H112 as metalloregulatory zinc ligands in S. pneumoniae AdcR...
  17. pmc Molecular insights into the metal selectivity of the copper(I)-sensing repressor CsoR from Bacillus subtilis
    Zhen Ma
    Department of Chemistry, Indiana University, Bloomington, Indiana 47405 7102, USA
    Biochemistry 48:3325-34. 2009
    ..The implications of these findings on the specificity of metal-sensing sites in CsoR/RcnR proteins are discussed...
  18. pmc Hydrogen peroxide sensing in Bacillus subtilis: it is all about the (metallo)regulator
    David P Giedroc
    Department of Chemistry, Indiana University, Bloomington, IN 47405 7002, USA
    Mol Microbiol 73:1-4. 2009
    ....
  19. pmc Unnatural amino acid substitution as a probe of the allosteric coupling pathway in a mycobacterial Cu(I) sensor
    Zhen Ma
    Department of Chemistry, Indiana University, Bloomington, Indiana 47405 7102, USA
    J Am Chem Soc 131:18044-5. 2009
    ..A model analogous to the allosteric switch mechanism in Staphylococcus aureus CzrA, a zinc sensor protein with a completely different fold, is proposed...
  20. pmc Energetics of allosteric negative coupling in the zinc sensor S. aureus CzrA
    Nicholas E Grossoehme
    Department of Chemistry, Indiana University, Bloomington, Indiana 47405 7102, USA
    J Am Chem Soc 131:17860-70. 2009
    ..1 kcal mol(-1), i.e., DeltaS(c)(t) > 0). The implications of these findings for other metalloregulatory proteins are discussed...
  21. pmc Mycobacterium tuberculosis NmtR harbors a nickel sensing site with parallels to Escherichia coli RcnR
    Hermes Reyes-Caballero
    Department of Chemistry, Indiana University, Bloomington, Indiana 47405 7102, USA
    Biochemistry 50:7941-52. 2011
    ....
  22. pmc Interplay between manganese and zinc homeostasis in the human pathogen Streptococcus pneumoniae
    Faith E Jacobsen
    Department of Chemistry, Indiana University, Bloomington, IN 47405 7102, USA
    Metallomics 3:38-41. 2011
    ..We conclude that Zn(II) toxicity results in a cytoplasmic Mn(II) deficiency, possibly caused by competition at the Mn(II) uptake transporter protein PsaA...
  23. pmc Control of copper resistance and inorganic sulfur metabolism by paralogous regulators in Staphylococcus aureus
    Nicholas Grossoehme
    Department of Chemistry, Indiana University, Bloomington, Indiana 47405 7102, USA
    J Biol Chem 286:13522-31. 2011
    ..CsoR is unreactive toward sulfite under the same conditions. We conclude that CsoR and CstR are paralogs in S. aureus that function in the same cytoplasm to control distinct physiological processes...
  24. pmc Metalloregulatory proteins: metal selectivity and allosteric switching
    Hermes Reyes-Caballero
    Department of Chemistry, Indiana University, Bloomington, 47405 7102, United States
    Biophys Chem 156:103-14. 2011
    ....
  25. pmc Simulations of allosteric motions in the zinc sensor CzrA
    Dhruva K Chakravorty
    Department of Chemistry and the Quantum Theory Project, 2328 New Physics Building, P O Box 118435, University of Florida, Gainesville, Florida 32611 8435, USA
    J Am Chem Soc 134:3367-76. 2012
    ..Overall, our simulations provide molecular-level insights into the mechanism of allosteric regulation by CzrA and demonstrate the importance of protein motion in its biological activity...
  26. pmc Structure of Thermotoga maritima TM0439: implications for the mechanism of bacterial GntR transcription regulators with Zn2+-binding FCD domains
    Meiying Zheng
    Department of Molecular Physiology and Biological Physics, Integrated Center for Structure Function Innovation, University of Virginia, Charlottesville, VA 22908 0736, USA
    Acta Crystallogr D Biol Crystallogr 65:356-65. 2009
    ..Finally, the TM0439 structure is compared with two other FadR-family structures recently deposited by structural genomics consortia. The results call for a revision in the classification of the FadR family of transcription factors...
  27. pmc A Cu(I)-sensing ArsR family metal sensor protein with a relaxed metal selectivity profile
    Tong Liu
    Department of Biochemistry and Biophysics, Texas A and M University, College Station, Texas 77843 2128, USA
    Biochemistry 47:10564-75. 2008
    ..The evolutionary implications of these findings for the ArsR metal sensor family are discussed...