Heavy metal poisoning by elements such as mercury, lead, cadmium, and arsenic is a significant human health problem. Understanding the interaction of heavy metals with proteins is essential for defining the mechanism of toxicity, developing ways to minimize human exposure and to provide therapeutic regimens for the removal of toxic ions. Our goals are (1) to develop peptide systems that provide a foundation for understanding metal binding by metalloregulatory proteins and metallochaperones, (2) to understand the thermodynamics and kinetics of heavy metals in helical assemblies and (3) to prepare new designed peptides utilizing different ligands (e.g., histidine or D-amino acids) or asymmetric binding sites in single polypeptides that fold into 1-helical bundles. To achieve these goals we will use a de novo peptide system based on the three-stranded coiled coil peptide aggregate motif that encapsulates with high affinity single heavy metal ions and provides spectroscopic models of mercury, cadmium, and arsenic binding sites in biological systems. We will generate high resolution structures of this peptide system in the presence and absence of these heavy metals, elucidate the kinetic and thermodynamic mechanisms of heavy metal encapsulation, and expand the array of characterized systems to include single chain peptides that encapsulate heavy metals, coiled coils that provide different coordination environments than the original design and those that encapsulate more than one heavy metal ion. These studies will expand the foundation of knowledge that has been laid by the scientific community investigating metallopeptide design, metalloregulatory proteins and heavy metal detoxification. These objectives will develop insight into the interplay between metal coordination and apopeptide structure in defining the overall metallopeptide fold, an important aspect of metallopeptide design.

Public Health Relevance

Relevance to Human Health Heavy metals such as PbII, HgII, AsII and CdII cause severe health concerns due both to their acute toxicity and the long term effects of chronic exposure (e.g., the EPA estimated in 2002 that over 300,000 children in the USA had blood levels exceeding 10 ?g/dL). Our studies address these concerns in numerous ways: including making mimics of heavy metal binding sites in proteins in order to understand how heavy metals bind to proteins, defining the rates at which reactions occur and establishing the thermodynamic preferences of these metals to different sites. We are also relating this work to studies on proteins used to recognize or detoxify heavy metals in bacteria, fungi and humans (e.g., assessing how HgII interacts with the human copper chaperone HAH1).

Agency
National Institute of Health (NIH)
Institute
National Institute of Environmental Health Sciences (NIEHS)
Type
Research Project (R01)
Project #
5R01ES012236-09
Application #
8230719
Study Section
Macromolecular Structure and Function A Study Section (MSFA)
Program Officer
Balshaw, David M
Project Start
2003-06-09
Project End
2014-02-28
Budget Start
2012-03-01
Budget End
2013-02-28
Support Year
9
Fiscal Year
2012
Total Cost
$267,374
Indirect Cost
$90,956
Name
University of Michigan Ann Arbor
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
073133571
City
Ann Arbor
State
MI
Country
United States
Zip Code
48109
Cangelosi, Virginia M; Deb, Aniruddha; Penner-Hahn, James E et al. (2014) A de?novo designed metalloenzyme for the hydration of CO2. Angew Chem Int Ed Engl 53:7900-3
Zastrow, Melissa L; Pecoraro, Vincent L (2014) Designing hydrolytic zinc metalloenzymes. Biochemistry 53:957-78
Peacock, Anna F A; Pecoraro, Vincent L (2013) Natural and artificial proteins containing cadmium. Met Ions Life Sci 11:303-37
Luczkowski, Marek; Zeider, Brian A; Hinz, Alia V H et al. (2013) Probing the coordination environment of the human copper chaperone HAH1: characterization of Hg(II)-bridged homodimeric species in solution. Chemistry 19:9042-9
Zastrow, Melissa L; Pecoraro, Vincent L (2013) Designing functional metalloproteins: from structural to catalytic metal sites. Coord Chem Rev 257:2565-2588
Zastrow, Melissa L; Pecoraro, Vincent L (2013) Influence of active site location on catalytic activity in de novo-designed zinc metalloenzymes. J Am Chem Soc 135:5895-903
Chakraborty, Saumen; Iranzo, Olga; Zuiderweg, Erik R P et al. (2012) Experimental and theoretical evaluation of multisite cadmium(II) exchange in designed three-stranded coiled-coil peptides. J Am Chem Soc 134:6191-203
Zastrow, Melissa L; Peacock, Anna F A; Stuckey, Jeanne A et al. (2012) Hydrolytic catalysis and structural stabilization in a designed metalloprotein. Nat Chem 4:118-23
Zampella, Giuseppe; Neupane, Kosh P; De Gioia, Luca et al. (2012) The importance of stereochemically active lone pairs for influencing Pb(II) and As(III) protein binding. Chemistry 18:2040-50
Neupane, Kosh P; Pecoraro, Vincent L (2011) Pb-207 NMR spectroscopy reveals that Pb(II) coordinates with glutathione (GSH) and tris cysteine zinc finger proteins in a PbS3 coordination environment. J Inorg Biochem 105:1030-4

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