Superoxide dismutases (SODs) are metalloenzymes containing Mn, Fe, Cu/Zn, or Ni active sites that defend biological systems against oxidative damage mediated by the superoxide radical anion (O2), a product of aerobic metabolism. SODs have also been shown to protect against inflammation and are involved in a range of anti-cancer and anti-aging mechanisms. This proposal focuses on the structurally similar Mn- and Fe-dependent SODs, which accomplish their function through disproportionation of O2 to 02 and H202.The long-term objectives of the research outlined in this proposal are- to identify key geometric and electronic structure contributions to the reactivities of Mn- and Fe-dependent SODs and- to obtain molecular-level insight into the reaction mechanisms of these enzymes.With these goals in mind, the following specific aims have been formulated:- Generate electronic structure descriptions of the oxidized and reduced Mn- and Fe-SOD active sites.- Explore the factors responsible for SOD metal specificity.- Assess the role of second-sphere amino acids in tuning active site properties.- Define the nature of the substrate-metal interaction for Mn- and Fe-SODs and evaluate key steps in the corresponding catalytic cycles on experimental and theoretical levels.Our approach involves using a combination of spectroscopic tools (absorption, CD, MCD, EPR, and rR) and computational methods (DFT and NBO) to study the native Mn- and Fe-SOD proteins, the catalytically inactive metal-substituted Mn- and Fe-SOD species, and several mutant proteins. These studies will systematically explore geometric and electronic factors contributing to SOD activity.As SOD enzymes demonstrate therapeutic efficacy in animal models of disease states involving superoxide, low molecular weight SOD enzyme mimics (synzymes) have been proposed for the treatment of a variety of diseases. Synzymes could have distinct advantages over natural SOD enzymes as pharmaceutical agents, such as cellular permeability, lack of immunogenicity, longer lifetimes, potential for oral delivery, and lower production costs. Thus, understanding the principles by which the Mn- and Fe-SOD enzymes achieve their remarkably high catalytic rates, in particular the influence of second coordination shell amino acids on active site electronics, could aid significantly in the rational design of SOD mimics for pharmaceutical applications.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM064631-02
Application #
6640333
Study Section
Metallobiochemistry Study Section (BMT)
Program Officer
Preusch, Peter C
Project Start
2002-06-05
Project End
2007-05-31
Budget Start
2003-06-01
Budget End
2004-05-31
Support Year
2
Fiscal Year
2003
Total Cost
$145,500
Indirect Cost
Name
University of Wisconsin Madison
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
161202122
City
Madison
State
WI
Country
United States
Zip Code
53715
Ryan, Kelly C; Guce, Abigail I; Johnson, Olivia E et al. (2015) Nickel superoxide dismutase: structural and functional roles of His1 and its H-bonding network. Biochemistry 54:1016-27
Blaesi, Elizabeth J; Fox, Brian G; Brunold, Thomas C (2015) Spectroscopic and Computational Investigation of the H155A Variant of Cysteine Dioxygenase: Geometric and Electronic Consequences of a Third-Sphere Amino Acid Substitution. Biochemistry 54:2874-84
Broering, Ellen P; Dillon, Stephanie; Gale, Eric M et al. (2015) Accessing Ni(III)-thiolate versus Ni(II)-thiyl bonding in a family of Ni-N2S2 synthetic models of NiSOD. Inorg Chem 54:3815-28
Blaesi, Elizabeth J; Fox, Brian G; Brunold, Thomas C (2014) Spectroscopic and computational investigation of iron(III) cysteine dioxygenase: implications for the nature of the putative superoxo-Fe(III) intermediate. Biochemistry 53:5759-70
Gutman, Craig T; Guzei, Ilia A; Brunold, Thomas C (2013) Structural, spectroscopic, and computational characterization of the azide adduct of Fe(III)(2,6-diacetylpyridinebis(semioxamazide)), a functional analogue of iron superoxide dismutase. Inorg Chem 52:8909-18
Blaesi, Elizabeth J; Gardner, Jessica D; Fox, Brian G et al. (2013) Spectroscopic and computational characterization of the NO adduct of substrate-bound Fe(II) cysteine dioxygenase: insights into the mechanism of O2 activation. Biochemistry 52:6040-51
Jackson, Timothy A; Gutman, Craig T; Maliekal, James et al. (2013) Geometric and electronic structures of manganese-substituted iron superoxide dismutase. Inorg Chem 52:3356-67
Gutman, Craig T; Brunold, Thomas C (2012) Spectroscopic and computational studies of a small-molecule functional mimic of iron superoxide dismutase, iron 2,6-diacetylpyridinebis(semioxamazide). Inorg Chem 51:12729-37
Johnson, Olivia E; Ryan, Kelly C; Maroney, Michael J et al. (2010) Spectroscopic and computational investigation of three Cys-to-Ser mutants of nickel superoxide dismutase: insight into the roles played by the Cys2 and Cys6 active-site residues. J Biol Inorg Chem 15:777-93
Van Heuvelen, Katherine M; Cho, Jaeheung; Dingee, Timothy et al. (2010) Spectroscopic and computational studies of a series of high-spin Ni(II) thiolate complexes. Inorg Chem 49:6535-44

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