Nickel Superoxide Dismutase

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This award in the Inorganic, Bioinorganic and Organometallic Chemistry program supports research by Professor Michael Maroney at the University of Massachusetts to understand the role of the protein in creating the active site in recombinant Streptomyces coelicolor nickel superoxide dismutase (NiSOD). Point mutations will be employed to address the following questions: 1. Do residues in the second coordination sphere of Ni play a critical role in optimizing the redox potential of the active site? 2. Is the quaternary structure required for catalysis? 3. What are the roles of conserved tyrosine residues near the active site? 4. How does the protein structure moderate the interaction of the active site with hydrogen peroxide, a molecule that is a reaction product, an active site reductant, and readily oxidizes thiolates, such as the cysteine ligands found in the active site? Mutant enzymes will be characterized using a strategy that employs a variety of complementary biophysical techniques including EPR and x-ray absorption spectroscopy. The effect of mutations on catalysis will be addressed by measuring the catalytic rate constant and examining the reaction mechanism using pulse radiolytic generation of superoxide and UV-Vis spectroscopy, and by assessing the redox potential of the enzyme using redox titrations. Mutations that result in a perturbed mechanism will be further examined using crystallography. Catalytic intermediates that have been detected in a number of existing mutant NiSODs will be freeze-trapped and examined spectroscopically The study of NiSOD will contribute to the understanding of redox catalysis by biological nickel sites. The research will provide multidisciplinary training at the interface of chemistry and biology for students at all levels (undergraduate through post-doctoral).