There exists a fundamental gap in our understanding of the relationship between pyranopterin molybdenum enzyme structure and function. The long-term goals of our research are to understand enzyme mechanism in order to improve the quality of human health and the environment. Our objective in pursuit of these goals is to develop a comprehensive understanding of how active site geometric and electronic structure contributes to proper enzyme function. This will be accomplished through a combination of detailed spectroscopic (electronic absorption, MCD, Raman, XAS, EPR, etc.) and bonding studies on enzymes from all three pyranopterin Mo enzyme families. This work will be complemented by parallel studies on small molecule analogues. Our combined spectroscopic approach is designed to provide detailed insight into key electronic structure contributions to catalysis and our computational studies will be calibrated to spectroscopic and reactivity data in order to obtain a high level of mechanistic detail. The central hypothesis is that a complex interplay exists between active site geometric and electronic structure that functions to facilitate the unique reactions these enzymes catalyze. The rationale for this research is that a comprehensive understanding of how electronic structure contributes to reactivity in pyranopterin Mo enzymes will lead to greater insight into innovative drug and pro-drug design, understanding disease states related to Mo enzyme activity, and generally improving human health and the environment. We will test our central hypothesis in order to accomplish the stated objective of this proposal through the successful pursuit of the three Specific Aims 1) Determine the reaction coordinate for the molybdenum hydroxylases, 2) Develop a comprehensive understanding of active site contributions to catalysis in the sulfite oxidase family enzymes YedY and mARC, and 3) Identify key molybdenum-sulfur covalency contributions to electron transfer (ET) and redox potential modulation in dimethylsulfoxide reductase family enzymes. Our research plan is innovative because it 1) utilizes a combined spectroscopic approach coupled with sophisticated computational studies to probe key enzyme states with minimal or no interference from endogenous chromophores, 2) proposes to study a new Mo enzyme found in humans (mARC), and 3) contributes to a greater understanding of the pyranopterin dithiolene in catalysis. This proposed research is significant because it will lead to a markedly greater understanding of how active site geometric and electronic structure directly affect molybdoenzyme substrate specificity and the nature of the reaction coordinate, the nature of the reaction catalyzed (oxidation/reduction), and the role of the pyranopterin dithiolene cofactor in catalysis.
The proposed research is related to public health because, in humans, these enzymes are involved in drug metabolism, oxidative stress and fatal diseases. Therefore, the work detailed in this proposal is relevant to NIH's mission at it pertains to the pursuit of fundamental knowledge regarding the nature and behavior of living systems and the application of that knowledge to extend healthy life and reduce the burdens of illness.
|Giles, Logan J; Ruppelt, Christian; Yang, Jing et al. (2014) Molybdenum site structure of MOSC family proteins. Inorg Chem 53:9460-2|
|Dong, Chao; Yang, Jing; Leimkühler, Silke et al. (2014) Pyranopterin dithiolene distortions relevant to electron transfer in xanthine oxidase/dehydrogenase. Inorg Chem 53:7077-9|
|Stein, Benjamin W; Kirk, Martin L (2014) Orbital contributions to CO oxidation in Mo-Cu carbon monoxide dehydrogenase. Chem Commun (Camb) 50:1104-6|
|Cutsail 3rd, George E; Stein, Benjamin W; Subedi, Deepak et al. (2014) EPR, ENDOR, and electronic structure studies of the Jahn-Teller distortion in an Fe(V) nitride. J Am Chem Soc 136:12323-36|
|Sugimoto, Hideki; Sato, Masanori; Giles, Logan J et al. (2013) Oxo-carboxylato-molybdenum(VI) complexes possessing dithiolene ligands related to the active site of type II DMSOR family molybdoenzymes. Dalton Trans 42:15927-30|
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|Matz, Kelly G; Mtei, Regina P; Rothstein, Rebecca et al. (2011) Study of molybdenum(4+) quinoxalyldithiolenes as models for the noninnocent pyranopterin in the molybdenum cofactor. Inorg Chem 50:9804-15|
|Mtei, Regina P; Lyashenko, Ganna; Stein, Benjamin et al. (2011) Spectroscopic and electronic structure studies of a dimethyl sulfoxide reductase catalytic intermediate: implications for electron- and atom-transfer reactivity. J Am Chem Soc 133:9762-74|
|Sempombe, Joseph; Galinato, Mary Grace I; Elmore, Bradley O et al. (2011) Mutation in the flavin mononucleotide domain modulates magnetic circular dichroism spectra of the iNOS ferric cyano complex in a substrate-specific manner. Inorg Chem 50:6859-61|
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