Structure and Mechanism of Myo-Inositol Oxygenase

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DESCRIPTION (provided by applicant): Myo-inositol oxygenase (MIOX) catalyzes the first step in the only known pathway in humans for breakdown of myo-inositol (Ml), the sugar backbone of cell-signaling phosphoinositides. Evidence suggests that increased expression or activity of MIOX may contribute to pathologies commonly associated with diabetes mellitus, marking MIOX as a potential drug target. Our preliminary data show that the MIOX reaction proceeds via an unprecedented chemical mechanism. A non-heme diiron cluster in its mixed-valent, ll/lll, oxidation state, which probably coordinates one or more oxygen atoms of the substrate, reacts with molecular oxygen to generate a formally diiron(lll/lll)-superoxide complex, which abstracts hydrogen from the substrate. We seek to determine by biophysical and biochemical methods: (1) the structure of this unique enzyme and its diiron cofactor; (2) how the protein promotes initial formation and stability of the active, mixed-valent form (which is unstable in many other non-heme diiron proteins); (3) how the substrate interacts with the protein and cofactor; (4) whether and how this interaction activates the cofactor or substrate (or both) for subsequent reaction with O2; and (5) the nature and rate of each chemical step leading from addition of oxygen to the enzyme*substrate complex through release of the product along with the structures of two reactive intermediates that we have already discovered and additional intermediates that remain to be discovered in this sequence. By so doing, we hope to provide the necessary tools for rational design of MIOX inhibitors that could be useful in combating complications from diabetes mellitus.