Second-Coordination-Sphere Effects Reveal Electronic Structure Differences between the Mitochondrial Amidoxime...

A combination of X-ray absorption and low-temperature electronic absorption spectroscopies has been used to probe the geometric and electronic structures of the human mitochondrial amidoxime reducing component enzyme (hmARC1) in the oxidized Mo(VI) and reduced Mo(IV) forms. Extended X-ray absorption fine structure analysis revealed that oxidized enzyme possesses a 5-coordinate [MoO₂(S_Cys)(PDT)]^- (PDT = pyranopterin dithiolene) active site with a cysteine coordinated to Mo. A 5-coordinate geometry is retained in the reduced state, with the equatorial oxo being protonated. Low-temperature electronic absorption spectroscopy of hmARC1 reveals a spectrum for the oxidized enzyme that is significantly different from what has been reported for sulfite oxidase family enzymes. Time-dependent density functional theory computations on oxidized and reduced hmARC1, and a small molecule analogue for hmARC1_ox, have been used to assist us in making detailed band assignments and developing a greater understanding of enzyme electronic structure contributions to reactivity. Our understanding of the hmARC_red HOMO and the LUMO of the benzamidoxime substrate reveal a potential π-bonding interaction between these redox orbitals, with two-electron occupation of the substrate LUMO along the reaction coordinate activating the O-N bond for cleavage and promoting oxygen atom transfer to the Mo site.