Non-heme Fe(II) and alpha-ketoglutarate dependent dioxygenases are a relatively large family of enzymes important in a wide range of biologically and medically relevant processes. Enzymes of this family are believed to use a generally conserved catalytic mechanism and possess a common 2-histidine, 1-carboxylate (typically supplied by Asp/Glu) Fe(II) coordination motif known as a "facial triad." One member of this family, Taurine Hydroxylase (TauD), catalyzes the oxidation of taurine and cofactor alpha-ketogluteric acid (α-KG) to succinate, carbon dioxide (CO2), and aminoacetylaldehyde using molecular oxygen (O2). To study the catalytically relevant active site structure with Electron Paramagnetic Resonance Spectroscopy (EPR) enzyme samples were prepared with nitric oxide (NO) as a surrogate to O2 forming a S=3/2 {FeNO}7 centered active site. Using Electron Spin Echo Envelope Modulation (ESEEM) we have measured the position and orientation of taurine in relation to the magnetic axes of the {FeNO}7 complex at TauD's active site center. Using Hyperfine Subevel Correlation (HYSCORE), a 2 dimensional version of the ESEEM experiment, we have provided insight into the orientation of directly coordinated His ligands providing a frame of reference for the structural relationship of the substrate and ligands. HYSCORE spectra for samples prepared without substrate or α-KG aided in the understanding of ligand coordination to the {FeNO}7 complex. Xanthine Hydroxylase (XanA) is a poorly understood member of this family of enzymes that catalyzes the conversion of xanthine and α-KG to succinate, CO2, and uric acid. XanA has been shown to allow for growth of Aspergillus nidulans on xanthine as the sole source of nitrogen in the absence of xanthine dehydrogenase activity, the enzyme usually responsible for this function. Homology models based on sequence similarity to TauD have been constructed but are not well studied. The methodology for studying TauD was applied to XanA to provide spectroscopic support for the homology model.
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