RUI: Novel Heme Chemistry of Cytochrome c'

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This project will investigate the novel heme chemistry of Alcaligenes xylosoxidans cytochrome c' (AXCP) with a particular focus on undergraduate involvement. The PI and coworkers have previously reported the unprecedented reactivity of AXCP, whereby nitric oxide (NO) coordinates to the proximal heme face by way of a putative dinitrosyl intermediate. This alternative heme-NO binding mode, involving both distal and proximal heme faces, is challenging previous assumptions of how heme proteins react with NO, and suggests novel mechanisms for modulating ligand interactions that may be relevant to heme-based sensors, such as soluble guanylate cyclase. The novel heme-NO chemistry of AXCP takes on further significance in the light of reports that cytochrome c' may protect bacteria against NO toxicity, either as an NO reductase, or else as a reversible NO-transporter. The PIs group will perform a detailed structure reactivity based dissection of the proximal and distal heme pockets using a combination of kinetics, spectroscopy, mutagenesis, high-resolution x-ray crystallography, and electrochemistry. Specific aims are to (i) understand the mechanism of proximal NO binding (ii) determine the role of the proximal and distal pockets on heme-NO release, and (iii) investigate heme redox reactivity.The broader impacts of this project include the exceptional opportunities for undergraduate students at Eastern Oregon University (EOU)--a small, public liberal arts college--to be engaged in cutting-edge research. In addition to carrying out protein purification and kinetic measurements at EOU, students will travel to collaborating laboratories to gain additional experience in protein characterization, as well as insights into graduate research environments. Overall, the project will support the thriving undergraduate research culture at EOU (exemplified by the in-house Eastern Oregon Science Journal) and will build on the previous successes of the PI in encouraging students to pursue science at the graduate level.