Elucidation of the Role of the Heme Regulatory Motif in Heme Oxygenase-2

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DESCRIPTION (provided by applicant): Heme oxygenase catalyzes the conversion of heme to biliverdin, CO, and iron. The two forms of heme oxygenase (heme oxygenase-1 and heme oxygenase-2) share similar physical and kinetic properties but exhibit different physiological roles and organ locations. The linkage between heme oxygenase-2 and a Caactivated high conductance potassium channel (the BK or Slopoke channel) has been strongly implicated in the control of ventilation by the carotid body in response to changes in the blood oxygen concentration. The major apparent distinction between the sequences of heme oxygenase-1 and heme oxygenase-2 is the occurrence of heme responsive (or regulatory) motifs (HRMs) in heme oxygenase-2. The interaction of heme with HRMs has been proposed to control the activity or stability of several regulatory proteins. Based on recent results, HRM does not bind heme per se or affect HO-2 stability, but appears to act as a redox switch involved in regulating the affinity of heme oxygenase-2 for heme and the spin state of the ferric heme. We plan to use spectroscopic, kinetic, molecular genetic, and electrophysiology studies to determine the role of the HRM in controlling heme oxygenase-2 structure and function. We will characterize the heme and HRM thiol/disulfide redox centers in heme oxygenase-2 by spectroscopic and kinetic methods and determine the redox state of the HRM in vitro and in vivo under various physiological conditions. We also will perform spectroscopic and electrophysiology measurements to determine the mechanism by which heme oxygenase-2 influences BK channel function. PUBLIC HEALTH RELEVANCE Heme oxygenase is the only mammalian protein known to degrade heme and sits at the nexus of several major redox and metal regulatory systems. Two Heme Regulatory Motifs at the C- terminal end of heme oxygenase-2 (HO-2) differentiate it from heme oxygenase-1. This proposal is aimed at determining how(s) these Heme Regulatory Motifs regulate the enzymatic and regulatory properties of HO-2.