DESIGN OF INHIBITORS OF EPINEPHRINE BIOSYNTHESIS

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Phenylethanolamine N-Methyltransferase (PNMT,E.C.2.1.1.28) is the enzyme that catalyzes the terminal step in the biosynthesis of epinephrine. Increased levels of this enzyme are found during periods of stress and in hypertension. Epinephrine has been implicated in a number of neuroregulatory processes in the brain. we have demonstrated that inhibitors of PNMT can lower blood pressure in spontaneously hypertensive rats; however, all of the inhibitors presently available show alpha 2-adrenergic receptor antagonism that could also produce some of the observed pharmacological effects. Therefore, we propose to design more selective inhibitors of this enzyme by mapping out the PNMT active site (1) with a few carefully selected structural and conformational probes (substrate analogues to probe the possibility of multiple aromatic ring binding sites and to probe the manner in which norepinephrine interacts at the active site of PNMT; multisubstrate inhibitors to enhance selectivity by occupying both the norepinephrine and AdoMet substrate sites, dead end inhibitors to probe the conformational requirements for binding of the ethanolamine side chain; and alternate substrate inhibitors to explore regional lipophilic effects upon binding), (2) with transferred NOE techniques to probe the conformation of ligands when bound to PNMT, and (3) by affinity labeling of amino acid residues at the active site. Results from all three sub- projects will be combined to refine a computer graphics model of the active site of PNMT, a model developed in our laboratories that (when integrated with a similar model to be developed for the alpha 2-adrenegic receptor) will allow the design and synthesis of a selective inhibitor of the enzyme. Such an inhibitor would be a useful pharmacological tool to probe the role played by epinephrine in the central nervous system.