Identification of Active Site Amino Acid Residues of Phenylethanolamine N-Methyltransferase

Abstract

Phenylethanolamine N-methyltransferase (PNMT; EC2.1.1.28) is an enzyme which catalyzes the formation of epinephrine from norepinephrine, the last step of the catecholamine biosynthesis pathway (Axelrod, 1962). PNMT is a monomeric enzyme, and similarly to other mammalian small-molecule N-methyltransferase, S-adenosyl-L-methionine is required for the enzyme reaction as a methyl donor. In addition to its major role in producing adrenomedullary hormones, PNMT has some important roles in blood pressure and neuroendocrine regulation in the central nervous system. Studies using PNMT inhibitors that act in vivo suggested involvement of epinephrine neurons in blood pressure regulation. However, PNMT inhibitors, more or less, possesses a₂-adrenoceptor blocking activity. Therefore, information regarding the structure of the active site may be useful for development of selective and potent inhibitors for possible use as antihypertensive agents (Fuller, 1982). We have previously isolated a full-length cDNA encoding human PNMT, and revealed the primary structure of the enzyme (Kaneda et al., 1988). Human PNMT consists of 282 amino acid residues with a predicted molecular weight of 30,853 including the initial methionine. Using PCR-based site-directed mutagenesis, we investigated amino acid residues located at the active site of the enzyme.