Synthesis and characterization of an immobilized phenylethanolamine N-methyltransferase liquid chromatographic stationary phase

Norepinephrine is N-methylated to epinephrine by the catalytic effect of th e terminal enzyme in catecholamine biosynthesis, phenylethanolamine N-methy l-transferase (PNMT). PNMT has been covalently immobilized onto a silica-ba sed liquid chromatographic support, glutaraldehyde-P (Glut-P). The resultin g PNMT-Glut-P stationary phase (PNMT-SP) was enzymatically active, stable, and reusable. Standard Michaelis-Menten kinetic studies were performed with both free and immobilized PNMT and known substrates and inhibitors were ex amined. The results demonstrate that the PNMT-SP can be utilized for the ra pid screening of potential PNMT substrates as well as the screening of comp ounds for PNMT inhibitory activity. (C) 2001 Academic Press.