Synthesis and Characterization of an Immobilized Phenylethanolamine N-Methyltransferase Liquid Chromatographic Stationary Phase

• Published in: Analytical biochemistry Vol. 288; no. 1; pp. 83 - 88
• Main Authors: Markoglou, Nektaria, Wainer, Irving W.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.01.2001
• Subjects: Animals; Benzylamines - pharmacology; chromatography; Chromatography, Liquid - methods; Enzyme Inhibitors - pharmacology; Enzymes, Immobilized - metabolism; Ethylmaleimide - pharmacology; Glutaral - analogs & derivatives; Glutaral - metabolism; glutaraldehyde-P; Humans; Hydrogen-Ion Concentration; immobilization; Kinetics; Normetanephrine - metabolism; Organomercury Compounds - pharmacology; phenylethanolamine N-methyltransferase; Phenylethanolamine N-Methyltransferase - chemistry; Phenylethanolamine N-Methyltransferase - metabolism; S-Adenosylmethionine - metabolism; Temperature; immobilization; phenylethanolamine N-methyltransferase; glutaraldehyde-P; chromatography
• ISSN: 0003-2697; 1096-0309
• DOI: 10.1006/abio.2000.4884

Norepinephrine is N-methylated to epinephrine by the catalytic effect of the terminal enzyme in catecholamine biosynthesis, phenylethanolamine N-methyltransferase (PNMT). PNMT has been covalently immobilized onto a silica-based liquid chromatographic support, glutaraldehyde-P (Glut-P). The resulting 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 examined. The results demonstrate that the PNMT-SP can be utilized for the rapid screening of potential PNMT substrates as well as the screening of compounds for PNMT inhibitory activity.