Spin-label assay for phospholipase A2

✍ Scribed by Ching-San Lai; Jian-Zhong Zhang; Joy Joseph

• Publisher: Elsevier Science
• Year: 1988
• Tongue: English
• Weight: 483 KB
• Volume: 172
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1016/0003-2697(88)90461-7

✦ Synopsis

A rapid and continuous method for measuring phospholipase A2 activity using electron spin resonance spectroscopy and a spin-labeled phospholipid as a substrate has been developed. The substrate, 1-palmitoyl-2-(4-doxylpentanoyl)glycerophosphocholine, gives rise principally to a broad ESR line in aqueous solution due to strong spin-spin interactions, probably resulting from its micellar formation. Upon addition of bee venom phospholipase A2, the water-soluble product, 4-doxylpentanoic acid, is released which brings about a sharp three-line spectrum. Thus, the kinetics of phospholipase A2 activity can be followed by monitoring the increase in the ESR signal amplitude of the three-line spectrum, which is linearly proportional to the amount of 4-doxylpentanoic acid produced; no separation of the product from the substrate is needed during the measurement. The rate of hydrolysis of 1 nmol min-1 can be accurately measured within a 5-min period of time in a sample volume of 100 microliters. This new method should be useful for assaying phospholipase A2 activities in various biological systems.