PMID: 1194274Dec 10, 1975Paper

Kinetic analysis of phospholipase A2 activity toward mixed micelles and its implications for the study of lipolytic enzymes

The Journal of Biological Chemistry
R A DeemsE A Dennis

Abstract

A detailed kinetic scheme is proposed for the action of phospholipase A2 on mixed micelles of phospholipid and surfactant: see article. where E is the enzyme, A is the mixed micelle, and B is the phospholipid substrate in the mixed micelle. This scheme takes into account quantitatively the involvement of the lipid-water interface in the action of this enzyme toward substrate in macromolecular lipid complexes. The kinetic equation for this scheme is derived and four simplifying assumptions which are necessary for its practical application are described. Kinetic data are reported for the action of cobra venom phospholipase A2 (Naja naja naja) on 1,2-dipalmitosyl-sn-glycero-3-phosphorylcholine in mixed micelles with the nonionic surfactant Triton X-100, and these data are analyzed in terms of the kinetic equation presented. At 40 degrees, pH 8.0, and in the presence of 10 mM Ca2+, V was found to be about 4 X 10(3) mumol min(-1) mg of protein(-1). KsA, which is the dissociation constant for the enzyme-mixed micelle complex, is about 5 X 10(-4) M. KmB, the Michaelis constant for the catalytic step, which is (k-2 + k3)/k2, is 1 to 2 X 10(-10) mol cm-2. This kinetic treatment, together with the fact that the mixed micelle system allow...Continue Reading

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