Heparin prevents the binding of phospholipase A2 to phospholipid micelles: importance of the amino-terminus.
The activity of the major isoform of porcine pancreatic phospholipase A2 (PLA2), designated B-PLA2, against micellar substrates is inhibited by heparin. Inhibition is a consequence of binding of the enzyme to heparin, documented by a heparin-induced alteration in the intrinsic fluorescence of B-PLA2 and in the 8-anilino-1-naphthalene sulfonate fluorescence and by the enhanced rate of chemical modification of the active site residue His-48. As a consequence of heparin binding, the conformation of B-PLA2 at the active site and at the amino-terminus is altered, and the enzyme does not bind to phospholipid micelles. In spite of the heparin-induced conformational changes, B-PLA2 retains its ability to catalyze the hydrolysis of monomeric phospholipid. Other glycosaminoglycans can bind to and inhibit the activity of B-PLA2 toward organized phospholipids, but none tested is as effective as heparin. An isoform of the pancreatic enzyme, designated UB-PLA2 and which corresponds to iso-pig PLA2, does not bind to nor is its catalytic activity influenced by heparin. A peptide corresponding to the amino-terminal 26 residues of B-PLA2 can rescue PLA2 from heparin inhibition. A similar peptide corresponding to the amino-terminus of UB-PLA2 has no effect on heparin inhibition. A model for the inhibition of B-PLA2 by heparin is proposed in which the catalytically significant effect of heparin is to interact directly with the amino-terminus of B-PLA2, the interfacial recognition site, to prevent the enzyme from binding to micellar substrates.
Diccianni
M B
MB
Department of Pharmacology and Cell Biophysics, College of Medicine, University of Cincinnati, Ohio 45267-0575.
Lilly-Stauderman
M
M
McLean
L R
LR
Balasubramaniam
A
A
Harmony
J A
JA
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites / Q000187:drug effects; D002384:Catalysis; D004347:Drug Interactions; D006493:Heparin / Q000494:pharmacology*; D006868:Hydrolysis; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000187:drug effects; D008823:Micelles; D008969:Molecular Sequence Data; D010741:Phospholipases A / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000187:drug effects; D054467:Phospholipases A2; D010743:Phospholipids / Q000737:chemistry*; D011485:Protein Binding / Q000187:drug effects; D011487:Protein Conformation; D012996:Solutions; D013329:Structure-Activity Relationship; D013552:Swine
D007527:Isoenzymes; D008823:Micelles; D010743:Phospholipids; D012996:Solutions; D006493:Heparin; D010741:Phospholipases A; D054467:Phospholipases A2