MELITTIN BINDS TO SECRETORY PHOSPHOLIPASE A(2) AND INHIBITS ITS ENZYMATIC-ACTIVITY

Synthetic melittin inhibited the enzymatic activity of secretory-phosp holipase A(2) (PLA(2)) from various sources, including bee and snake v enoms, bovine pancreas, and synovial fluid from rheumatoid arthritis p atients, irrespective of substrate (e.g., [C-14]-phosphatidylcholine o r phosphatidylethanolamine vesicles and [H-3]-oleic acid-labeled E.col i). A Lineweaver-Burk analysis showed that melittin was a noncompetiti ve inhibitor of bee venom PLA(2), causing a change in V-max from 200 t o 50 units/min/mg of protein. The K-m remained unchanged (0.75 nmole). Melittin inhibited approximately 50% of purified bee venom PLA(2) act ivity in a 30:1 molar ratio (melittin:enzyme). Because the enzyme kine tics indicated a PLA(2)-melittin interaction, a melittin-sepharose aff inity column was used to purify a PLA(2) from human serum. Further, an enzyme-linked assay was developed to quantitate PLA(2) activity in bi ological fluids using avidin-peroxidase and ELISA plates coated with b iotinylated melittin. These observations may have potential therapeuti c significance, as well as provide a convenient basis for the isolatio n and quantitation of PLA(2). (C) 1997 Academic Press.