PURIFICATION AND CHARACTERIZATION OF A NOVEL PHOSPHOLIPASE A(2) FROM KING COBRA (OPHIOPHAGUS HANNAH) VENOM

A novel phospholipase A(2), designated as Oh-DE-2, was isolated from t he venom of Ophiophagus hannah (king cobra) by successive chromatograp hy on SP-Sephadex C-25, DE-52, and Q-Sepharose columns. Oh-DE-2 with p I 5.1 showed an apparent molecular weight of 14 kD as revealed by SDS- PAGE and gel filtration. The amino acid sequence was homologous with t hose of PLA(2)s from Elapidae venoms. Oh-DE-2 was effectively inactiva ted by p-bromophenacyl bromide, indicating that the conserved His-48 i s essential for its enzymatic activity. However, modification of the c onserved Trp-19 did not cause a precipitous drop in the enzymatic acti vity of Oh-DE-2 as observed with PLA(2)s from Naja naja atra and Bunga rus multicinctus venoms. A quenching study showed that the microenviro nment of Trp in Oh-DE-2 was inaccessible to acrylamide, iodide, or ces ium, a finding which was different from those observed with PLA(2)s fr om N. naja atra and B. multicinctus venoms. These results might sugges t that, unlike other PLA(2) enzymes, Trp-19 in Oh-DE-2 is not directly involved in its enzymatic mechanisms.