Two phospholipase A(2) inhibitors from the plasma of Cerrophidion (Bothrops) godmani which selectively inhibit two different group-II phospholipase A(2) myotoxins from its own venom: isolation, molecular cloning and biological properties

Myotoxic phospholipases A(2) (PLA(2)s; group II) account for most of the mu scle-tissue damage that results from envenomation by viperid snakes. In the venom of the Godman's viper (Cerrophidion godmani, formerly Bothrops godma ni), an enzymically active PLA(2) (myotoxin I) and an inactive, Lys-49 vari ant (myotoxin II) induce extensive muscle damage and oedema. In this study, two distinct myotoxin inhibitor proteins of C. godmani, CgMIP-I and CgMIP- II, were purified directly from blood plasma by selective binding to affini ty columns containing either myotoxin I or myotoxin II, respectively. Both proteins are glycosylated, acidic (pI = 4) and composed of 20-25-kDa subuni ts that form oligomers of 110 kDa (CgMIP-I) or 180 kDa (CgMIP-II). In inhib ition studies, CgMIP-I specifically neutralized the PLA(2) and the myotoxic , oedema-forming and cytolytic activities of myotoxins I, whereas CgMIP-II selectively inhibited the toxic properties of myotoxin II. N-terminal amino acid sequence analysis and sequencing of cDNAs encoding the two inhibitors revealed that CgMIP-I is similar to gamma-type inhibitors, which share a p attern of cysteine residues present in the Ly-6 superfamily of proteins, wh ereas CgMIP-II shares sequence identity with alpha-type inhibitors that con tain carbohydrate-recognition-like domains, also found in C-type lectins an d mammalian PLA(2) receptors. N-terminal sequencing of myotoxin I revealed a different primary structure from myotoxin II [De Sousa, Morhy, Arni, Ward , Diaz and Gutierrez (1998) Biochim. Biophys. Acta 1384, 204-208], which pr ovides insight into the nature of such pharmacological specificity.