Isolation, characterization and crystallization of a phospholipase A 2 myotoxin from the venom of the prairie rattlesnake ( Crotalus viridis viridis)

Abstract

A myotoxin with phospholipase A 2 (PLA 2) activity was isolated from the venom of the prairie rattlesnake ( Crotalus viridis viridis, CVV) by cation-exchange chromatography. The toxin contains 123 amino acids and has an estimated mol. wt of 14,000. It is basic, with a p I above 9. Comparison of the N-terminal 33 residues of this myotoxin with other PLA 2 proteins from snake venoms showed that CVV myotoxin has highest homology (91%) to one isoform of the B component of crotoxin from Crotalus durissus terrificus venom, and less homology (73–75%) to mojave toxin from Crotalus scutulatus scutulatus venom and agkistrotoxin from Agkistrodon halys Pallas venom. It has the least homology (40–43%) to PLA 2s from venom of two other snakes in the Crotalus genus which are neither neurotoxic nor myotoxic. CVV myotoxin induces the type of myonecrosis typical of snake venom myotoxins with the PLA 2 structure, i.e. rapid disruption of the plasma membrane as indicated by the presence of delta lesions, hypercontraction and clumping of the myofilaments, and necrosis of affected skeletal muscle cells. Inhibition of the phospholipase activity of the toxin with p-bromophenacyl bromide inhibits the myotoxic activity, indicating that for some myotoxins with the PLA 2 structure, the catalytic activity is important for myotoxic activity. This is the first report of the isolation of a non-neurotoxic, single-chain PLA 2 myotoxin from the venom of a snake from the Crotalus genus.