Cloning of a novel acidic phospholipase A2 from the venom gland of Crotalus durissus cascavella (Brazilian northeastern rattlesnake)

Abstract

The phospholipase A2 superfamily encompasses 15 groups that are classified into: secreted PLA2 (sPLA2); cytosolic PLA2 (cPLA2); Ca2+-independent intracellular PLA2 (iPLA2); platelet-activating factor acetylhydrolase (PAF-AH); and lysosomal PLA2. Currently, approximately 700 PLA2 sequences are known, of which 200 are obtained from the venom gland of Crotalinae snakes. However, thus far, little information is available on cloning, purification and structural characterization of PLA2 from Crotalus durisssus cascavela venom gland. In the present work, we report the molecular cloning of a novel svPLA2 from C. d. cascavella (Cdc), a predominant rattlesnake subspecies in northeastern Brazil. The Cdc svPLA2 cDNA precursor is 689 nucleotides long and encodes a protein of 138 amino acid residues, with a calculated molecular mass of approximately 13,847 Da and an estimated isoelectric point of 5.14. Phylogenetic analysis of Crotalinae PLA2 reveals that Cdc PLA2 clustered with other acidic type IIA PLA2 homologues is also present in the venom of North American rattlesnakes. Hitherto, this study presents a novel PLA2 cDNA precursor from C. d. cascavella and data reported herein will be useful for further steps in svPLA2 purification and analysis

Biological characterization of a myotoxin phosphoplipase A2 homologue purified from the venom of the snake Bothrops moojeni

“…In the present work, a myotoxin phospholipase A 2 homologue from Bothrops moojeni venom, denominated BmooMtx, was purified by a combination of ion exchange and gel filtration chromatography (Figure 1 -A and B). The molecular weight observed at SDS-PAGE (16,500) was similar to those reported for MjTX-I (13,400), M-IV (14,000), BmTX-I (14,240) and BmooTX-I (15,000), from the same venom, a presynaptic phospholipase A 2 , neuwieditoxin-I (14,000), isolated from the venom of Bothrops neuwiedi pauloensis and an acidic phospholipase A 2 (14,000) from the venom of Crotalus durissus cascavella (35,36 (Figure 2 -A). These results show that the B. moojeni venom shares some common myotoxins that induce antibodies that crossreact with crude venoms of B. alternatus and B. leucurus.…”

Proteomic analysis of the rare Uracoan rattlesnake Crotalus vegrandis venom: Evidence of a broad arsenal of toxins

“…1). Its sequence matches to a PLA 2 cloned from the C. d. cascavella venom gland (UniProtKB: C9E7C4) (Guarnieri et al, 2009). The identification of this protein species is supported by two UPs.…”

Purification procedure for the isolation of a P-I metalloprotease and an acidic phospholipase A2 from Bothrops atrox snake venom

“…PLA 2 s are classified into 15 different subgroups according to specific features, with snake venom PLA 2 s belonging to groups I and II [ 60 , 61 ]. Group IIA PLA 2 s are further divided into Asp49 PLA 2 s, enzymes that usually exhibit high catalytic activity, and Lys49 PLA 2 s with low or no enzymatic activity on artificial substrates [ 62 , 63 ]. The catalytic site of PLA 2 s is composed of the residues His48, Asp49, Tyr52 and Asp99, and the majority of these enzymes have conserved His48 and Asp49 residues that present a catalytically essential water molecule attached to their side-chains by hydrogen bonds [ 64 , 65 ].…”