cDNA and deduced primary structure of basic phospholipase A(2) with neurotoxic activity from the venom secretion of the Crotalus durissus collilineatus rattlesnake

Author(s)

Fagundes, F. H. R. ; Oliveira, M. ^[1] ; Huancahuire-Vega, S. ; Romero-Vargas, F. F. ; Ponce-Soto, L. A. ^[2] ; Marangoni, S.

Affiliation

^[1] Feldmann Labs, Sao Paulo - Brazil

^[2] Univ Estadual Campinas, Univ Estadual Campinas, Inst Biol, Dept Bioquim, BR-13083970 Campinas, SP - Brazil

• Document type: Journal article
• Source: Brazilian Journal of Medical and Biological Research; v. 43, n. 3, p. 262-270, MAR 2010.
• Web of Science Citations: 1

Abstract

To illustrate the construction of precursor complementary DNAs, we isolated mRNAs from whole venom samples. After reverse transcription polymerase chain reaction (RT-PCR), we amplified the cDNA coding for a neurotoxic protein, phospholipase A(2) D49 (PLA(2) D49), from the venom of Crotalus durissus collilineatus (Cdc PLA(2)). The cDNA encoding Cdc PLA(2) from whole venom was sequenced. The deduced amino acid sequence of this cDNA has high overall sequence identity with the group II PLA(2) protein family. Cdc PLA(2) has 14 cysteine residues capable of forming seven disulfide bonds that characterize this group of PLA(2) enzymes. Cdc PLA(2) was isolated using conventional Sephadex G75 column chromatography and reverse-phase high performance liquid chromatography (RP-HPLC). The molecular mass was estimated using matrix-assisted laser desorption ionization-time-of-flight (MALDI-TOF) mass spectrometry. We tested the neuromuscular blocking activities on chick biventer cervicis neuromuscular tissue. Phylogenetic analysis of Cdc PLA(2) showed the existence of two lines of N6-PLA(2), denominated F24 and S24. Apparently, the sequences of the New World's N6-F24-PLA(2) are similar to those of the agkistrodotoxin from the Asian genus Gloydius. The sequences of N6-S24-PLA(2) are similar to the sequence of trimucrotoxin from the genus Protobothrops, found in the Old World. (AU)