Biological and biochemical characterization of new basic phospholipase A2 BmTX-I isolated from Bothrops moojeni snake venom

Andrana K. Calgarotto, Daniela C.S. Damico, L. A. Ponce-Soto, Paulo A. Baldasso, Saulo L. Da Silva, Gustavo H.M.F. Souza, Marcos N. Eberlin, Sergio Marangoni

Research output: Contribution to journalArticlepeer-review

41 Scopus citations

Abstract

BmTX-I, an Asp49 phospholipase A2, was purified from Bothrops moojeni venom after only one chromatographic step using reverse-phase HPLC on μ-Bondapak C-18 column. A molecular mass of 14238.71 Da was determined by MALDI-TOF mass spectrometry. Amino acid analysis showed a high content of hydrophobic and basic amino acids as well as 14 half-cysteine residues. The BmTX-I PLA2 had a sequence of 121 residues of amino acids: DLWQFNKMIK KEVGKLPFPF YGAYGCYCGW GGRGEKPKDG TDRCCFVHDC CYKKLTGCPK WDDRYSYSWK DITIVCGEDL PCEEICECDR AAAVCFYENL GTYNKKYMKH LKPCKKADYP C and pI value 7.84, and showed a high degree of homology with basic Asp49 PLA2 myotoxins from other Bothrops venoms. BmTX-I presented PLA2 activity in the presence of a synthetic substrate and showed a minimum sigmoidal behavior, reaching its maximal activity at pH 8.0 and 35-45 °C. Maximum PLA2 activity required Ca2+ and in the presence of Mg2+, Cd2+ and Mn2+ it was reduced in presence or absence of Ca2+. Crotapotin from Crotalus durissus colillineatus rattlesnake venom has significantly inhibited (P<0.05) the enzymatic activity of BmTX-I. In vitro, the whole venom and BmTX-I caused a blockade of the neuromuscular transmission in young chick biventer cervicis preparations in a similar way to other bothrops species. In mice, BmTX-I and the whole venom-induced myonecrosis and a systemic interleukin-6 response upon intramuscular injection. Edema-forming activity was also analyzed through injection of the venom and the purified BmTX-I into the subplantar region of the right footpad. Since BmTX-I exert a strong proinflammatory effect; the enzymatic phospholipids hydrolysis might be relevant for these phenomena.

Original languageEnglish
Pages (from-to)1509-1519
Number of pages11
JournalToxicon
Volume51
Issue number8
DOIs
StatePublished - 15 Jun 2008
Externally publishedYes

Keywords

  • Asp49 phospholipase A2
  • Characterization
  • Edema-forming activity
  • Myonecrosis and interleukin-6 response
  • Neuromuscular blockade
  • Snake venom

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