N-terminal amino acid sequenceS of A PRE SYNAPTICALLY-ACTIVE NEUROTOXIC phospholipaseS from Bothropsneuwiedi pauloensis (jararaca-pintada) snake venom 

BORJA-OLIVEIRA, C.R.(1), DURIGON, A.M.(1), KASSAB, B.H.(2), TOYAMA, M.H.(2), NOVELLO, J.C.(2), MARANGONI, S.(2), RODRIGUES-SIMIONI, L.(1)

(1)Departamento de Farmacologia, FCM, UNICAMP, Campinas, SP, (2)Departamento de Bioquímica, IB, UNICAMP, Campinas, SP, Brasil.

OBJECTIVES: A presynaptically-activeneurotoxicphospholipaseA₂(PLA₂) have been isolated from Bothrops neuwiedi pauloensis snake venom. The toxin have PLA₂ activity and produce complete neuromuscular blockade in chick isolated biventer cervicis preparation, without inhibiting acetylcholine-induced contractures (Borja-Oliveira et al., Revista Brasileira de Toxicologia2001,14:113). In this work, we determined the N-terminal sequence of this PLA2, named neuwiedi toxin-I, and compared it with those from other presynaptic and myotoxic PLA₂ from snake venoms. METHODS AND RESULTS: B. n. pauloensis venom was fractionated by molecular exclusion HPLC (Protein Pack 300SW column) followed by reverse phase HPLC (C-18 m-Bondapak column).Tricine SDS-PAGE in the absence or presence of dithiothreitol (DTT) showed that neuwiedi toxin-I had a molecular mass of 30 kDa and 15 kDa, respectively. The N-terminal sequence  DLVQFGQMILKVAGRSLPKSYGAYGCYCGWGGRGK... showed 66% homology to the Asp49 PLA₂ myotoxin bothropstoxin-II and 54% homology with the presynaptic PLA₂ caudoxin and taipoxin (a-subunit). CONCLUSION: The fact that neuwiedi toxin-I have Gln-4, Phe-5 and Tyr-28, amino acid residues conserved in all Asp49 PLA₂ variants purified so far, and that their amino acid sequences were similar to Asp49 PLA₂ strongly suggests that neuwiedi toxin-I is a Asp49 PLA₂.

Financial support: FAPESP

CORRESPONDENCE TO:

Rodrigues-Simioni, L., Department of Pharmacology, Faculty of Medical Sciences, State University of Campinas (UNICAMP), 13083-970, Campinas, SP, Brazil, Email: simioni@unicamp.br