Molecular characterization of an acidic phospholipase A(2) from Bothrops pirajai snake venom: synthetic C-terminal peptide identifies its antiplatelet region

• Published in: Archives of toxicology 2011-10, Vol.85 (10), p.1219
• Main Authors: Teixeira, Sabrina S, Silveira, Lucas B, da Silva, Franco M N, Marchi-Salvador, Daniela P, Silva, Jr, Floriano P, Izidoro, Luiz Fernando M, Fuly, André L, Juliano, Maria A, dos Santos, Camila R, Murakami, Mário T, Sampaio, Suely V, da Silva, Saulo L, Soares, Andreimar M
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Animals; Base Sequence; Bayes Theorem; Bothrops; Cloning, Molecular; Crotalid Venoms - enzymology; DNA, Complementary; Dose-Response Relationship, Drug; Humans; Hypotension - chemically induced; Male; Mice; Models, Molecular; Molecular Sequence Data; Peptide Fragments - pharmacology; Phospholipases A2 - chemistry; Phospholipases A2 - genetics; Phospholipases A2 - isolation & purification; Phospholipases A2 - metabolism; Phylogeny; Platelet Aggregation Inhibitors - pharmacology; Rabbits
• ISSN: 1432-0738
• DOI: 10.1007/s00204-011-0665-6

This paper describes a biochemical and pharmacological characterization of BpirPLA(2)-I, the first acidic Asp49-PLA(2) isolated from Bothrops pirajai. BpirPLA(2)-I caused hypotension in vivo, presented phospholipolytic activity upon artificial substrates and inhibitory effects on platelet aggregation in vitro. Moreover, a synthetic peptide of BpirPLA(2)-I, comprising residues of the C-terminal region, reproduced the antiplatelet activity of the intact protein. A cDNA fragment of 366 bp encompassing the mature form of BpirPLA(2)-I was cloned by reverse transcriptase-PCR of B. pirajai venom gland total RNA. A Bayesian phylogenetic analysis indicated that BpirPLA(2)-I forms a clade with other acid Asp49-PLA(2) enzymes from the Bothrops genus, which are characterized by the high catalytic activity associated with anticoagulant or hypotensive activity or both. Comparison of the electrostatic potential (EP) on the molecular surfaces calculated from a BpirPLA(2)-I homology model and from the crystallographic models of a group of close homologues revealed that the greatest number of charge inversions occurred on the face opposite to the active site entrance, particularly in the Ca(2+) ion binding loop. This observation suggests a possible relationship between the basic or acid character of PLA(2) enzymes and the functionality of the Ca(2+) ion binding loop.