title: Molecular cloning and pharmacological properties of an acidic PLA2 from Bothrops pauloensis snake venom.
abstract: In this work, we describe the molecular cloning and pharmacological properties of an acidic phospholipase A(2) (PLA(2)) isolated from Bothrops pauloensis snake venom. This enzyme, denominated BpPLA(2)-TXI, was purified by four chromatographic steps and represents 2.4% of the total snake venom protein content. BpPLA(2)-TXI is a monomeric protein with a molecular mass of 13.6 kDa, as demonstrated by Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF) analysis and its theoretical isoelectric point was 4.98. BpPLA(2)-TXI was catalytically active and showed some pharmacological effects such as inhibition of platelet aggregation induced by collagen or ADP and also induced edema and myotoxicity. BpPLA(2)-TXI displayed low cytotoxicity on TG-180 (CCRF S 180 II) and Ovarian Carcinoma (OVCAR-3), whereas no cytotoxicity was found in regard to MEF (Mouse Embryonic Fibroblast) and Sarcoma 180 (TIB-66). The N-terminal sequence of forty-eight amino acid residues was determined by Edman degradation. In addition, the complete primary structure of 122 amino acids was deduced by cDNA from the total RNA of the venom gland using specific primers, and it was significantly similar to other acidic D49 PLA(2)s. The phylogenetic analyses showed that BpPLA(2)-TXI forms a group with other acidic D49 PLA(2)s from the gender Bothrops, which are characterized by a catalytic activity associated with anti-platelet effects.
authors.affiliation: Institute of Genetics and Biochemistry, Federal University of Uberlândia, UFU, 38400902 Uberlândia-MG, Brazil. veridiana@ingeb.ufu.br.
authors.forename: Francis Barbosa
authors.initials: FB
authors.lastname: Ferreira
authors.forename: Mário Sérgio Rocha
authors.initials: MS
authors.lastname: Gomes
authors.forename: Dayane Lorena Naves
authors.initials: DL
authors.lastname: de Souza
authors.forename: Sarah Natalie Cirilo
authors.initials: SN
authors.lastname: Gimenes
authors.forename: Letícia Eulalio
authors.initials: LE
authors.lastname: Castanheira
authors.forename: Márcia Helena
authors.initials: MH
authors.lastname: Borges
authors.forename: Renata Santos
authors.initials: RS
authors.lastname: Rodrigues
authors.forename: Kelly Aparecida Geraldo
authors.initials: KA
authors.lastname: Yoneyama
authors.forename: Maria Inês Homsi
authors.initials: MI
authors.lastname: Brandeburgo
authors.forename: Veridiana M
authors.initials: VM
authors.lastname: Rodrigues
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D017837:Bothrops; D002460:Cell Line; D045744:Cell Line, Tumor; D003001:Cloning, Molecular; D003402:Creatine Kinase / Q000097:blood; D018076:DNA, Complementary / Q000235:genetics; D004487:Edema / Q000139:chemically induced; D006801:Humans; D008297:Male; D051379:Mice; D008969:Molecular Sequence Data; D018482:Muscle, Skeletal / Q000187:drug effects / Q000473:pathology; D054467:Phospholipases A2 / Q000235:genetics / Q000302:isolation & purification / Q000494:pharmacology; D010974:Platelet Aggregation / Q000187:drug effects; D010975:Platelet Aggregation Inhibitors / Q000302:isolation & purification / Q000494:pharmacology; D014757:Viper Venoms / Q000737:chemistry* / Q000494:pharmacology
chemical_list: D018076:DNA, Complementary; D010975:Platelet Aggregation Inhibitors; D014757:Viper Venoms; D003402:Creatine Kinase; D054467:Phospholipases A2