RESUMO
The Lansberg's hognose pitviper, Porthidium lansbergii lansbergii, inhabits northern Colombia. A recent proteomic characterization of its venom (J. Proteomics [2015] 114, 287-299) revealed the presence of phospholipases A2 (PLA2) accounting for 16.2% of its proteins. The two most abundant PLA2s were biochemically and functionally characterized. Pllans-I is a basic, dimeric enzyme with a monomer mass of 14,136 Da, while Pllans-II is an acidic, monomeric enzyme of 13,901 Da. Both have Asp49 in their partial amino acid sequences and, accordingly, are catalytically active upon natural or synthetic substrates. Nevertheless, these two enzymes differ markedly in their bioactivities. Pllans-I induces myonecrosis, edema, and is lethal by intracerebro-ventricular injection in mice, as well as cytolytic and anticoagulant in vitro. In contrast, Pllans-II is devoid of these effects, except for the induction of a moderate edema. In spite of lacking myotoxicity, Pllans-II enhances the muscle damaging action of Pllans-I in vivo. Altogether, results further illustrate the divergent functional profiles of basic and acidic PLA2s in viperid venoms, and suggest that Pllans-I plays a myotoxic role in envenomings by P. l. lansbergii, whereas Pllans-II, apparently devoid of toxicity, enhances muscle damage caused by Pllans-I.
Assuntos
Fosfolipases A2/metabolismo , Venenos de Víboras/enzimologia , Sequência de Aminoácidos , Animais , Linhagem Celular , Cromatografia Líquida de Alta Pressão , Cromatografia de Fase Reversa , Eletroforese em Gel de Poliacrilamida , Feminino , Concentração de Íons de Hidrogênio , Masculino , Camundongos , Fosfolipases A2/química , Fosfolipases A2/toxicidade , Homologia de Sequência de Aminoácidos , Venenos de Víboras/toxicidade , ViperidaeRESUMO
The venom of the Lansberg's hognose pitviper, Porthidium lansbergii lansbergii, a species found in the northern region of Colombia, is poorly known. Aiming to increase knowledge on Porthidium species venoms, its proteomic analysis and functional evaluation of in vitro and in vivo activities relevant to its toxicity were undertaken. Out of 51 protein components resolved by a combination of RP-HPLC and SDS-PAGE, 47 were assigned to 12 known protein families. In similarity with two previously characterized venoms from species within this genus, Porthidium nasutum and Porthidium ophryomegas, that of P. lansbergii lansbergii was dominated by metalloproteinases, although in lower proportion. A common feature of the three Porthidium venoms appears to be a high content of disintegrins. Proteins not previously observed in Porthidium venoms belong to the vascular endothelium growth factor, phosphodiesterase, and phospholipase B families. P. lansbergii lansbergii venom showed relatively weak lethal activity to mice, and induced a moderate local myotoxicity, but considerable hemorrhage. Its isolated VEGF component showed potent edema-inducing activity in the mouse footpad assay. Significant thrombocytopenia, but no other major hematological changes, were observed in envenomed mice. In vitro, this venom lacked coagulant effect on human plasma, and induced a potent inhibition of platelet aggregation which was reproduced by its purified disintegrin components. Phospholipase A2 and proteolytic activities were also demonstrated. Overall, the compositional and functional data herein described for the venom of P. lansbergii lansbergii may contribute to a better understanding of envenomings by this pitviper species, for which specific clinical information is lacking. BIOLOGICAL SIGNIFICANCE: Porthidium lansbergii lansbergii is estimated to be responsible for nearly 20% of snakebite envenoming cases at the Atlantic Department of Colombia, but the identity and functional properties of its venom components are largely unknown. This study provides the first combined proteomic and functional analyses of the venom of this pitviper, which may contribute to a better understanding of the features of envenomings by this species.