Prospection, structural analysis and phylogenetic relationships of endogenous γ-phospholipase A 2 inhibitors in Brazilian Bothrops snakes (Viperidae, Crotalinae)

Abstract

During the last 20 years, there have been an increasing number of reports on endogenous phospholipase A 2 inhibitors (PLIs) in the sera of snakes. These studies have demonstrated the existence of three different structural classes of PLIs (α, β and γ). The γ class members are potent inhibitors of phospholipases A 2 (PLA 2) from the venom of Viperidae snakes. These enzymes, together with the mammalian pro-inflammatory PLA 2, belong to the IIA class of the PLA 2-superfamily. Although coming from distinct sources, these phospholipases A 2 share main structural features. For this reason, γPLIs have been considered as potential models for the development of selective inhibitors of pro-inflammatory PLA 2 in humans. In spite of the rich diversity of the ophidian fauna in Brazil, only two γPLI representatives, from Crotalus durissus terrificus and Lachesis muta, have been described in Brazilian snakes so far. Here we investigated the presence of transcripts of novel γPLIs in six Bothrops species (Viperidae, Crotalinae) commonly found in our country: Bothrops alternatus, Bothrops erythromelas, Bothrops jararaca, Bothrops jararacussu, Bothrops moojeni and Bothrops neuwiedi. γPLI transcripts were present in every species analysed. The deduced mature proteins possessed 181 amino acid residues following a 19-residue signal peptide, similar to the γPLIs from C. d. terrificus taken as our model, with the exception of the deduced proteins from B. erythromelas and B. neuwiedi snakes. In these particular cases, an insertion of 4-amino acid residues was consistently present. A Bayesian tree was obtained for the Brazilian Bothrops γPLIs, showing four clusters: (1) L. muta and B. jararacussu, (2) B. alternatus, (3) B. erythromelas and B. neuwiedi, (4) B. jararaca and B. moojeni. Detailed structural analysis and further comparisons of these novel Bothrops inhibitors with γPLIs from New and Old World snakes are provided.