Identification, chimerical construction and properties of the lethal factor from plumieri

Costa, F. L. S.1; De Lima, M. E.1; Figueiredo, S. G.2; Kalapothakis, E.3; Salas, C. E.1

1Departamento de Bioquímica e Imunologia, Instituto de Ciências Biológicas, Universidade Federal de Minas Gerais, Belo Horizonte, Brazil; 2Departamento de Ciências Fisiológicas, Universidade Federal do Espírito Santo, Vitória, Brazil; 3Departamento de Biologia Geral, Instituto de Ciências Biológicas, Universidade Federal de Minas Gerais, Belo Horizonte, Brazil.

Introduction: Lethal factor is a multifunctional protein from venom elaborated by the : Scorpaenoidei order. It comprises two heterodimers, termed α- and β-subunit; which elicits an array of biological responses, particularly a potent hypotensive effect that appears to be mediated by the nitric oxide pathway. Objective: The present work describes the identification, cDNA cloning, sequencing, chimerical construction and evaluating the in silico properties of Sp- Tx, the putative lethal factor from the venom gland of the scorpionfish Scorpaena plumieri. Materials and methods: PCR was used to amplify α- and β-subunits from the excised cDNA library using primers designed from previously identified stonefish toxins, conserved sequences. The DNA structure was elucidated and fragments obtained were assembled into contiguous sequences using tools available in BLAST. Results and Discussion: The deduced amino acid sequence of each subunit has 702 amino acid residues and neither of them shows significant homology with any known protein, except for toxins described in Scorpaeniformes. Protein sequence alignment between subunits displays an overall identity of 54% and a similarity of 76%, this high homology implies that they may have arisen from a common ancestor. The subunits of this novel toxin lack typical N-terminal signal sequences, but, potential glycosylation sites were identified. As reported in Scorpaeniformes toxins, the α and β toxin also display a B30.2/SPRY domain (comprising nearly 200 amino acid residues) in the C-terminal region. Conclusion: These data indicate the presence of a putative toxin protein whose primary structure is alike other toxins and with potential for production of antivenom against scorpionfish envenomation in Brazil. As a prelude to structure- function studies and to the best of our knowledge, this is the first report of a fully identified putative sequence of scorpionfish-derived toxin.

Keywords: Scorpionfish, Scorpaena plumieri, cDNA libray, lethal factor. Financial support: CNPq, INCTTOX-FAPESP, MCT/FUNDEP.