IDENTIFICATION AND CHARACTERIZATION OF THREE SERPINS IN FASCIOLA HEPATICA.
Autor(es): Lucía Laura Sánchez Di Maggio, Lucas Tirloni, Dinorah Gambetta, Uruguaysito Benavides, Carlos Carmona, Itabajara da Silva Vaz Junior, Monica Patricia Berasain Brandolini
IDENTIFICATION AND CHARACTERIZATION OF THREE SERPINS IN FASCIOLA HEPATICA.
» Área de pesquisa: HELMINTOLOGIA
» Instituição: Instituto de Higiene - UDELAR
» Agência de fomento e patrocinadores: Dicyt-CNPq, CAPES-UDELAR, PEDECIBA, INCT-Entomologia Molecular, FAPERGS
The liver fluke Fasciola hepatica is the causative agent of fascioliasis. It leads to significant economic losses in the livestock industries in addition to becoming an emerging pathogen for humans in many countries. Serpins are a superfamily of inhibitors regulating inflammation, complement activation, blood coagulation and fibrinolysis. Here we present sequence and in silico analysis of three F. hepatica serpins regarding their role in parasite-host relationship. Serpin encoding sequences were retrieved from a public database and specific primers were designed. Total RNA were extracted from eggs, newly excysted juvenile (NEJ) and adults and cDNA were synthetized. Amplicons from PCR were purified and cloned into pGEM-T vector and identity confirmed by DNA sequencing. The deduced amino acid sequences were scanned against Genebank using the BLASTp algorithm. Alignment was performed using ClustalW algorithm in the MEGA6.06 program. In silico models for tertiary structure were built with Phyre2 and Swiss-Model softwares. It was demonstrated the existence of three serpin encoding genes (FhSPN1, FhSPN2 and FhSPN4) with 1125pb, 1205 pb and 1227pb long ORFs, respectively. FhSPN4 (409aa) has similarity with serpins from Schistosoma japonicum while FhSPN1 (375aa) and FhSPN2 (384aa) show similarity with sequences from Clonorchis sinensis. The 3D models obtained are consistently with the predicted tertiary structure for serpins. Peptide sequences compatible with FhSPN1 and FhSPN2 have been identified by proteomics assays for excretory/secretory products. According with the avaliable data , FhSPN1 y FhSPN2 are secreted during the NEJ and adult stage, respectively, suggesting a role of these proteins in parasite-host relationship. Expression of the recombinant proteins of these three serpins are under development for biochemical and functional characterization.
