Aplicação da proteômica de Leptospira e ferramentas de imunoinformática para a identificação de candidatos ao desenvolvimento de uma vacina contra leptospirose

Abstract

Leptospirosis is a zoonosis caused by spirochetes of the Leptospira genus, which causes a significant impact on public health, with more than one million new infections and 59,000 deaths each year, moreover major economic losses in agriculture. Lack of an fully effective vaccine against the disease is one of the main factors that compromise its control, so it is urgent to develop new vaccines that is able to overcoming the limitations of vaccines currently available, that can be used in human populations globally. Therefore, the aim of this study was to fully evaluate the protein profile of Leptospira interrogans serovar Copenhageni strain M20, with in silico characterization of uncharacterized proteins and proteins that is able to induce immune response and compose a subunit vaccine. To perform proteomics, a simplified protocol for obtaining protein extract was developed followed by mass spectrometry LTQ-Orbitrap Velos coupled to liquid chromatography EASY-nLC II. The proteins found undego a full evaluation, like a taxonomic analysis to identify common proteins to other species and serovars of the bacterium; description of functional categories; design of the three-dimensional structure of important proteins in the Leptospira study and in silico characterization of uncharacterized proteins and proteins already identified as induce an immune response. Then, many servers were used to perform immunoinformatics analysis, aiming to identify and select the proteins with the major potential for vaccine target and subsequent selection of the best epitopes able to stimulating humoral and cellular immunity. Proteins Loa 22, LipL32, Flagellin, Elongation fator Tu and Elongation fator Ts had their three-dimensional structure developed and validated. Among the selected proteins with the major vaccine target potential, most are uncharacterized proteins until then. This study also identified epitopes of B cell, of cytotoxic T lymphocyte, of helper T lymphocyte, of TCD4 cell and of IFN-γ inducers. This epitopes have been assembled through rational design to constitute a multi-epitope chimeric protein that can be used as a vaccine target with potential for overcome the bacterin limitations. The detailed study of Leptospira protein profile associated with the screening of vaccine targets by immunoinformatics resulted in the design of a chimeric protein, with desirable characteristics for an immunogen in the vaccine development process.