ProPred analysis and experimental evaluation of promiscuous T-cell epitopes of three major secreted antigens of Mycobacterium tuberculosis.

In the search for safe vaccine candidates against tuberculosis (TB), subunit vaccines including peptide-based candidates deserve consideration. However, an important requirement for such vaccine candidates is their promiscuous presentation to Th1 cells mediating protective immunity against TB, i.e. Th1 cells secreting IFN-gamma. The aim of the present study was to identify promiscuous Th1 cell epitopes of three major secreted antigens of Mycobacterium tuberculosis, i.e. ESAT-6, CFP10 and MPT70 by using a virtual matrix-based prediction program (ProPred) for peptide binding to 51 HLA-DR alleles. The ProPred analysis of these proteins was performed using the server (http:www.imtech.res.in/raghava/ProPed/). The peptides predicted to bind > 50% HLA-DR alleles included in the ProPred were considered promiscuous for binding predictions. Based on this criteria, one region in ESAT-6 (aa 69-77), two regions in CFP10 (aa 55-66 and aa 76-84) and four regions in MPT70 (aa 1-11, aa 81-95, aa 124-140 and aa 182-191) were considered promiscuous HLA-DR binders. The experimental evaluation of these regions, by using overlapping synthetic peptides for presentation to T-cells, confirmed the promiscuous nature of peptides covering the regions aa 69-77, aa 76-84 and aa 182-191 of ESAT-6, CFP10 and MPT70, respectively. These results demonstrate that the ProPred analysis can facilitate the selection of promiscuous peptides recognized by Th1 cells, and thus it can be useful in the identification of peptide-based vaccine candidates against TB.