Identification of HLA-A*0201-Restricted T-Cell Epitope on MPT51 Protein, a Major Secreted Protein Derived from Mycobacterium tuberculosis using MPT51 Overlapping Peptides Screening

Department of Infectious Diseases, Department of Health Science, Department of Internal Medicine, Second Division, Hamamatsu University School of Medicine, Hamamatsu 431-3192, Japan

^* To whom correspondence should be addressed. Email: tnagata{at}hama-med.ac.jp.

	Abstract

CD8⁺ T cells play a pivotal role in protection against Mycobacterium tuberculosis infection. We identified a novel HLA-A*0201-restricted CD8⁺ T-cell epitope on a dominant secreted antigen of M. tuberculosis, MPT51, in HLA-A*0201-transgenic HHD mice. HHD mice were immunized with plasmid DNA encoding MPT51 with gene gun bombardment and interferon (IFN)- production from the immune splenocytes was analyzed. In response to overlapping synthetic peptides covering the mature MPT51 sequence, only one peptide, p51-70, stimulated the splenocytes to produce IFN-. Three-color flow cytometric analysis of intracellular IFN- and cell-surface CD4 and CD8 staining revealed that MPT51 p51-70 peptide contains an immunodominant CD8⁺ T-cell epitope. Further analysis using computer algorithms permitted identification of a bona fide T-cell epitope, p53-62. MHC class I stabilization assay using T2 cells confirmed that the epitope binds to HLA-A*0201. The T cells were capable of lysing MPT51 p53-62 peptide-pulsed T2 cells. In addition, MPT51 p53-62-specific memory CD8⁺ T cells were found in tuberculin skin test-positive HLA-A*0201⁺ healthy individuals. This HLA-A*0201-restricted CD8⁺ T-cell epitope would be feasible for analysis of the role of MPT51-specific T cells in M. tuberculosis infection and for future vaccine design against tuberculosis.