The infected cells present viral antigens to the CTLs through MHC class I, leading to the generation of immune response and destruction of infected cells. Antigen presentation to the CTL involve, proteasomal cleavage, transport to the ER, HLA binding, and presentation to TCR. Response of the CTLs with particular HLA.
Objectives: To study Mutations in gag gene of HIV-1 and their effect on proteosomal degradation of peptides.Association of HLA types with selected viral mutations and its effect on the immune response.
Methodology: Samples were collected from patients followed by DNA extraction which was used in HLA typing and in Nested PCR to amplify the HIV gag gene followed by sequencing . The sequences were analyzed using bioinformatics tools to identify mutations and for potential proteosomal degradation sites using Netchop. The selected peptides containing the proteosomal degradation sites were synthesized and used in protoeosomal degradation assay. Digested peptides were run on SDS gel to observe the degradation pattern . The digest products were send for MALDI-MS to confirm the exact cutting sites of the peptides.
We identified T303V mutation which is a stronger proteosomal degradation site as compared to T303T in majority of the sequences that we retrieved from database . The in-vitro proteosomal degradation assay and MALDI-MS also support these results. The HLA types of the patients were relatively diverse but HLA A*68 was found more prevalent than other HLA types.
The HIV infection in the region is relatively new and the virus is evolving according to the local environment and immune pressure by the patients. We have identified T303V mutation in locally spread virus. We intend to use the peptide containing this particular residue in ELISpot experiments to further elucidate the interaction between HIV and the host immune system.