Non-Enzymatic Structural Modifications Reshape Peptide Presentation and Antigen Recognition

For cytotoxic T-cells to identify and eliminate infected and transformed cells, the adaptive immune system requires the presentation of antigenic peptides by major histocompatibility complex class I (MHC-I) molecules. These peptides, typically derived from intracellular proteins, undergo a rigorous selection process to ensure self-tolerance. However, post-translational modifications (PTMs) and non-enzymatic chemical modifications can alter peptide structure and chemistry, potentially affecting immune recognition. In this study, we investigated the impact of non-enzymatic PTMs on antigen presentation and T cell recognition. Using the well-characterized model epitope SIINFEKL, we synthesized peptide variants incorporating common non-enzymatic PTMs and assessed their effects on MHC-I binding interactions and T cell recognition. We demonstrated that non-enzymatic PTMs markedly alter MHC-I affinity of peptides in a cancer-associated immunopeptidome. Additionally, we developed a novel enrichment strategy using an alkyne-modified probe to identify sites of non-enzymatic acylation prone to MHC-I display. These findings underscore the significance of non-enzymatic PTMs in altering the immunopeptidome and modulating immune responses.