Direct evidence for conformational dynamics in major histocompatibility complex class I molecules

Abstract

Major histocompatibility complex class I molecules (MHC I) help protect jawed vertebrates by binding and presenting im-munogenic peptides to cytotoxic T lymphocytes. Peptides are selected from a large diversity present in the endoplasmic reticulum. However, only a limited number of peptides complement the polymorphic MHC specificity determining pockets in a way that leads to high-affinity peptide binding and efficient antigen presentation. MHC I molecules possess an intrinsic ability to discriminate between peptides, which varies in efficiency between allotypes, but the mechanism of selection is unknown. Elucidation of the selection mechanism is likely to benefit future immune-modulatory therapies. Evidence suggests peptide selection involves transient adoption of alternative, presumably higher energy conformations than native peptide–MHC complexes. However, the instability of peptide-receptive MHC molecules has hindered characterization of such conformational plasticity. To investigate the dynamic nature of MHC, we refolded MHC proteins with peptides that can be hydrolyzed by UV light and thus released. We compared the resultant peptide-receptive MHC molecules with non-hydrolyzed peptide-loaded MHC complexes by monitoring the exchange of hydrogen for deuterium in solution. We found differences in hydrogen– deuterium exchange between peptide-loaded and peptide-receptive molecules that were negated by the addition of peptide to peptide-receptive MHC molecules. Peptide hydrolysis caused significant increases in hydrogen– deuterium exchange in sub-regions of the peptide-binding domain and smaller increases elsewhere, including in the 3 domain and the non-covalently associated β2-microglobulin molecule, demonstrating long-range dynamic communication. Comparing two MHC allotypes revealed allotype-specific differences in hydrogen– deuterium exchange, consistent with the notion that MHC I plasticity underpins peptide selection.