Universal open MHC-I molecules for rapid peptide loading and enhanced complex stability across HLA allotypes.
Yi SunMichael C YoungClaire H WoodwardJulia N DanonHau V TruongSagar GuptaTrenton J WintersJoan Font-BurgadaGeorge M BurslemNikolaos G SgourakisPublished in: Proceedings of the National Academy of Sciences of the United States of America (2023)
The polymorphic nature and intrinsic instability of class I major histocompatibility complex (MHC-I) and MHC-like molecules loaded with suboptimal peptides, metabolites, or glycolipids presents a fundamental challenge for identifying disease-relevant antigens and antigen-specific T cell receptors (TCRs), hindering the development of autologous therapeutics. Here, we leverage the positive allosteric coupling between the peptide and light chain (β 2 microglobulin, β 2 m) subunits for binding to the MHC-I heavy chain (HC) through an engineered disulfide bond bridging conserved epitopes across the HC/β 2 m interface, to generate conformationally stable, peptide-receptive molecules named "open MHC-I." Biophysical characterization shows that open MHC-I molecules are properly folded protein complexes of enhanced thermal stability compared to the wild type when loaded with low- to moderate-affinity peptides. Using solution NMR, we characterize the effects of the disulfide bond on the conformation and dynamics of the MHC-I structure, ranging from local changes in β 2 m-interacting sites of the peptide-binding groove to long-range effects on the α 2-1 helix and α 3 domain. The interchain disulfide bond stabilizes MHC-I molecules in an open conformation to promote peptide exchange across multiple human leukocyte antigen (HLA) allotypes, covering representatives from five HLA-A supertypes, six HLA-B supertypes, and oligomorphic HLA-Ib molecules. Our structure-guided design, combined with conditional β-peptide ligands, provides a universal platform to generate ready-to-load MHC-I systems of enhanced stability, enabling a range of approaches to screen antigenic epitope libraries and probe polyclonal TCR repertoires covering highly polymorphic HLA-I allotypes, as well as oligomorphic nonclassical molecules.