THE EFFECT OF HUMAN BETA(2)-MICROGLOBULIN ON MAJOR HISTOCOMPATIBILITYCOMPLEX-I PEPTIDE LOADING AND THE ENGINEERING OF A HIGH-AFFINITY VARIANT - IMPLICATIONS FOR PEPTIDE-BASED VACCINES

The ability to directly load cell surface major histocompatibility com plex (MHC) class I molecules with peptides provides a potentially powe rful approach toward the development of vaccines to generate cell-medi ated immunity. We demonstrate that exogenous beta(2)-microglobulin (be ta(2)m) stabilizes human cell surface MHC I molecules and facilitates their loading with exogenous peptides, Additionally, using three-dimen sional crystal structures and known interaction sites between MHC I he avy chains and beta(2)m, we engineered variants of human beta(2)m (h b eta(2)m) with a single serine substitution at residue 55, This alterat ion was predicted to promote hydrophobic interactions at the MHC I hea vy chain/beta(2)m interface and displace an ordered water molecule. Co mpared with h beta(2)m, the serine to valine substitution at residue 5 5 had improved ability to bind to cell surface HLA-A1, HLA-A2, and HLA -A3 molecules, facilitate exogenous peptide loading, and promote recog nition by peptide-specific T cells. The inclusion of h beta(2)m or hig her affinity variants when pulsing cells with MHC-restricted peptides increases the efficiency of peptide loading 50-80-fold. Therefore, the inclusion of h beta(2)m in peptide-based vaccines may increase cell s urface antigen densities above thresholds that allow recognition of pe ptide antigens by the immune system, particularly for cryptic, subdomi nant, or marginally antigenic peptides.