Clustering of class IHLA oligomers with CD8 and TCR: Three-dimensional models based on fluorescence resonance energy transfer and crystallographic data

Fluorescence resonance energy transfer (FRET) data, in accordance with late ral mobility measurements, suggested the existence of class I HLA dimers an d oligomers at the surface of live human cells, including the B lymphoblast cell line (JY) used in the present study. Intra- and intermolecular class I HLA epitope distances were measured on JY B cells by FRET using fluoropho re-conjugated Ag-binding fragments of mAbs W6/32 and L368 directed against structurally well-characterized heavy and light chain epitopes, respectivel y. Out-of-plane location of these epitopes relative to the membrane-bound B ODIPY-PC (2-(4,4-difluoro-5-(4-phenyl-1,3-butadienyl)-4-bora-3a,4a-diaza-s- indacene-3-pentanoyl)-1-hexadecanoyl-sn-glycero-3-phosphocholine) was also determined by FRET. Computer-simulated docking of crystallographic structur es of class I HLA and epitope-specific Ag-binding fragments, with experimen tally determined interepitope and epitope to cell surface distances as cons traints, revealed several sterically allowed and FRET-compatible class I HL A dimeric and tetrameric arrangements. Extension of the tetrameric class I HLA model with interacting TCR and CD8 resulted in a model of a supramolecu lar cluster that may exist physiologically and serve as a functionally sign ificant unit for a network of CD8-HLA-I complexes providing enhanced signal ing efficiency even at low MHC-peptide concentrations at the interface of e ffector and APCs. The Journal of Immunology, 2001.