Cytotoxic T lymphocytes recognize a reconstituted class I histocompatibility antigen (HLA-A2) as an allogeneic target molecule.
Recent findings suggest that peptide fragments of newly synthesized proteins may associate intracellularly with nascent chains of class I histocompatibility antigens (termed MHC-I proteins because they are encoded by genes of the major histocompatibility complex) and that these peptide adducts may be required for the folding or stability and perhaps even the transport of these proteins to the cell surface. To determine whether these proteins can be reconstituted from their separated subunits into ostensibly native molecules in the absence of added peptides, we denatured a purified human MHC-I protein (HLA-A2) with 4 M NaSCN, separated its heavy (alpha) and light (beta 2-microglobulin) chains by gel filtration, and then mixed them in the presence of a 3-fold molar excess of beta 2-microglobulin and absence of added peptides. The reconstituted protein, recovered in 10% yield, was indistinguishable from native A2 in its reactivity with a monoclonal antibody (BB7.7) and its ability to specifically activate A2-specific CD8+ T cells. Inasmuch as the reconstituted A2 contained no detectable peptide adducts (we estimate less than 1 per 100 on a molar basis, assuming peptides of 2-5 kDa), the results suggest that peptide-free A2 can be recognized by CD8+ T cells and that peptide adducts are not essential for the MHC-I protein to maintain an ostensibly native structure.
Elliott
T J
TJ
Center for Cancer Research, Massachusetts Institute of Technology, Cambridge 02139.
Eisen
H N
HN
D002460:Cell Line; D002850:Chromatography, Gel; D003165:Complement System Proteins / Q000276:immunology; D003602:Cytotoxicity, Immunologic; D004247:DNA / Q000096:biosynthesis; D004261:DNA Replication; D015789:HLA-A2 Antigen / Q000276:immunology* / Q000302:isolation & purification; D006801:Humans; D007700:Kinetics; D008213:Lymphocyte Activation; D046911:Macromolecular Substances; D008970:Molecular Weight; D011489:Protein Denaturation; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology*
D015789:HLA-A2 Antigen; D046911:Macromolecular Substances; D003165:Complement System Proteins; D004247:DNA