Dimerization of soluble disulfide trap single-chain major histocompatibility complex class I molecules dependent on peptide binding affinity.
Stable presentation of peptide epitope by major histocompatibility complex (MHC) class I molecules is a prerequisite for the efficient expansion of CD8(+) T cells. The construction of single-chain MHC class I molecules in which the peptide, β(2)-microglobulin, and MHC heavy chain are all joined together via flexible linkers increases peptide-MHC stability. We have expressed two T cell epitopes that may be useful in leukemia treatment as single-chain MHC class I molecules, aiming to develop a system for the expansion of antigen-specific CD8(+) T cells in vitro. Disulfide trap versions of these single-chain MHC molecules were also created to improve anchoring of the peptides in the MHC molecule. Unexpectedly, we observed that soluble disulfide trap single-chain molecules expressed in eukaryotic cells were prone to homodimerization, depending on the binding affinity of the peptide epitope. The dimers were remarkably stable and efficiently recognized by conformation-specific antibodies, suggesting that they consisted of largely correctly folded molecules. However, dimerization was not observed when the disulfide trap molecules were expressed as full-length, transmembrane-anchored molecules. Our results further emphasize the importance of peptide binding affinity for the efficient folding of MHC class I molecules.
Department of Immunology, Wright-Fleming Institute, Imperial College London, London, United Kingdom.
Eleni
E
Kotsiou
Joanna
J
Brzostek
Izabela
I
Lenart
Antony N
AN
Antoniou
Julian
J
Dyson
Keith G
KG
Gould
D018528:ATP-Binding Cassette Transporters / Q000378:metabolism; D000818:Animals; D000918:Antibody Specificity / Q000276:immunology; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology; D016466:CHO Cells; D002460:Cell Line; D006224:Cricetinae; D003412:Cricetulus; D000939:Epitopes / Q000737:chemistry / Q000378:metabolism*; D057809:HEK293 Cells; D015789:HLA-A2 Antigen / Q000276:immunology / Q000378:metabolism; D015395:Histocompatibility Antigens Class I / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D010455:Peptides / Q000276:immunology / Q000378:metabolism; D017510:Protein Folding; D055503:Protein Multimerization; D001613:beta 2-Microglobulin / Q000276:immunology / Q000378:metabolism
D018528:ATP-Binding Cassette Transporters; D000939:Epitopes; D015789:HLA-A2 Antigen; D015395:Histocompatibility Antigens Class I; D010455:Peptides; D001613:beta 2-Microglobulin; C477327:transporter associated with antigen processing (TAP)