title: Disulfide bond engineering to trap peptides in the MHC class I binding groove.
abstract: Immunodominant peptides in CD8 T cell responses to pathogens and tumors are not always tight binders to MHC class I molecules. Furthermore, antigenic peptides that bind weakly to the MHC can be problematic when designing vaccines to elicit CD8 T cells in vivo or for the production of MHC multimers for enumerating pathogen-specific T cells in vitro. Thus, to enhance peptide binding to MHC class I, we have engineered a disulfide bond to trap antigenic peptides into the binding groove of murine MHC class I molecules expressed as single-chain trimers or SCTs. These SCTs with disulfide traps, termed dtSCTs, oxidized properly in the endoplasmic reticulum, transited to the cell surface, and were recognized by T cells. Introducing a disulfide trap created remarkably tenacious MHC/peptide complexes because the peptide moiety of the dtSCT was not displaced by high-affinity competitor peptides, even when relatively weak binding peptides were incorporated into the dtSCT. This technology promises to be useful for DNA vaccination to elicit CD8 T cells, in vivo study of CD8 T cell development, and construction of multivalent MHC/peptide reagents for the enumeration and tracking of T cells-particularly when the antigenic peptide has relatively weak affinity for the MHC.
authors.affiliation: Pathology and Immunology, Washington University School of Medicine, St. Louis, MO 63110, USA.
authors.forename: Steven M
authors.initials: SM
authors.lastname: Truscott
authors.forename: Lonnie
authors.initials: L
authors.lastname: Lybarger
authors.forename: John M
authors.initials: JM
authors.lastname: Martinko
authors.forename: Vesselin E
authors.initials: VE
authors.lastname: Mitaksov
authors.forename: David M
authors.initials: DM
authors.lastname: Kranz
authors.forename: Janet M
authors.initials: JM
authors.lastname: Connolly
authors.forename: Daved H
authors.initials: DH
authors.lastname: Fremont
authors.forename: Ted H
authors.initials: TH
authors.lastname: Hansen
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001667:Binding, Competitive / Q000235:genetics / Q000276:immunology; D004220:Disulfides / Q000737:chemistry* / Q000378:metabolism; D015395:Histocompatibility Antigens Class I / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D007739:L Cells; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D011485:Protein Binding / Q000235:genetics / Q000276:immunology; D015202:Protein Engineering / Q000379:methods*
chemical_list: D004220:Disulfides; D015395:Histocompatibility Antigens Class I; D010455:Peptides