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Disulfide Bond Engineering to Trap Peptides in the MHC Class I Binding Groove¹

Steven M. Truscott^*, Lonnie Lybarger, John M. Martinko, Vesselin E. Mitaksov^*, David M. Kranz, Janet M. Connolly^*, Daved H. Fremont^* and Ted H. Hansen^2,*

^* Pathology and Immunology, Washington University School of Medicine, St. Louis, MO 63110; Cell Biology and Anatomy, University of Arizona Health Sciences Center, Tucson, AZ 85724; Microbiology, Southern Illinois University, Carbondale, IL 62901; and Biochemistry, University of Illinois, Urbana, IL 61801

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.

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

¹ This work was supported by National Institutes of Health Grants AI055849, AI27568, and AI057160.

² Address correspondence and reprint requests to Dr. Ted H. Hansen, Department of Pathology and Immunology, Box 8118, Washington University School of Medicine, 660 South Euclid Avenue, St. Louis, MO 63110. E-mail address: hansen{at}pathology.wustl.edu

³ Abbreviations used in this paper: pMHC, peptide/MHC; 3KO, K^b–/–D^b–/–₂m^–/– fibroblast; ₂m, ₂-microglobulin; dtSCT, disulfide-trap single-chain trimer; EndoH, endoglycosidase H; ER, endoplasmic reticulum; [G₄S]_n, n multiples of Gly-Gly-Gly-Gly-Ser; HPV, human papillomavirus; MCMV, murine cytomegalovirus; MFI, mean fluorescence intensity; SCT, single-chain trimer.

This article has been cited by other articles:

				S. M. Truscott, X. Wang, L. Lybarger, W. E. Biddison, C. McBerry, J. M. Martinko, J. M. Connolly, G. P. Linette, D. H. Fremont, T. H. Hansen, et al. Human Major Histocompatibility Complex (MHC) Class I Molecules with Disulfide Traps Secure Disease-related Antigenic Peptides and Exclude Competitor Peptides J. Biol. Chem., March 21, 2008; 283(12): 7480 - 7490. [Abstract] [Full Text] [PDF]