Enhanced immune presentation of a single-chain major histocompatibility complex class I molecule engineered to optimize linkage of a C-terminally extended peptide

Lonnie Lybarger, Y. Y. Lawrence Yu, Michael J. Miley, Daved H. Fremont, Nancy Myers, Tina Primeau, Steven M. Truscott, Janet M. Connolly, Ted H. Hansen

Research output: Contribution to journalArticlepeer-review

41 Scopus citations

Abstract

Major histocompatibility complex class I molecules can be expressed as single polypeptides wherein the antigenic peptide, β2-microglobulin, and heavy chain are attached by flexible linkers. These molecules, single-chain trimers (SCTs), are remarkably stable at the cell surface compared with native (noncovalently attached) class I molecules. In this study, we used a structure-based approach to engineer an F pocket variant SCT of the murine class I molecule Kb that presents the SIIN-FEKL epitope of ovalbumin. Mutation of heavy chain residue Tyr84 (Y84A) in the SCT resulted in enhanced serological and cytolytic CD8 T cell recognition of the covalently linked peptide due to better accommodation of the linker extending from the C terminus of the peptide. These SCTs exhibit significant cell-surface stability, which we hypothesize is rendered by their ability to continuously and efficiently rebind the covalently attached peptide. In addition, we demonstrate that SCT technology can be applied to tetramer construction using recombinant SCTs expressed in Escherichia coli. SCT-based tetramers could have applications for the enumeration of T and natural killer cells that recognize peptide-class I complexes prone to dissociation.

Original languageEnglish
Pages (from-to)27105-27111
Number of pages7
JournalJournal of Biological Chemistry
Volume278
Issue number29
DOIs
StatePublished - Jul 18 2003

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