High affinity T cell receptors from yeast display libraries block T cell activation by superantigens.

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TitleHigh affinity T cell receptors from yeast display libraries block T cell activation by superantigens.
Publication TypeJournal Article
Year of Publication2001
AuthorsKieke, MC, Sundberg, E, Shusta, EV, Mariuzza, RA, Wittrup, KD, Kranz, DM
JournalJ Mol Biol
Volume307
Issue5
Pagination1305-15
Date Published2001 Apr 13
ISSN0022-2836
KeywordsBinding Sites, Directed Molecular Evolution, Enterotoxins, Flow Cytometry, Lymphocyte Activation, Models, Molecular, Mutation, Peptide Library, Protein Conformation, Protein Engineering, Receptors, Antigen, T-Cell, Solubility, Staphylococcus aureus, Superantigens, Surface Plasmon Resonance, T-Lymphocytes, Cytotoxic, Yeasts
Abstract

The alphabeta T cell receptor (TCR) can be triggered by a class of ligands called superantigens. Enterotoxins secreted by bacteria act as superantigens by simultaneously binding to an MHC class II molecule on an antigen- presenting cell and to a TCR beta-chain, thereby causing activation of the T cell. The cross-reactivity of enterotoxins with different Vbeta regions can lead to stimulation of a large fraction of T cells. To understand the molecular details of TCR-enterotoxin interactions and to generate potential antagonists of these serious hyperimmune reactions, we engineered soluble TCR mutants with improved affinity for staphylococcal enterotoxin C3 (SEC3). A library of randomly mutated, single-chain TCRs (Vbeta-linker-Valpha) were expressed as fusions to the Aga2p protein on the surface of yeast cells. Mutants were selected by flow cytometric cell sorting with a fluorescent-labeled SEC3. Various mutations were identified, primarily in Vbeta residues that are located at the TCR:SEC3 interface. The combined mutations created a remodeled SEC3-binding surface and yielded a Vbeta domain with an affinity that was increased by 1000-fold (K(D)=7 nM). A soluble form of this Vbeta mutant was a potent inhibitor of SEC3-mediated T cell activity, suggesting that these engineered proteins may be useful as antagonists.

DOI10.1006/jmbi.2001.4560
Alternate JournalJ. Mol. Biol.
PubMed ID11292343
Grant ListAI42937 / AI / NIAID NIH HHS / United States
GM52801 / GM / NIGMS NIH HHS / United States
GM55767 / GM / NIGMS NIH HHS / United States
T32 GM07283 / GM / NIGMS NIH HHS / United States