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

Printer-friendly versionPrinter-friendly versionPDF versionPDF version
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
Date Published2001 Apr 13
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

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.

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