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T Cell Receptor Engineering and Analysis Using the Yeast Display Platform

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Yeast Surface Display

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1319))

Abstract

The αβ heterodimeric T cell receptor (TCR) recognizes peptide antigens that are transported to the cell surface as a complex with a protein encoded by the major histocompatibility complex (MHC). T cells thus evolved a strategy to sense these intracellular antigens, and to respond either by eliminating the antigen-presenting cell (e.g., a virus-infected cell) or by secreting factors that recruit the immune system to the site of the antigen. The central role of the TCR in the binding of antigens as peptide-MHC (pepMHC) ligands has now been studied thoroughly. Interestingly, despite their exquisite sensitivity (e.g., T cell activation by as few as 1–3 pepMHC complexes on a single target cell), TCRs are known to have relatively low affinities for pepMHC, with K D values in the micromolar range. There has been interest in engineering the affinity of TCRs in order to use this class of molecules in ways similar to now done with antibodies. By doing so, it would be possible to harness the potential of TCRs as therapeutics against a much wider array of antigens that include essentially all intracellular targets. To engineer TCRs, and to analyze their binding features more rapidly, we have used a yeast display system as a platform. Expression and engineering of a single-chain form of the TCR, analogous to scFv fragments from antibodies, allow the TCR to be affinity matured with a variety of possible pepMHC ligands. In addition, the yeast display platform allows one to rapidly generate TCR variants with diverse binding affinities and to analyze specificity and affinity without the need for purification of soluble forms of the TCRs. The present chapter describes the methods for engineering and analyzing single-chain TCRs using yeast display.

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Acknowledgments

This work was supported by various NIH grants over the years (DMK), including current NIH grants P01 CA097296 (DMK), T32 GM070421 (SNS), and F30 CA180723 (DTH), and a grant from the Melanoma Research Alliance (DMK). We thank Dane Wittrup for helpful discussions, and past and current members of the Kranz lab.

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Authors and Affiliations

  1. Department of Biochemistry, University of Illinois, 600 S. Matthews Ave., Urbana, IL, 61801, USA

    Sheena N. Smith, Daniel T. Harris & David M. Kranz

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  1. Sheena N. Smith
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  2. Daniel T. Harris
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  3. David M. Kranz
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Correspondence to David M. Kranz .

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  1. UCSF, San Francisco, California, USA

    Bin Liu

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Smith, S.N., Harris, D.T., Kranz, D.M. (2015). T Cell Receptor Engineering and Analysis Using the Yeast Display Platform. In: Liu, B. (eds) Yeast Surface Display. Methods in Molecular Biology, vol 1319. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2748-7_6

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  • DOI: https://doi.org/10.1007/978-1-4939-2748-7_6

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-2747-0

  • Online ISBN: 978-1-4939-2748-7

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