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Construction and Screening of an Antigen-Derived Peptide Library Displayed on Yeast Cell Surface for CD4+ T Cell Epitope Identification

  • Fei Wen
  • Mason R. Smith
  • Huimin ZhaoEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 2024)

Abstract

Antigenic peptides (termed T cell epitopes) are assembled with major histocompatibility complex (MHC) molecules and presented on the surface of antigen-presenting cells (APCs) for T cell recognition. T cells engage these peptide-MHCs using T cell receptors (TCRs). Because T cell epitopes determine the specificity of a T cell immune response, their prediction and identification are important steps in developing peptide-based vaccines and immunotherapies. In recent years, a number of computational methods have been developed to predict T cell epitopes by evaluating peptide-MHC binding; however, the success of these methods has been limited for MHC class II (MHCII) due to the structural complexity of MHCII antigen presentation. Moreover, while peptide-MHC binding is a prerequisite for a T cell epitope, it alone is not sufficient. Therefore, T cell epitope identification requires further functional verification of the MHC-binding peptide using professional APCs, which are difficult to isolate, expand, and maintain. To address these issues, we have developed a facile, accurate, and high-throughput method for T cell epitope mapping by screening antigen-derived peptide libraries in complex with MHC protein displayed on yeast cell surface. Here, we use hemagglutinin and influenza A virus X31/A/Aichi/68 as examples to describe the key steps in identification of CD4+ T cell epitopes from a single antigenic protein and the entire genome of a pathogen, respectively. Methods for single-chain peptide MHC vector design, yeast surface display, peptide library generation in Escherichia coli, and functional screening in Saccharomyces cerevisiae are discussed.

Key words

CD4+ T cell epitope mapping Peptide library Major histocompatibility complex MHC-binding peptides Human leukocyte antigen DR1 (HLA-DR1) Yeast display Influenza A virus Flow cytometry High-throughput screening 

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Department of Chemical and Biomolecular EngineeringUniversity of Illinois at Urbana-ChampaignUrbanaUSA
  2. 2.Department of Chemical EngineeringUniversity of MichiganAnn ArborUSA
  3. 3.Carl R. Woese Institute for Genomic BiologyUniversity of Illinois at Urbana-ChampaignUrbanaUSA
  4. 4.Department of Chemistry, BiochemistryUniversity of Illinois at Urbana-ChampaignUrbanaUSA
  5. 5.Department of BioengineeringUniversity of Illinois at Urbana-ChampaignUrbanaUSA

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