Cell surface receptors perform crucial functions in the communication of extracellular information to the inside of the cell. It is possible to study the mechanisms that dictate biological effects by engineering the affinities or kinetics of such interactions. Recently, yeast display has been used to engineer T cell receptors (TCRs) with improved stability and/or affinity. Creative Biolabs offers the most comprehensive portfolio of yeast display related services, including yeast display-based T cell receptor engineering.
T Cell Receptors and Antibodies
The αβ heterodimeric T cell receptors (TCRs) recognize peptide antigens that are conveyed to the cell surface as a complex with a protein encoded by the major histocompatibility complex (MHC). Thus, T cells have 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. T cell receptors and antibodies have developed to recognize different kinds of ligands. Antibodies function as membrane-bound and soluble proteins that bind to soluble antigens, whereas TCRs function only as membrane-bound molecules that bind to cell-associated peptide MHC antigens.
T Cell Receptor Engineering Using Yeast Display
Yeast display provides a highly effective system for engineering high-affinity proteins via a fluorescent-labeled ligand and fluorescence-activated cell sorting (FACS). Recently, yeast display has been used to engineer T cell receptors (TCRs) with improved stability and/or affinity. In the yeast display system, the TCR has been cloned as a Vβ-linker-Vα single chain (scTCR) fused to the gene for the yeast cell surface protein Aga2. This kind of fusion construct would be displayed on the surface of yeast cells, where it is amenable to in vitro engineering for higher affinity binding to the peptide-MHC ligand.
Fig 1. Diagram of scTv fragment displayed on the surface of yeast. A scTv fragment with an N-terminus HA tag and a C-terminus c-myc is depicted. The scTv construct is expressed as a fusion as an N-terminal fusion to the AGA-2 yeast mating protein, which forms covalent linkages to AGA-1 on the yeast cell surface. Expression of the construct can be monitored through antibodies against either tags (HA or c-myc) or antibodies specific for the scTv variable domains (Smith et al., 2015).
The brief procedure of yeast display-based T cell receptor engineering is performed in following steps:
1) Random Mutagenesis and Expression of 2C TCR
2) Selection of Mutant TCRs by Using Flow Cytometry
3) Estimation of TCR per Cell
4) Peptide MHC Functional Binding Assay
As mentioned above, it has been demonstrated that the yeast surface-display system could be used for genetic engineering of T cell receptors by combinatorial library screening. With years of extensive experience and advanced technology, scientists at Creative Biolabs are willing to offer T cell receptor engineering services for our worldwide clients based on our legendary yeast display platform which can improve your results and save your time in your research.
If you are interested in learning more about Creative Biolabs’ yeast display services, please contact us for more details.
All listed services and products are For Research Use Only. Do Not use in any diagnostic or therapeutic applications.