High throughput pMHC-I tetramer library production using chaperone-mediated peptide exchange
| Autor: | Peter Smibert, Stephanie Hao, Nikolaos G. Sgourakis, Sara M. O'Rourke, Michael R. Betts, Giora I. Morozov, Son Nguyen, Sarah A. Overall, Mark Yarmarkovich, Alberto Sada Japp, Danai Moschidi, John M. Maris, Jugmohit S. Toor, Nicolas Gonzalez |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2020 |
| Předmět: |
0301 basic medicine
Models Molecular T-Lymphocytes Science General Physics and Astronomy Translational immunology Peptide chemical and pharmacologic phenomena Computational biology Human leukocyte antigen Major histocompatibility complex General Biochemistry Genetics and Molecular Biology Article 03 medical and health sciences Mice 0302 clinical medicine Antigen Tetramer Animals Humans Genomic library Protein Interaction Domains and Motifs lcsh:Science Alleles Gene Library chemistry.chemical_classification Immunity Cellular Multidisciplinary biology Immunochemistry T-cell receptor Histocompatibility Antigens Class I Membrane Transport Proteins General Chemistry Cellular immunity 030104 developmental biology chemistry Chaperone (protein) biology.protein MHC class I lcsh:Q Carrier Proteins Peptides 030215 immunology Molecular Chaperones |
| Zdroj: | Nature Communications, Vol 11, Iss 1, Pp 1-13 (2020) Nature Communications |
| ISSN: | 2041-1723 |
| Popis: | Peptide exchange technologies are essential for the generation of pMHC-multimer libraries used to probe diverse, polyclonal TCR repertoires in various settings. Here, using the molecular chaperone TAPBPR, we develop a robust method for the capture of stable, empty MHC-I molecules comprising murine H2 and human HLA alleles, which can be readily tetramerized and loaded with peptides of choice in a high-throughput manner. Alternatively, catalytic amounts of TAPBPR can be used to exchange placeholder peptides with high affinity peptides of interest. Using the same system, we describe high throughput assays to validate binding of multiple candidate peptides on empty MHC-I/TAPBPR complexes. Combined with tetramer-barcoding via a multi-modal cellular indexing technology, ECCITE-seq, our approach allows a combined analysis of TCR repertoires and other T cell transcription profiles together with their cognate antigen specificities in a single experiment. The new approach allows TCR/pMHC interactions to be interrogated easily at large scale. Peptide-MHC (pMHC) tetramers are important tools for probing T cell repertoire and adaptive immune responses. Here the authors use a molecular chaperone, TAPBPR, to develop a high-throughput, multiplexible platform for pMHC tetramer generation to facilitate simultaneous assessments of T cell repertoire/antigen specificity and transcriptome. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|