High throughput pMHC-I tetramer library production using chaperone-mediated peptide exchange.

Autor:	Overall SA; Department of Chemistry and Biochemistry, University of California Santa Cruz, 1156 High St., Santa Cruz, CA, 95064, USA., Toor JS; Department of Chemistry and Biochemistry, University of California Santa Cruz, 1156 High St., Santa Cruz, CA, 95064, USA., Hao S; Technology Innovation Lab, New York Genome Center, 101 6th Ave, New York, NY, 10013, USA., Yarmarkovich M; Division of Oncology, Center for Childhood Cancer Research, Children's Hospital of Philadelphia and Department of Pediatrics, Perelman School of Medicine, University of Pennsylvania, 3400 Civic Center Blvd, Philadelphia, PA, 19104, USA., Sara M O'Rourke; Department of Chemistry and Biochemistry, University of California Santa Cruz, 1156 High St., Santa Cruz, CA, 95064, USA., Morozov GI; Department of Chemistry and Biochemistry, University of California Santa Cruz, 1156 High St., Santa Cruz, CA, 95064, USA., Nguyen S; Department of Microbiology, Perelman School of Medicine, University of Pennsylvania, 3400 Civic Center Blvd, Philadelphia, PA, 19104, USA., Japp AS; Department of Microbiology, Perelman School of Medicine, University of Pennsylvania, 3400 Civic Center Blvd, Philadelphia, PA, 19104, USA., Gonzalez N; Department of Chemistry and Biochemistry, University of California Santa Cruz, 1156 High St., Santa Cruz, CA, 95064, USA., Moschidi D; Department of Chemistry and Biochemistry, University of California Santa Cruz, 1156 High St., Santa Cruz, CA, 95064, USA., Betts MR; Department of Microbiology, Perelman School of Medicine, University of Pennsylvania, 3400 Civic Center Blvd, Philadelphia, PA, 19104, USA., Maris JM; Division of Oncology, Center for Childhood Cancer Research, Children's Hospital of Philadelphia and Department of Pediatrics, Perelman School of Medicine, University of Pennsylvania, 3400 Civic Center Blvd, Philadelphia, PA, 19104, USA., Smibert P; Technology Innovation Lab, New York Genome Center, 101 6th Ave, New York, NY, 10013, USA., Sgourakis NG; Department of Chemistry and Biochemistry, University of California Santa Cruz, 1156 High St., Santa Cruz, CA, 95064, USA. nsgourak@ucsc.edu.
Jazyk:	angličtina
Zdroj:	Nature communications [Nat Commun] 2020 Apr 20; Vol. 11 (1), pp. 1909. Date of Electronic Publication: 2020 Apr 20.
DOI:	10.1038/s41467-020-15710-1
Abstrakt:	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.
Databáze:	MEDLINE
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