Journal article
Characterization of binding specificities of bovine leucocyte class I molecules: impacts for rational epitope discovery
University of Copenhagen1
International Livestock Research Institute2
United States Department of Agriculture3
University of Vermont4
Department of Systems Biology, Technical University of Denmark5
Center for Biological Sequence Analysis, Department of Systems Biology, Technical University of Denmark6
Immunological Bioinformatics, Center for Biological Sequence Analysis, Department of Systems Biology, Technical University of Denmark7
The binding of peptides to classical major histocompatibility complex (MHC) class I proteins is the single most selective step in antigen presentation. However, the peptide-binding specificity of cattle MHC (bovine leucocyte antigen, BoLA) class I (BoLA-I) molecules remains poorly characterized. Here, we demonstrate how a combination of high-throughput assays using positional scanning combinatorial peptide libraries, peptide dissociation, and peptide-binding affinity binding measurements can be combined with bioinformatics to effectively characterize the functionality of BoLA-I molecules.
Using this strategy, we characterized eight BoLA-I molecules, and found the peptide specificity to resemble that of human MHC-I molecules with primary anchors most often at P2 and P9, and occasional auxiliary P1/P3/P5/P6 anchors. We analyzed nine reported CTL epitopes from Theileria parva, and in eight cases, stable and high affinity binding was confirmed.
A set of peptides were tested for binding affinity to the eight BoLA proteins and used to refine the predictors of peptide-MHC binding NetMHC and NetMHCpan. The inclusion of BoLA-specific peptide-binding data led to a significant improvement in prediction accuracy for reported T. parva CTL epitopes.
For reported CTL epitopes with weak or no predicted binding, these refined prediction methods suggested presence of nested minimal epitopes with high-predicted binding affinity. The enhanced affinity of the alternative peptides was in all cases confirmed experimentally. This study demonstrates how biochemical high-throughput assays combined with immunoinformatics can be used to characterize the peptide-binding motifs of BoLA-I molecules, boosting performance of MHC peptide-binding prediction methods, and empowering rational epitope discovery in cattle.
Language: | English |
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Publisher: | Springer Berlin Heidelberg |
Year: | 2014 |
Pages: | 705-718 |
ISSN: | 14321211 and 00937711 |
Types: | Journal article |
DOI: | 10.1007/s00251-014-0802-5 |
ORCIDs: | Nielsen, Morten and 0000-0001-8363-1999 |
Allergology Amino Acid Motifs Amino Acid Sequence Animals Binding Sites Biomedical and Life Sciences Biomedicine Cattle Cell Biology Cross Reactions Epitope Mapping Epitopes Gene Function Histocompatibility Antigens Class I Human Genetics Humans Immunology Ligands Peptide Library Peptides Position-Specific Scoring Matrices Protein Binding Recombinant Proteins Reproducibility of Results beta 2-Microglobulin