Cytotoxic T-lymphocytes (CTL) can lyse infected or transformed cells through recognition of peptides presented on human leukocyte antigen (HLA) molecules. A thorough understanding of peptide-HLA interactions is needed for improvement of CTL-based immunotherapies. We observed that aspartic acid (D) and glutamic acid (E) at peptide position 4 are highly prevalent in HLA-I peptide ligands, and discovered that they interact with arginine (R) in position 65 and lysine (L) in position 66 of the α1 helix of the binding groove in HLA-A*0201 and HLA-A*2402. Since this interaction differed from well-characterized peptide-HLA anchor interactions mediated by peptide position 2 and the C-terminus, we investigated if the charged interactions between D/E in position 4 and R65 and K66 on the α1 helix of A*0201 and A*2402 are important for peptide antigen presentation. Mutations to R65 and K66 caused loss of HLA cell surface expression, reduced peptide repertoire diversity, and loss of charged interactions between D/E4 and p65-66 in both A*0201 and A*2402. In addition, mutating R65 and/or K66 altered the peptide motifs of both A*0201 and A*2402 specifically in peptide positions 1, 2 and the C-terminus. Peptide binding and stability assays revealed that D/E residues in position 4 contribute significantly to both peptide binding and stability, particularly in peptides with weaker N and C-terminal anchors. Several other HLA-I alleles and MHC molecules from different animal species also showed conservation of R65 and K66, in addition to D/E4 in bound peptide ligands, suggesting that charged interactions between D/E4 and R65/K66 represent a conserved, non-canonical pan-MHC-I anchor interaction that should be accounted for when identifying peptide targets for immunotherapies.
","abstract_html":"<p>Cytotoxic T-lymphocytes (CTL) can lyse infected or transformed cells through recognition of peptides presented on human leukocyte antigen (HLA) molecules. A thorough understanding of peptide-HLA interactions is needed for improvement of CTL-based immunotherapies. We observed that aspartic acid (D) and glutamic acid (E) at peptide position 4 are highly prevalent in HLA-I peptide ligands, and discovered that they interact with arginine (R) in position 65 and lysine (L) in position 66 of the α1 helix of the binding groove in HLA-A*0201 and HLA-A*2402. Since this interaction differed from well-characterized peptide-HLA anchor interactions mediated by peptide position 2 and the C-terminus, we investigated if the charged interactions between D/E in position 4 and R65 and K66 on the α1 helix of A*0201 and A*2402 are important for peptide antigen presentation. Mutations to R65 and K66 caused loss of HLA cell surface expression, reduced peptide repertoire diversity, and loss of charged interactions between D/E4 and p65-66 in both A*0201 and A*2402. In addition, mutating R65 and/or K66 altered the peptide motifs of both A*0201 and A*2402 specifically in peptide positions 1, 2 and the C-terminus. Peptide binding and stability assays revealed that D/E residues in position 4 contribute significantly to both peptide binding and stability, particularly in peptides with weaker N and C-terminal anchors. Several other HLA-I alleles and MHC molecules from different animal species also showed conservation of R65 and K66, in addition to D/E4 in bound peptide ligands, suggesting that charged interactions between D/E4 and R65/K66 represent a conserved, non-canonical pan-MHC-I anchor interaction that should be accounted for when identifying peptide targets for immunotherapies.</p>","abstract_has_math":false,"creators":["Jackson, Kyle","https://orcid.org/0000-0002-3711-2861
"],"institution":null,"degree_name":"Doctor of Philosophy (PhD)","degree_level":"Dissertation (PhD)","degree_discipline":null,"degree_department":null,"school":null,"contributors":["Gregory Lizee","Jagannadha K Sastry","Juan Fueyo"],"advisors":[],"committee_chairs":[],"committee_members":[],"year":2021,"date_issued":"2021-08-01T07:00:00Z","date_published":"2021-08-01T07:00:00Z","updated_at":"2026-07-24T05:50:24Z","subjects":["Antigen presentation","Human Leukocyte Antigens","HLA Class I","peptide binding stability","mass spectrometry","computational modeling","immunopeptidome","peptide repertoire diversity","molecular interactions","molecular dynamics","Immunity","Medicine and Health Sciences","Other Immunology and Infectious Disease"],"languages":[],"rights":[],"rights_urls":[],"identifier_entries":[]},"links":{"outbound_url":"https://digitalcommons.library.tmc.edu/utgsbs_dissertations/1135","outbound_label":"Repository record","outbound_source":"dc:identifier"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor","label":"Contributor","values":["Gregory Lizee","Jagannadha K Sastry","Juan Fueyo"]},{"key":"dc:creator","label":"Author","values":["Jackson, Kyle","https://orcid.org/0000-0002-3711-2861
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"]},{"key":"dc:title","label":"Title","values":["Conserved Non-Pocket Interactions Drive The Diversity of Peptide Presentation By Mhc Class I Molecules"]}]}],"canonical_facts":{"dc:contributor":["Gregory Lizee","Jagannadha K Sastry","Juan Fueyo"],"dc:creator":["Jackson, Kyle","https://orcid.org/0000-0002-3711-2861
"],"dc:date.available":["2022-08-09T07:00:00Z"],"dc:description.abstract":["Cytotoxic T-lymphocytes (CTL) can lyse infected or transformed cells through recognition of peptides presented on human leukocyte antigen (HLA) molecules. A thorough understanding of peptide-HLA interactions is needed for improvement of CTL-based immunotherapies. We observed that aspartic acid (D) and glutamic acid (E) at peptide position 4 are highly prevalent in HLA-I peptide ligands, and discovered that they interact with arginine (R) in position 65 and lysine (L) in position 66 of the α1 helix of the binding groove in HLA-A*0201 and HLA-A*2402. Since this interaction differed from well-characterized peptide-HLA anchor interactions mediated by peptide position 2 and the C-terminus, we investigated if the charged interactions between D/E in position 4 and R65 and K66 on the α1 helix of A*0201 and A*2402 are important for peptide antigen presentation. Mutations to R65 and K66 caused loss of HLA cell surface expression, reduced peptide repertoire diversity, and loss of charged interactions between D/E4 and p65-66 in both A*0201 and A*2402. In addition, mutating R65 and/or K66 altered the peptide motifs of both A*0201 and A*2402 specifically in peptide positions 1, 2 and the C-terminus. Peptide binding and stability assays revealed that D/E residues in position 4 contribute significantly to both peptide binding and stability, particularly in peptides with weaker N and C-terminal anchors. Several other HLA-I alleles and MHC molecules from different animal species also showed conservation of R65 and K66, in addition to D/E4 in bound peptide ligands, suggesting that charged interactions between D/E4 and R65/K66 represent a conserved, non-canonical pan-MHC-I anchor interaction that should be accounted for when identifying peptide targets for immunotherapies.
"],"dc:identifier":["https://digitalcommons.library.tmc.edu/utgsbs_dissertations/1135"],"dc:subject":["Antigen presentation","Human Leukocyte Antigens","HLA Class I","peptide binding stability","mass spectrometry","computational modeling","immunopeptidome","peptide repertoire diversity","molecular interactions","molecular dynamics","Immunity","Medicine and Health Sciences","Other Immunology and Infectious Disease"],"dc:title":["Conserved Non-Pocket Interactions Drive The Diversity of Peptide Presentation By Mhc Class I Molecules"],"thesis:degree_level":["Dissertation (PhD)"],"thesis:degree_name":["Doctor of Philosophy (PhD)"]},"updated_at":"2026-07-24T05:50:24Z"}