title: Probing HLA-B7 conformational shifts induced by peptide-binding groove mutations and bound peptide with anti-HLA monoclonal antibodies.
abstract: To determine the influence of peptide-binding groove residues and MHC-bound peptide on HLA-B7 conformation, we investigated the binding sites of nine locus- or allele-specific mAbs using a panel of 82 HLA-B7 variants. The functional mAb epitopes encircle the HLA-B7 peptide-binding groove. Three mAbs are affected by mutations at solvent-accessible peptide-binding groove mutations. Mutations in peptide-binding groove residues 45, 63, and 150 affect multiple nonoverlapping mAb epitopes, probably by interaction with other MHC residues or bound peptide. However, 18 of 24 peptide-binding groove mutations do not affect mAb binding, indicating that the conformation of solvent-accessible HLA-B7 structures is largely dissociated from changes in the peptide-binding groove. To test whether bound peptides alter HLA-B7 conformation, we loaded HLA-B7 heavy chains on acid-stripped cells with beta2-microglobulin and 20 individual synthetic peptides. Two of eight mAbs are sensitive to HLA-B7-bound peptides. A likely interpretation of these data is that the conformational flexibility of HLA-B7 is due to peptide-induced conformational shifts in MHC side chains, rather than major shifts in the MHC main chain. These results suggest that HLA-B7 conformation is largely maintained in the context of different bound peptides and different peptide-binding grooves.
authors.affiliation: Department of Pathology, University of Iowa College of Medicine, Iowa City 52242, USA.
authors.forename: K D
authors.initials: KD
authors.lastname: Smith
authors.forename: B E
authors.initials: BE
authors.lastname: Mace
authors.forename: A
authors.initials: A
authors.lastname: Valenzuela
authors.forename: J L
authors.initials: JL
authors.lastname: Vigna
authors.forename: J A
authors.initials: JA
authors.lastname: McCutcheon
authors.forename: J A
authors.initials: JA
authors.lastname: Barbosa
authors.forename: E
authors.initials: E
authors.lastname: Huczko
authors.forename: V H
authors.initials: VH
authors.lastname: Engelhard
authors.forename: C T
authors.initials: CT
authors.lastname: Lutz
mesh_terms: D000911:Antibodies, Monoclonal / Q000494:pharmacology*; D001667:Binding, Competitive / Q000235:genetics / Q000276:immunology; D002460:Cell Line; D000939:Epitopes / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D015793:HLA-B7 Antigen / Q000737:chemistry / Q000235:genetics* / Q000276:immunology*; D006801:Humans; D009154:Mutation / Q000276:immunology*; D010446:Peptide Fragments / Q000276:immunology* / Q000378:metabolism / Q000494:pharmacology*; D011485:Protein Binding / Q000235:genetics / Q000276:immunology; D011487:Protein Conformation / Q000187:drug effects*; D017510:Protein Folding
chemical_list: D000911:Antibodies, Monoclonal; D000939:Epitopes; D015793:HLA-B7 Antigen; D010446:Peptide Fragments