title: Monoclonal antibodies to HLA-E bind epitopes carried by unfolded β2 m-free heavy chains.
abstract: Since HLA-E heavy chains accumulate free of their light β2 -microglobulin (β2 m) subunit, raising mAbs to folded HLA-E heterodimers has been difficult, and mAb characterization has been controversial. Herein, mAb W6/32 and 5 HLA-E-restricted mAbs (MEM-E/02, MEM-E/07, MEM-E/08, DT9, and 3D12) were tested on denatured, acid-treated, and natively folded (both β2 m-associated and β2 m-free) HLA-E molecules. Four distinct conformations were detected, including unusual, partially folded (and yet β2 m-free) heavy chains reactive with mAb DT9. In contrast with previous studies, epitope mapping and substitution scan on thousands of overlapping peptides printed on microchips revealed that mAbs MEM-E/02, MEM-E/07, and MEM-E/08 bind three distinct α1 and α2 domain epitopes. All three epitopes are linear since they span just 4-6 residues and are "hidden" in folded HLA-E heterodimers. They contain at least one HLA-E-specific residue that cannot be replaced by single substitutions with polymorphic HLA-A, HLA-B, HLA-C, HLA-F, and HLA-G residues. Finally, also the MEM-E/02 and 3D12 epitopes are spatially distinct. In summary, HLA-E-specific residues are dominantly immunogenic, but only when heavy chains are locally unfolded. Consequently, the available mAbs fail to selectively bind conformed HLA-E heterodimers, and HLA-E expression may have been inaccurately assessed in some previous oncology, reproductive immunology, virology, and transplantation studies.
authors.affiliation: Laboratory of Immunology, Regina Elena National Cancer Institute, Rome, Italy.
authors.forename: Elisa
authors.initials: E
authors.lastname: Tremante
authors.affiliation: Laboratory of Immunology, Regina Elena National Cancer Institute, Rome, Italy.
authors.forename: Elisa
authors.initials: E
authors.lastname: Lo Monaco
authors.affiliation: Laboratory of Immunology, Regina Elena National Cancer Institute, Rome, Italy.
authors.forename: Tiziano
authors.initials: T
authors.lastname: Ingegnere
authors.affiliation: Laboratory of Immunology, Regina Elena National Cancer Institute, Rome, Italy.
authors.forename: Camilla
authors.initials: C
authors.lastname: Sampaoli
authors.affiliation: Laboratory of Immunology, Regina Elena National Cancer Institute, Rome, Italy.
authors.forename: Rocco
authors.initials: R
authors.lastname: Fraioli
authors.affiliation: Laboratory of Immunology, Regina Elena National Cancer Institute, Rome, Italy.
authors.forename: Patrizio
authors.initials: P
authors.lastname: Giacomini
mesh_terms: D000911:Antibodies, Monoclonal / Q000737:chemistry* / Q000276:immunology; D002460:Cell Line; D018604:Epitope Mapping; D000939:Epitopes / Q000737:chemistry* / Q000276:immunology; D015395:Histocompatibility Antigens Class I / Q000737:chemistry* / Q000276:immunology; D006801:Humans; D017510:Protein Folding; D001613:beta 2-Microglobulin / Q000737:chemistry* / Q000276:immunology; D000097625:HLA-E Antigens
chemical_list: D000911:Antibodies, Monoclonal; D000939:Epitopes; D015395:Histocompatibility Antigens Class I; D001613:beta 2-Microglobulin; D000097625:HLA-E Antigens
keywords: -B; -C; Conformation; HLA-A; HLA-E; mAbs; β2-microglobulin (β2m)