title: Total chemical synthesis, characterization, and immunological properties of an MHC class I model using the TASP concept for protein de novo design.
abstract: The design, total chemical synthesis, and immunological properties of a four-alpha-helix bundle template-assembled synthetic protein (TASP) mimicking some of the structural features of the major histocompatibility complex (MHC) class I is described. In a first approach, the native sequence 58-74 of the alpha 1 heavy chain domain of HLA-A2 was modeled in order to increase helix stability and amphiphilicity of the 17-mer peptide, preserving the residues for potential T-cell receptor (TcR) binding properties. According to the TASP concept, these helical segments were covalently attached to a cyclic template molecule designed for the induction of a four-helix-bundle topology of the assembled peptide blocks. After extensive HPLC purification, stepwise solid-phase synthesis resulted in a TASP molecule of high chemical purity as demonstrated by analytical HPLC, mass spectrometry, and amino acid analysis. CD spectroscopic investigations are consistent with the onset of a partial alpha-helical conformation in aqueous buffer as well as in TFE. Antibodies raised directly against this four-alpha-helix bundle TASP molecule (without prior conjugation to a carrier molecule) were detected by ELISA. Flow cytometry studies showed that these antibodies recognize the native MHC class I molecule on the surface of HLA-A2-positive cells. The results indicate that the TASP approach represents a versatile tool for mimicking conformational epitopes.
authors.affiliation: Salk Institute, La Jolla, California 92037.
authors.forename: G
authors.initials: G
authors.lastname: Tuchscherer
authors.forename: C
authors.initials: C
authors.lastname: Servis
authors.forename: G
authors.initials: G
authors.lastname: Corradin
authors.forename: U
authors.initials: U
authors.lastname: Blum
authors.forename: J
authors.initials: J
authors.lastname: Rivier
authors.forename: M
authors.initials: M
authors.lastname: Mutter
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000276:immunology; D002460:Cell Line; D002942:Circular Dichroism; D005260:Female; D005434:Flow Cytometry; D015789:HLA-A2 Antigen / Q000276:immunology; D015395:Histocompatibility Antigens Class I / Q000737:chemistry* / Q000276:immunology; D013058:Mass Spectrometry; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D011487:Protein Conformation
chemical_list: D000906:Antibodies; D015789:HLA-A2 Antigen; D015395:Histocompatibility Antigens Class I