Total chemical synthesis, characterization, and immunological properties of an MHC class I model using the TASP concept for protein de novo design.
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.
Tuchscherer
G
G
Salk Institute, La Jolla, California 92037.
Servis
C
C
Corradin
G
G
Blum
U
U
Rivier
J
J
Mutter
M
M
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
D000906:Antibodies; D015789:HLA-A2 Antigen; D015395:Histocompatibility Antigens Class I