Self-Association of a Synthetic Peptide from the N Terminus of the Hepatitis Delta Virus Protein Into an Immunoreactive α-Helical Multimer

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 92; no. 2; pp. 382 - 386
• Main Authors: Rozzelle, James E., Wang, Jia-Gang, Wagner, David S., Erickson, Bruce W., Lemon, Stanley M.
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences of the United States of America 17.01.1995; National Acad Sciences; National Academy of Sciences
• Subjects: Amino Acid Sequence; Animals; Antibodies; Antigens, Viral - immunology; Antigens, Viral - metabolism; Biochemistry; Centrifugation, Density Gradient; Chromatography; Circular Dichroism; Cross-Linking Reagents; Deuterium; Enzyme linked immunosorbent assay; Hepatitis antigens; hepatitis D virus; Hepatitis delta Antigens; Hepatitis delta virus; Hepatitis Delta Virus - immunology; Hydrogen; Immunoassay; Kinetics; Marmota - virology; Mass spectroscopy; Molecular Sequence Data; Molecules; Peptide Fragments - chemical synthesis; Peptide Fragments - immunology; Protein Conformation; Proteins; Sandwiches; Structure-Activity Relationship; Synthetic products
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.92.2.382

The formation of hepatitis delta antigen (HDAg) multimers is required for full biologic activity of this protein and for replication of the hepatitis delta virus. To determine the residues responsible for multimerization, three peptides [δ12-49, δ25-60(Y), δ12-60(Y)] from the putative coiled-coil multimer-forming domain of HDAg were chemically synthesized and biophysically characterized by circular dichroic spectroscopy, deuterium-exchange mass spectrometry, gel filtration, chemical crosslinking, and ultracentrifugation. By circular dichroism the 50-residue peptide δ12-60(Y) was half-denatured above 80⚬C and was 97% α-helical at 5°C and 84% α-helical at 37°C. By deuterium exchange, peptide δ12-60(Y) was 93% α-helical at 25°C. Its high α-helicity and melting temperature are due to the formation of an α-helical multimer consisting of four or more chains. All three synthetic peptides reacted with human anti-HDAg antibodies in an enzyme-linked immunosorbent assay, but only peptide δ12-60(Y) was detected in a sandwich radioimmunoassay in which successful antigens must display at least two antibody-binding sites, which correlates with the ability of this peptide to form multimers. Peptide δ12-60(Y) also interfered with the self-association of natural HDAg into multimers. These results have significant practical implications for development of improved diagnostic tests, antiviral agents, and possibly even vaccines for prevention of hepatitis delta virus disease.