Self-association of a synthetic peptide from the N terminus of the hepatitis delta virus protein into an immunoreactive alpha-helical multimer.
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 [ delta 12-49, delta 25-60(Y), delta 12-60(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 delta 12-60(Y) was half-denatured above 80 degrees C and was 97% alpha-helical at 5 degrees C and 84% alpha-helical at 37 degrees C. By deuterium exchange, peptide delta 12-60(Y) was 93% alpha-helical at 25 degrees C. Its high alpha-helicity and melting temperature are due to the formation of an alpha-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 delta 12-60(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 delta 12-60(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.
Department of Chemistry, University of North Carolina at Chapel Hill 27599.
J E
JE
Rozzelle
J G
JG
Wang
D S
DS
Wagner
B W
BW
Erickson
S M
SM
Lemon
D000595:Amino Acid Sequence; D000818:Animals; D000956:Antigens, Viral / Q000276:immunology* / Q000378:metabolism; D002499:Centrifugation, Density Gradient; D002845:Chromatography; D002942:Circular Dichroism; D003432:Cross-Linking Reagents; D003903:Deuterium; D003698:Hepatitis Delta Virus / Q000276:immunology*; D035923:Hepatitis delta Antigens; D007118:Immunoassay; D007700:Kinetics; D008392:Marmota / Q000821:virology; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000276:immunology*; D011487:Protein Conformation; D013329:Structure-Activity Relationship
D000956:Antigens, Viral; D003432:Cross-Linking Reagents; D035923:Hepatitis delta Antigens; D010446:Peptide Fragments; C442244:hepatitis delta virus large antigen; D003903:Deuterium