A unique conformation at the carboxyl terminus of the small hepatitis delta antigen revealed by a specific monoclonal antibody.
Two forms of the hepatitis delta antigen (HDAg), a small (24 kDa) and a large (27 kDa) one, have different functions in the hepatitis delta virus (HDV) replication cycle. The small HDAg trans-activates RNA replication, while the large one inhibits RNA replication. The lack of the trans-acting activity in the large HDAg, even though it contains the complete sequence of small HDAg, suggests that the large HDAg lacks a certain functional conformation. To test such a possibility, monoclonal antibodies (MAbs) were generated from mice immunized with recombinant baculovirus-expressed small HDAg. As expected, most of the MAbs recognized both small and large HDAg. In addition, one MAb (9E4) was obtained which recognized only the small HDAg, but not the large one, in Western blot and immunoprecipitation analysis, suggesting that it recognized an epitope unique to small HDAg. However, MAb 9E4 detected both forms of HDAg in virus-infected cells by immunofluorescence and reacted with TrpE-large HDAg fusion proteins expressed in Escherichia coli, suggesting that this MAb recognizes a conformation-dependent epitope which is not present in the native large HDAg molecule but is detectable in LHDAg when its conformation is altered. The 9E4 epitope was mapped within a region of 32 amino acids at the carboxyl-terminus of small HDAg, indicating that this region contains a unique conformation not present in the native molecule of large HDAg. Since this is the only structure identified that is unique to small HDAg, the C-terminal region may contain the domain associated with the biological activities unique to the small HDAg.
Hwang
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SB
Howard Hughes Medical Institute, University of Southern California School of Medicine, Los Angeles 90033-1054.
Lai
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MM
D000818:Animals; D000911:Antibodies, Monoclonal; D000918:Antibody Specificity; D000956:Antigens, Viral / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D016367:Baculoviridae / Q000235:genetics; D002460:Cell Line; D005455:Fluorescent Antibody Technique; D005822:Genetic Vectors; D003698:Hepatitis Delta Virus / Q000235:genetics*; D035923:Hepatitis delta Antigens; D015151:Immunoblotting; D009036:Moths; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000276:immunology
D000911:Antibodies, Monoclonal; D000956:Antigens, Viral; D035923:Hepatitis delta Antigens; D011993:Recombinant Fusion Proteins; C442244:hepatitis delta virus large antigen