title: Recombinant hepatitis delta antigen from E. coli promotes hepatitis delta virus RNA replication only from the genomic strand but not the antigenomic strand.
abstract: Hepatitis delta antigen (HDAg) of hepatitis delta virus (HDV) typically consists of two related protein species. The small HDAg (S-HDAg) is a 24-kDa protein of 195 amino acids and the large HDAg (L-HDAg) is a 27-kDa protein with an additional 19 amino acids at its C-terminus. These two proteins have distinct functions in the HDV life cycle. We have developed conditions for expressing S-HDAg and L-HDAg in E. coli as soluble proteins to facilitate large-scale purification. These proteins were purified to homogeneity and shown to be biologically active. Transfection of the purified recombinant S-HDAg together with HDV genomic RNA resulted in viral RNA replication. Surprisingly, the purified S-HDAg could not initiate replication from the antigenomic-sense HDV RNA, even though the latter led to RNA replication when transfected with an mRNA encoding the S-HDAg. These results suggest that initiation of HDV RNA synthesis from the antigenomic RNA may require a form of HDAg that is modified in mammalian cells; in contrast, RNA synthesis from the genomic RNA could be initiated by the recombinant S-HDAg from E. coli. Interestingly, the purified L-HDAg appeared as multiple protein species, including one corresponding to S-HDAg, probably as a result of degradation. The partially proteolyzed L-HDAg also initiated HDV RNA replication under the same conditions. These results add to the mounting evidence that genomic- and antigenomic-strand HDV RNA syntheses are carried out by different mechanisms.
authors: Department of Molecular Microbiology and Immunology, Howard Hughes Medical Institute, Los Angeles, California 90033, USA.
authors: G T
authors: GT
authors: Sheu
authors: M M
authors: MM
authors: Lai
mesh_terms: D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D002846:Chromatography, Affinity; D002848:Chromatography, DEAE-Cellulose; D002850:Chromatography, Gel; D004926:Escherichia coli; D018963:Hepatitis Antigens / Q000302:isolation & purification / Q000378:metabolism*; D003698:Hepatitis Delta Virus / Q000235:genetics* / Q000502:physiology; D035923:Hepatitis delta Antigens; D016372:RNA, Antisense / Q000235:genetics; D012367:RNA, Viral / Q000235:genetics*; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D014158:Transcription, Genetic; D014779:Virus Replication
chemical_list: D018963:Hepatitis Antigens; D035923:Hepatitis delta Antigens; D016372:RNA, Antisense; D012367:RNA, Viral; D011994:Recombinant Proteins; C442244:hepatitis delta virus large antigen