Recombinant hepatitis delta antigen from E-coli promotes hepatitis delta virus RNA replication only from the genomic strand but not the antigenomic strand

Hepatitis delta antigen (HDAg) of hepatitis delta virus (HDV) typically con sists of two related protein species. The small HDAg (S-HDAg) is a 24-kDa p rotein of 195 amino acids and the large HDAg (L-HDAg) is a 27-kDa protein w ith 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. coil as soluble proteins to facilitate large-scale purification. These proteins were purified to homogeneity and s hown to be biologically active. Transfection of the purified recombinant S- HDAg together with HDV genomic RNA resulted in viral RNA replication. Surpr isingly, the purified S-HDAg could not initiate replication from the antige nomic-sense HDV RNA, even though the latter led to RNA replication when tra nsfected with an mRNA encoding the S-HDAg. These results suggest that initi ation of HDV RNA synthesis from the antigenomic RNA may require a form of H DAg that is modified in mammalian cells; in contrast, RNA synthesis from th e genomic RNA could be initiated by the recombinant S-HDAg from E. coli. In terestingly, the purified L-HDAg appeared as multiple protein species, incl uding 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- a nd antigenomic-strand HDV RNA syntheses are carried out by different mechan isms. (C) 2000 Academic Press.