DETECTION OF AN RNASE-H ACTIVITY ASSOCIATED WITH HEPADNAVIRUSES

Replication of the hepadnavirus DNA genome is accomplished via reverse transcription of an intermediate, pregenomic RNA molecule. This proce ss is likely to be carried out by a virally encoded, multifunctional p olymerase which possesses DNA- and RNA-dependent DNA polymerase and RN ase H activities. However, the nature of the product(s) of the polymer ase gene predicted to mediate these functions is unclear. Biochemical studies of the polymerase protein(s) have been limited by its apparent low abundance in virus particles and, until recently, the inability t o express active polymerase protein(s) heterologously. We have used ac tivity gel assays to detect DNA- and RNA-dependent DNA polymerase acti vities associated,vith highly purified duck hepatitis B virus (DHBV) c ore particles (S. M. Oberhaus and J. E. Newbold, J. Virol. 67:6558-656 6, 1993). Now we report that the same approach identifies a 35-kDa RNa se H activity in association with highly purified DHBV core particles and crude preparations of virions from DHBV-infected ducks and woodchu ck hepatitis virus-infected woodchucks. This is the first report of th e detection of an hepadnavirus-associated RNase H activity. Its appare nt size is smaller than any of the DNA polymerase activities that we d etected previously and significantly smaller than the full-length prot ein predicted from the polymerase open reading frame (p85 for DHBV). T hese data suggest that the viral polymerase and RNase H activities are separable and that these enzymes may coordinate their activities in v ivo by forming a complex.