EXPERIMENTAL CONFIRMATION OF A HEPATITIS-B VIRUS (HBV) EPSILON-LIKE BULGE-AND-LOOP STRUCTURE IN AVIAN HBV RNA ENCAPSIDATION SIGNALS

Hepatitis B viruses replicate via reverse transcription of an RNA inte rmediate. This RNA pregenome serves as mRNA and is packaged into capsi ds and reverse transcribed. Both processes require the interaction of the viral reverse transcriptase, P protein, with the 5'-proximal epsil on-signals on the pregenome, For epsilon of human hepatitis B virus (H BV), the presence of a functionally important stem-loop structure with a central bulge, part of which acts as template for a short primer of first-strand DNA synthesis, has been experimentally confirmed. Based on phylogeny and its functional similarities to epsilon, the D epsilon -signal of duck hepatitis B virus (DHBV) had been proposed to have a s imilar structure which does not, however, correspond to the most stabl e computer prediction. We have therefore experimentally determined the secondary structures of D epsilon and of the H epsilon-signal of hero n hepatitis B virus which differs considerably from D epsilon in prima ry sequence yet interacts productively with DHBV P protein. Our data s upper? an HBV epsilon-like structure for both D epsilon and H epsilon; in particular the bulge is highly conserved, in accord with its speci al function in replication. However, the apical loop in H epsilon is m uch enlarged suggesting that by an induced-fit mechanism, both RNAs ma y adopt a new, probably similar conformation in the complex with P pro tein. (C) 1997 Academic Press