Mutational analysis of the hepatitis B virus P gene product: domain structure and RNase H activity.
To correlate the hepatitis B virus P gene with the enzymatic activities predicted to participate in hepadnavirus reverse transcription, a series of P gene mutants containing missense mutations, in-phase insertions, and in-phase deletions was constructed by site-directed mutagenesis. These mutants were tested in the context of otherwise intact hepatitis B virus genomes for the ability to produce core particles containing the virus-associated polymerase activity. The results obtained suggest that the P protein consists of three functional domains and a nonessential spacer arranged in the following order: terminal protein, spacer, reverse transcriptase/DNA polymerase, and RNase H. The first two domains are separated by a spacer region which could be deleted to a large extent without significant loss of endogenous polymerase activity. In cotransfection experiments, all P gene mutants could be complemented in trans by constructs expressing the wild-type gene product but not by a second P gene mutant. This indicates that the multifunctional P gene is expressed as a single translational unit and independent of the core gene and furthermore that the gene product is freely diffusible and not processed before core assembly.
Radziwill
G
G
Zentrum für Molekulare Biologie, University of Heidelberg, Federal Republic of Germany.
Tucker
W
W
Schaller
H
H
D000595:Amino Acid Sequence; D001483:Base Sequence; D002460:Cell Line; D002872:Chromosome Deletion; D004722:Endoribonucleases / Q000235:genetics* / Q000378:metabolism; D005814:Genes, Viral; D005816:Genetic Complementation Test; D006515:Hepatitis B virus / Q000235:genetics*; D006801:Humans; D007438:Introns; D008969:Molecular Sequence Data; D009154:Mutation; D015345:Oligonucleotide Probes; D010957:Plasmids; D016914:Ribonuclease H; D015678:Viral Structural Proteins / Q000235:genetics*
D015345:Oligonucleotide Probes; D015678:Viral Structural Proteins; D004722:Endoribonucleases; D016914:Ribonuclease H