DNA-binding activity of hepatitis B e antigen polypeptide lacking the protaminelike sequence of nucleocapsid protein of human hepatitis B virus

Matsuda, Keizaburoh; Satoh, Shinji; Ohori, Hitoshi

doi:10.1128/jvi.62.9.3517-3521.1988

Cited by 21 publications

(12 citation statements)

References 37 publications

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“…The RNase H activity at 34 to 36 kDa was located just behind and in contact with a dark band identified as core protein by its characteristic size and prominence in core preparations (37,42,48). Its appearance as a dark band in this assay is consistent with its property of binding nucleic acid moieties (34,39). These results indicate that the major RNase H activity is efficiently renatured after the sample is boiled in SDS.…”

Section: Resultssupporting

confidence: 59%

Detection of an RNase H activity associated with hepadnaviruses

Oberhaus

Newbold

1995

J Virol

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Replication of the hepadnavirus DNA genome is accomplished via reverse transcription of an intermediate, pregenomic RNA molecule. This process is likely to be carried out by a virally encoded, multifunctional polymerase which possesses DNA-and RNA-dependent DNA polymerase and RNase H activities. However, the nature of the product(s) of the polymerase 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 to express active polymerase protein(s) heterologously. We have used activity gel assays to detect DNA-and RNA-dependent DNA polymerase activities associated with highly purified duck hepatitis B virus (DHBV) core particles (S. M. Oberhaus and J. E. Newbold, J. Virol. 67:6558-6566, 1993). Now we report that the same approach identifies a 35-kDa RNase H activity in association with highly purified DHBV core particles and crude preparations of virions from DHBV-infected ducks and woodchuck hepatitis virus-infected woodchucks. This is the first report of the detection of an hepadnavirus-associated RNase H activity. Its apparent size is smaller than any of the DNA polymerase activities that we detected previously and significantly smaller than the full-length protein predicted from the polymerase open reading frame (p85 for DHBV). These data suggest that the viral polymerase and RNase H activities are separable and that these enzymes may coordinate their activities in vivo by forming a complex.

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Section: Resultssupporting

confidence: 59%

Detection of an RNase H activity associated with hepadnaviruses

Oberhaus

Newbold

1995

J Virol

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“…I cannot rule out, however, the possibility that the faint signal originates from contaminating, i.e., nonencapsidated, genomic RNA. Obviously, in view of these results and previously reported, controversial in vitro data (8,18,27), the question of whether variant 144 retains a basal nucleic acid binding activity, i.e., a nucleic acid binding site independent of the poly-Arg region, needs further investigation.…”

Section: Discussionmentioning

confidence: 92%

The arginine-rich domain of the hepatitis B virus core protein is required for pregenome encapsidation and productive viral positive-strand DNA synthesis but not for virus assembly

Nassal

1992

J Virol

398

228

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Assembly of replication-competent hepatitis B virus (HBV) nucleocapsids requires the interaction of the core protein, the P protein, and the RNA pregenome. The core protein contains an arginine-rich C-terminal domain which is dispensable for particle formation in heterologous expression systems. Using transient expression in HuH7 cells of a series of C-terminally truncated core proteins, I examined the functional role of this basic region in the context of a complete HBV genome. All variants containing at least the 144 N-terminal amino acids were assembly competent, but efficient pregenome encapsidation was observed only with variants consisting of 164 or more amino acids. These data indicate that one function of the arginine-rich region is to provide the interactions between core protein and RNA pregenome. However, in cores from the variant ending with amino acid 164, the production of complete positive-strand DNA was drastically reduced. Moreover, almost all positive-strand DNA originated from in situ priming, whereas in wild-type particles, this type of priming not supporting the formation of relaxed circular DNA (RC-DNA) accounted for about one half of the positive strands. Further C-terminal residues to position 173 restored RC-DNA formation, and the corresponding variant did not differ from the full-length core protein in all assays used. The observation that RNA encapsidation and formation of RC-DNA can be genetically separated suggests that the core protein, via its basic C-terminal region, also acts as an essential auxiliary component in HBV replication, possibly like a histone, or like a single-stranded-DNA-binding protein. In contrast to their importance for HBV replication, sequences beyond amino acid 164 were not required for the formation of enveloped virions. Since particles from variant 164 did not contain mature DNA genomes, a genome maturation signal is apparently not required for HBV nucleocapsid envelopment.

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“…Antigenic determinants of the HBeAg are located between amino acids 76-89 [Salfeld et al, 19891 and 126-135 [Sallberg et al, 1991bl in the core region. Amino acids 124-133 may be exposed on the surface of native HBeAg [Matsuda et al, 1988;Sallberg et al, 19931, while amino acids 76-82 may be an epitope of the HBcAg [Sallberg et al, 1991al. Furthermore, mutations in the core promoter, which is thought to bind RNA polymerase for initiating transcription of preC mRNA, may also be related to the absence of serum HBeAg [Okamoto et al, 19941.…”

Section: Resultsmentioning

confidence: 99%

Prevalence of precore‐defective mutant of hepatitis B virus in HBV carriers

Niitsuma

Ishii

Saito

et al. 1995

Journal of Medical Virology

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Two hundred and seventy-three serum specimens from hepatitis B virus (HBV) carriers were examined for the presence of a characteristic one point mutation at nucleotide (nt) 1896 from the EcoRI site of the HBV genome in the precore region (the preC mutant) using restriction fragment length polymorphism (RFLP) analysis. This assay approach could detect preC mutants or wild-type sequences when either form constituted more than 10% of the total sample. Overall, 65.5% (76/116) of HBeAg-positive carriers had only the preC wild-type. All HBeAg-positive asymptomatic carriers (n = 14) had only the preC wild-type. In patients with chronic hepatitis B and in anti-HBe-positive asymptomatic carriers, increased prevalence of the preC mutant was associated with the development of anti-HBe antibodies and normalization of the serum alanine aminotransferase concentration. Furthermore, 27 (29.0%) of 93 HBeAg-negative carriers had unexpectedly preC wild-type sequences only. Direct sequencing of the HBV precore region of HBV specimens from 24 patients revealed no mutation at nt 1896, supporting the specificity of the RFLP analysis. These results suggest that RFLP analysis was accurate for the detection of the preC mutation and that the absence of serum HBeAg cannot be explained solely by the dominance of the preC mutant.

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DNA-binding activity of hepatitis B e antigen polypeptide lacking the protaminelike sequence of nucleocapsid protein of human hepatitis B virus

Cited by 21 publications

References 37 publications

Detection of an RNase H activity associated with hepadnaviruses

Detection of an RNase H activity associated with hepadnaviruses

The arginine-rich domain of the hepatitis B virus core protein is required for pregenome encapsidation and productive viral positive-strand DNA synthesis but not for virus assembly

Prevalence of precore‐defective mutant of hepatitis B virus in HBV carriers

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