Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0019163 (hepatitis B)
 38,309 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Hepadnaviruses, as well as other pararetroviruses, express their pol (P) gene product unfused to the preceding core gene implying that these retroelements have developed a mechanism for initiating assembly and replication that is principally different from the one used by retroviruses and retrotransposons. We have analysed this mechanism for the human hepatitis B virus by using a newly developed, highly sensitive detection method based upon radiolabelling of the P protein at newly introduced target sites for protein kinase A. The results obtained demonstrate that polymerase encapsidation depends on the concomittant encapsidation of the HBV RNA pregenome and that packaging of the viral RNA, in turn, depends on the presence of P protein. Loss of P protein encapsidation by mutations inactivating the HBV RNA encapsidation signal epsilon could be compensated by trans-complementation with recombinant RNA molecules carrying the epsilon sequence. Thus, in contrast to retroviral replication, the interaction of the hepadnaviral P protein and the RNA genome at its packaging signal appears to be crucial for initiating the formation of replication-competent nucleocapsids. Furthermore, RNA control of P protein packaging stringently limits the number of polymerase molecules that can be encapsidated.
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PMID:Hepadnaviral assembly is initiated by polymerase binding to the encapsidation signal in the viral RNA genome. 138 Apr 55

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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PMID: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. 160 35

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.
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PMID:Mutational analysis of the hepatitis B virus P gene product: domain structure and RNase H activity. 215 28

Antibodies against the RNase H domain of human hepatitis B virus P protein(s) are frequent markers of acute and chronic virus infection (T. Weimer, K. Weimer, Z.-X. Tu, M.-C. Jung, G. R. Pape, and H. Will, J. Immunol. 143:3750-3756, 1989). In the present study, these antibodies were determined in serial serum samples of experimentally infected chimpanzees and naturally infected human patients with acute and chronic hepatitis B virus infection. Anti-P antibodies were found in the sera of both chimpanzees and humans early in infection shortly after the immunoglobulin M anti-HBc response; they persisted in chronic carriers with ongoing viral replication but declined and disappeared at the time of virus clearance from the sera. These data demonstrate that antibodies to the RNase H domain of the hepatitis B virus P protein are early markers of infection and a signal of ongoing virus replication. Falling titers indicate the decline or end of active virus production and may therefore be a prognostic sign of virus elimination in natural infection and after antiviral therapy.
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PMID:Antibodies to the RNase H domain of hepatitis B virus P protein are associated with ongoing viral replication. 217 Jun 93

Encapsidation of the pregenomic RNA into nucleocapsids is a selective process which depends on specific RNA-protein interactions. The signal involved in the packaging of the hepatitis B virus (HBV) RNA pregenome was recently defined as a short sequence located near the 5' end of that molecule (Junker-Niepmann et al., EMBO J., in press), but it remained an open question which viral proteins are required. Using a genetic approach, we analyzed whether proteins derived from the HBV P gene play an important role in pregenome encapsidation. The results obtained with point mutations, deletions, and insertions scattered throughout the P gene clearly demonstrate that (i) a P gene product containing all functional domains is required both for the encapsidation of HBV pregenomic RNA and for packaging of nonviral RNAs fused to the HBV encapsidation signal, (ii) known enzymatic activities are not involved in the packaging reaction, suggesting that P protein is required as a structural component, and (iii) P protein acts primarily in cis, i.e., pregenomic RNAs from which P protein is synthesized are preferentially encapsidated.
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PMID:The P gene product of hepatitis B virus is required as a structural component for genomic RNA encapsidation. 221 19

Hepatitis B virus (HBV) is a small DNA virus that replicates by reverse transcription of a terminally redundant RNA, the pregenome. Specific packaging of this transcript into viral capsids is mediated by interaction of the reverse transcriptase, P protein, with the 5'-proximal encapsidation signal epsilon, epsilon-function is correlated with the formation of a hairpin structure containing a bulge and a loop, each consisting of 6 nt. To analyse the importance of primary sequence in these regions, we have combined selection of encapsidation competent individuals from pools of randomized epsilon-sequences in transfected cells with in vitro amplification, thus bypassing the current experimental limitations of the HBV system. While no alterations of the authentic loop sequence were detectable, many different sequences were tolerated in the 3'-part of the bulge. However, at the two 5'-proximal bulge positions the wt sequence was strongly selected for, indicating that for RNA packaging close contacts between protein and the 5'- but not the 3'-part of the bulge are important. Such a bipartite organisation provides a structural basis for the recently demonstrated special role of the 3'-part of the bulge as template for the first nucleotides of (-)-strand DNA in HBV reverse transcription.
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PMID:Distinct requirements for primary sequence in the 5'- and 3'-part of a bulge in the hepatitis B virus RNA encapsidation signal revealed by a combined in vivo selection/in vitro amplification system. 747 35

Hepadnavirus DNA minus strands are covalently linked at their 5' terminus to the viral P gene product, which has been taken to indicate that the hepadnaviral polymerase polypeptide itself also functions as a protein primer for initiating reverse transcription of the RNA pregenome. The present study confirms this indication by identifying the nucleotide-linked amino acid in the P protein sequence of the duck hepatitis B virus (DHBV). In a first set of experiments, mutational analysis of three phylogenetically conserved tyrosine residues in the DNA terminal (TP) domain indicated that of these, only tyrosine 96 was essential for both viral DNA synthesis in transfected cells and priming of DNA synthesis in a cell-free system. This assignment was confirmed by direct biochemical analysis: tryptic peptides from the DHBV P protein, 32P labelled at the priming amino acid by the initiating dGTP and additionally labelled internally by [35S]methionine, were isolated and analyzed in parallel to reference peptides synthesized chemically and 33P labelled by a tyrosine kinase. Mobility in high-performance liquid chromatography, as well as the release in stepwise amino acid sequencing of phospholabel and of [35S]methionine, identified the priming amino acid unequivocally as the tyrosine in the sequence 91KLSGLYQMK99, which is located in the center of the TP domain. Conserved sequence motifs surrounding Tyr-96 allow the prediction of the priming tyrosine in other hepadnaviruses. Weak sequence similarity to picornavirus genome-linked polypeptides (VPgs) and similar gene organization suggest a common origin for the mechanisms that use protein priming to initiate synthesis of viral DNA genomes or RNA genomes from an RNA template.
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PMID:Hepadnavirus P protein utilizes a tyrosine residue in the TP domain to prime reverse transcription. 751 55

Hepatitis B viruses encode a polymerase (P) protein with key roles in both reverse transcription and genomic RNA encapsidation. Genetic analysis of cis-acting signals required for viral replication implicates an RNA stem-loop structure in both RNA packaging and the initiation of reverse transcription, a process in which P protein also serves as the primer. We now show that duck hepatitis B virus (DHBV) polymerase binds specifically and with high affinity to this RNA stem-loop structure. Mutational analysis indicates that all mutations in the RNA target that inhibit the P protein-RNA interaction inhibit both in vivo RNA packaging and in vitro DNA priming to comparable extents. However, certain mutations in the loop region of the RNA have minimal impact on P protein-RNA binding but are nonetheless severely defective for packaging and DNA synthesis. Thus, P protein-RNA complex formation is necessary but not sufficient to initiate these activities. In addition, examination of RNA binding by truncated P proteins indicates that the C terminus of the polymerase, although required for RNA encapsidation in vivo, is dispensable for RNA binding and DNA priming.
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PMID:Site-specific RNA binding by a hepatitis B virus reverse transcriptase initiates two distinct reactions: RNA packaging and DNA synthesis. 752 92

The replication of hepatitis B virus DNA proceeds through reverse transcription of a pregenomic RNA intermediate, a reaction that takes place within viral nucleocapsids and is catalyzed by the viral P protein. P protein is involved in all phases of the reaction, serving as (a) a recognition factor for the selective encapsidation of the pregenomic RNA template; (b) the protein primer for the initiation of minus strand DNA synthesis; (c) the reverse transcriptase and DNA polymerase involved in strand elongation; and (d) the RNaseH activity required to remove RNA template prior to plus strand synthesis. P protein is capable of site-specific RNA recognition, specifically binding to a stem-loop structure at the 5' end of pregenomic RNA. This interaction is required for both RNA encapsidation and reverse transcription.
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PMID:Hepatitis B virus reverse transcriptase and its many roles in hepadnaviral genomic replication. 752 20

Replication of hepatitis B viruses proceeds by reverse transcription of an RNA intermediate, a reaction catalyzed by the virus-encoded polymerase (P protein). The reaction product is a partially duplex DNA whose (-)-strand is covalently linked to the P protein. Efforts to understand the mechanism of the reaction have been severely retarded by an inability to express functional polymerase outside of viral particles. Here we report the successful expression of enzymatically active polymerase in yeast cells, by fusing the P gene to coding sequences of the retrotransposon Ty1. The enzyme initiates correctly on viral RNA in yeast cells in vivo, producing nascent DNA chains covalently linked to protein, exactly as found in virus-infected cells. Replication complexes isolated from these yeast are enzymatically active in vitro, synthesizing DNA in a reaction that is actinomycin D-resistant but sensitive to RNase pretreatment. These results indicate that P protein is the sole viral protein required for the correct priming of reverse transcription and establish a tractable system for the biochemical dissection of the reaction.
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PMID:Expression of functional hepatitis B virus polymerase in yeast reveals it to be the sole viral protein required for correct initiation of reverse transcription. 768 22


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