Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.49 (reverse transcriptase)
 31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Primary duck hepatocyte (PDH) cultures, congenitally infected with the duck hepatitis B virus (DHBV), were grown on feeder cell layers of irradiated human embryonic lung fibroblasts and then exposed to a number of compounds with recognized or potential antiviral activity. These compounds included conventional antiviral agents, reverse transcriptase inhibitors, compounds with activity to supercoiled-DNA, and DNA-binding agents. Twenty-three compounds were evaluated, and 13 were found to inhibit significantly viral DNA replication. Seven of these compounds (ellipticine, amsacrine, coumermycin A1, Adriamycin, mitozantrone, chloroquine, and neocarzinostatin) acted at the level of viral SC DNA and significantly inhibited production of duck hepatitis B surface antigen (DHBsAg). Conventional agents that inhibited DHBV DNA replication included ganciclovir, acyclovir, bromovinyldeoxyuridine, ribavirin, phosphonoformate, and dideoxyadenosine. Except for dideoxyadenosine, these inhibitors of viral DNA synthesis did not significantly inhibit DHBsAg production. Two additional compounds, novobiocin and nalidixic acid, altered the pattern of viral DNA replication, especially the generation and processing of viral SC DNA, and also inhibited the production of DHBsAg. Several compounds acting at the level of viral SC DNA have now been identified and may offer potential for the management of chronic hepatitis B virus infection.
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PMID:Antiviral strategies in chronic hepatitis B virus infection: II. Inhibition of duck hepatitis B virus in vitro using conventional antiviral agents and supercoiled-DNA active compounds. 169 59

The polymerase (P) gene of hepadnaviruses encodes a large polypeptide that appears to participate in several steps in the viral life cycle: packaging of viral RNA, providing the primer for synthesis of minus-strand DNA, synthesizing minus-strand DNA from an RNA template and plus-strand DNA from a DNA template, and degrading viral RNA in RNA-DNA hybrids. To assist in the assignment of these functions to domains of the duck hepatitis B virus polymerase protein, we have constructed a series of substitution mutations and a large insertion mutation, based in part on amino acid sequence comparisons with other proteins known to exhibit reverse transcriptase (RT) and RNase H activities. We found that changes in highly conserved sequences in putative RT and RNase H domains in the carboxy-terminal half of the protein dramatically reduced synthesis of both strands of viral DNA without major effects on RNA packaging into subviral cores. Thus we can uncouple RNA packaging and DNA synthesis but cannot separate RT and RNase H activities as has been done with human hepatitis B virus. The viability of a mutant with a large insertion (123 amino acids) upstream of the RT and RNase H domain indicates that a hinge region may separate parts of the polymerase protein implicated in priming and polymerization.
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PMID:Effects of insertional and point mutations on the functions of the duck hepatitis B virus polymerase. 169 97

Constructs expressing the core, surface, X, or polymerase proteins of hepatitis B virus were transfected into human cells. In transient assays, only the polymerase inhibited the responses to interferons alpha and gamma (IFN-alpha and -gamma). Stable expression of the polymerase was achieved in the cell line 2fTGH, which carries an IFN-inducible marker gene, by growth under conditions that select for inhibition of the response to IFN-alpha, but the clones grew poorly. When expressed alone, the terminal protein domain of the polymerase gene inhibited the response to IFN-alpha and the reverse transcriptase plus RNase H domains appeared to be toxic. Clones of cells expressing terminal protein alone, selected for the loss of response to IFN-alpha, grew normally and had no detectable response to IFN-alpha, IFN-gamma, or double-stranded RNA. Binding of IFN-alpha to these cells was not impaired but did not lead to activation of the E alpha subunit of the IFN-induced transcription factor E. These observations are of potential importance in relation to the pathogenesis of chronic hepatitis B virus infection and the resistance of such infection to IFN-alpha therapy.
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PMID:Expression of the terminal protein region of hepatitis B virus inhibits cellular responses to interferons alpha and gamma and double-stranded RNA. 170 74

Duck hepatitis B virus mutants containing frameshift or stop codon mutations in a portion of the viral pol gene separating the terminal protein and reverse transcriptase domains had a leaky phenotype and, depending on the location and type of mutation, synthesized up to 10% as much viral DNA as did the wild type. This region of the pol gene had previously been reported to be refractory to missense mutations; in fact, the leakiness of most of our mutants appeared attributable to translational suppression, which would also be expected to introduce amino acid changes. However, at least one mutant (pH1093 + 2), which was ca. 10% as active as the wild type, appeared to use a novel pathway to express the viral pol gene. Our analyses indicated that pH1093 + 2 synthesized the viral reverse transcriptase as a fusion protein with the amino-terminal portion of the pre-S envelope protein. Thus, in this case, the products of the terminal-protein and reverse transcriptase domains of the pol gene would function as separate protein species, though perhaps noncovalently joined in a dimeric structure during assembly of DNA replication complexes. Evidence was also obtained that was consistent with the idea that the wild-type pol gene may, at least in certain instances, be expressed as functional, subgenic polypeptides.
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PMID:Evidence that less-than-full-length pol gene products are functional in hepadnavirus DNA synthesis. 170 80

Polymerase chain reaction (PCR) with a reverse transcriptase step characterized a specific transcription activity in hepatitis B virus (HBV)-infected peripheral blood mononuclear cells (PBMC) in two patients (1 and 2) with chronic hepatitis positive for antibody to hepatitis B core antigen (HBc). Patient 1 was also coinfected with human hepatitis delta virus. A patient who cleared HBV replication after antiviral treatment with vidarabine served as negative control. HBV-specific RNA poly A sequences were detected by PCR in PBMC of patients 1 and 2 even without detectable HBV DNA (patient 2) as shown by dot blot and PCR assays. RNA sequences were found in both the nucleus and cytoplasm. The demonstration of HBV mRNA sequences within PBMC suggests the transcription of viral DNA, in agreement with the findings of HBV surface antigen in PBMC. The results in patient 1 demonstrated HBV mRNA sequences in leukocytes even without PCR-detectable HBV DNA sequences, likely due to ongoing hepatitis delta virus replication.
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PMID:Detection of polyadenylated RNA in hepatitis B virus-infected peripheral blood mononuclear cells by polymerase chain reaction. 170 1

The hepatitis B virus, although containing a DNA genome, replicates by reverse transcription of an RNA pregenome. The viral Pol gene encodes the reverse transcriptase which catalyzes viral DNA synthesis. To study the interaction of this protein with HBV RNA, the entire Pol gene product was expressed except its eight amino-terminal codons in Escherichia coli as fusion protein with beta-galactosidase. In the absence of competing nucleic acids full-length expression products were able to nonspecifically bind in vitro synthesized HBV RNAs of different polarity and length. However, if competed with an excess of unspecific RNA, only those HBV RNAs were bound which contained besides the direct repeats 1 and 2 nucleotide sequences downstream of direct repeat 1. The corresponding binding site was found to be located within the adjacent 134 nucleotides downstream of DR1. We conclude from our data that this region which is in part homologous to the U5 region of retroviral genomes may be important for the binding of the HBV Pol gene product to the viral pregenome.
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PMID:Identification of a binding site in the hepatitis B virus RNA pregenome for the viral Pol gene product. 170 31

We have analyzed 11 independent mutations located at various domains of the polymerase gene (pol) of human hepatitis B virus. Surprisingly, one of the two missense mutants within the spacer/intron region appears to be lethal. This result further defines the N-terminal limit of the reverse transcriptase domain. Alternatively, it suggests the potential existence of a novel domain with an unknown function. Two missense mutations within the terminal protein (TP) domain appear to be replication-defective as well, suggesting a functionally essential role of the TP domain in DNA replication.
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PMID:Replication-defective missense mutations within the terminal protein and spacer/intron regions of the polymerase gene of human hepatitis B virus. 185 74

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

Hepatitis B virus (HBV) is the causative agent of hepatocellular carcinoma (HCC) in man. The HBV genome is a circular partially double-stranded DNA molecule of about 3.2 kb. The HBV genome contains four structural genes coding for the HBV envelope (HBsAg) and core (HBcAg/HBeAg) proteins, endogenous DNA-polymerase with the additional enzymatic activity of a reverse transcriptase and polypeptide X functioning as a trans-activator of cellular and viral genes. HBV DNA integration in the genomes of HCCs and hepatocytes of HBV carriers is an important evidence establishing a relationship between the HBV infection and the development of HCC. The mechanism of HBV DNA integration into the cellular genome and the possible role of integrated HBV DNA sequences in the malignant transformation of hepatocytes are discussed.
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PMID:[Human hepatitis B virus and hepatocellular carcinoma]. 215 10

Hepatitis B virus DNA replicates via its own polymerase that also acts as reverse transcriptase (Summers and Mason, 1982). This enzyme is encoded by a 3.5 Kb mRNA transcript covering the whole genome. Since the same transcript also codes for the core protein, and since the core open reading frame (ORF) is located upstream of the pol ORF, it has been suggested that the polymerase is first produced as a core-pol fusion protein that subsequently undergoes cleavage. This is already known to be the case with retrovirus reverse transcriptase, for which a gag-pol fusion protein is made first and the latter protein is liberated by proteolytic cleavage. We investigated this problem using mutants that were modified at the translation initiation codon for the core and precore ORF. Our findings suggested that polymerase translation occurred from the internal AUG codon independently of core protein synthesis, and that obligatory production of the core-pol fusion protein is accordingly unlikely.
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PMID:Translation of hepatitis B virus DNA polymerase from the internal AUG codon, not from the upstream AUG codon for the core protein. 222 32


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