Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.48 (transcriptase)
 9,479 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Hepatitis E virus is responsible for both sporadic and epidemic hepatitis in developing countries. The nonenveloped virus is 27-34 nm in diameter and has been shown to contain a single-strand, positive-sense, polyadenylylated RNA genome of approximately 7.5 kilobases. The nucleotide sequence of the Burma strain of hepatitis E virus has been reported and three open reading frames (ORFs) have been identified. The deduced amino acid sequence from each of these ORFs was used to synthesize overlapping peptides (decamers overlapping at every fourth amino acid) on a solid phase. These peptides were then tested in an ELISA with pooled acute-phase sera from known cases of enterically transmitted non-A, non-B hepatitis collected in the Sudan. Linear B-cell epitopes were identified in all three ORFs. Epitopes were identified throughout the polyprotein encoded by ORF1, but they appeared to be particularly concentrated in the region of the RNA-dependent RNA polymerase. Distinct epitopes were identified in the presumed structural protein encoded by ORF2, and one epitope was identified close to the carboxyl terminus of the protein encoded by ORF3. These data precisely pinpoint linear B-cell epitopes recognized by antibodies from patients with acute hepatitis E and identify an antibody response directed against the RNA-dependent RNA polymerase.
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PMID:Human linear B-cell epitopes encoded by the hepatitis E virus include determinants in the RNA-dependent RNA polymerase. 137 90

The hepatitis B virus (HBV) is a coated, incompletely double-stranded DNA virus with some outstanding features. (1) All three coat proteins of HBV contain HBsAg, which is highly immunogenic inducing anti-HBs. These antibodies are protective for HBV outer cells (humoural immunity). Structural viral proteins induce specific T-lymphocytes, which are able to eliminate HBV-infected cells (cytotoxic T-cells; cellular immunity). (2) Intracellular HBV primarily causes little or no damage (non-cytopathogenic), which is an excellent strategy of viral survival. However, viral oligo-peptides of 8-15 amino acids are loaded on host cell MHC-class 1 molecules and are transported to the cell surface. Thus, HBV-specific T-lymphocytes are able to detect infected cells and destroy them, an ingenious defence strategy. However, this cell deletion triggered by inflammation cells may result in acute hepatitis. If HBV is not eliminated, a delicate balance between viral replication and immunodefence prevails which may lead to chronic hepatitis and liver cirrhosis. (3) In chronically infected cells HBV may become partly cytopathogenic--a process still poorly understood--and the viral DNA may integrate into the host cell DNA (through a viral transcriptase). If integration leads to activation of crucial host genes a hepatocellular carcinoma results. These outstanding features are responsible for the highly variable course of HBV infection and its final outcome, e.g. when the load of HBV-infected cells is still low at the time when an efficient immune defence starts, the infection is self-limited and asymptomatic, and immunity results. When there is no immune defence or a defective immune defence (immune tolerance of new-borns or immunosuppressed individuals) the HBV infection very often becomes chronic. In these cases, no acute hepatitis occurs, but hepatocellular carcinoma may result. Treatment with Interferon has become accepted, resulting in up to 30 to 40% of cases in the elimination of the virus. However, treatment is laborious and expensive, and the mechanism of action is still poorly understood (anti-viral and/or immune-modulating).
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PMID:Hepatitis B: virus, pathogenesis and treatment. 991 26

Samples of serum, tissue and faeces from two pig herds in England were examined for hepatitis E virus by reverse-transcriptase PCR (RT-PCR), and a virus strain from each herd was partially sequenced. Eleven of 42 faecal samples and 16 of 21 tissue samples from two pigs were positive for the virus by RT-PCR. Analysis of two unique but closely related nucleotide sequences obtained from the two herds showed that the viruses clustered in genotype III (6) with a human strain of the virus from an autochthonously acquired case of acute hepatitis in the UK. An ELISA based on recombinant open reading frame 2 (ORF-2) was used to detect antibodies to hepatitis E virus in 256 pig sera from the UK; 85.5 per cent of the samples were positive, compared with 58 per cent of similar samples from Swedish pigs and 23.5 per cent of samples from Dutch pigs.
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PMID:Evidence for the presence of hepatitis E virus in pigs in the United Kingdom. 1500 46

Hepatitis E is an acute hepatitis casused by hepatitis E virus (HEV) in developing countries, where it occurs as cases sporadic and in epidemics form. The causative agent, hepatitis E virus, is transmitted primarily by the fecal-oral route. HEV is icosahedron non-enveloped virus, and its genome is a single-stranded, positive-sense, 3'-polyadenylated RNA about 7.5 kb in length. It contains three open reading frames (ORFs). Of which ORF1 codes for a polyprotein of 1693 amino acids and contain domains homologous to a viral methyltransferase, a papainlike cysteine protease, an RNA helicasre, and an RNA-dependent RNA polymerase, besides the most hypervariable region of the HEV genome. And ORF3 codes for a 123-amino-acide-long polypeptide with unknown function. While the major viral capsid protein (pORF2, ORF2 codes) of 660 amino acid was showed to contain the protective epitope. The bacterially expressed polypeptide disignated as NE2 has been proved to be a protective antige. And the anti-NE2 monoclonal antibodies (mAb) was screend, two of these mAbs 8C11 and 8H3 were showed to be against separate conformational neutralization epitope of hepatitis E virus (HEV). And these two mAb were used to screen for binding peptides from a 7-peptides phage display library. After four rounds of panning, tweenty-one positive monoclonal phages (11 for 8C11, and 10 for 8H3) were selected and the inserted fragments were sequenced. The DNA sequence coding for the obtained dominant peptide 8C11 (N'-His-Pro-Thr-Leu-Leu-Arg-Ile-C', named 8C11A) and 8H3 (N'-Ser-Ile-Leu-Pro-Tyr-Pro-Tyr-C', named 8H3A) were then synthesized and cloned to insert between amino acid 78 to 83 of hepatitis B core antigen (HBcAg), then expressed in E. coli. The recombinant proteins aggregate into homodimer or polymer on SDS-PAGE, and could bind with mAb 8C11 and 8H3 in Western blotting. Respectively, the recombinant protein C8C11A showed to be dimer mainly, which can bind with mAb 8C11. The monomer and dimer of C8H3A are in the same amount on SDS-PAGE, but only the dimer could bind with mAb 8H3 on Western blotting. The renatured recombinant proteins were all showed to aggregate into virus like particles which were similar as HBcAg on transmission electron micrograph. The dominant peptide 8H3A (N'-Ser-Ile-Leu-Pro-Tyr-ProTyr-C') that selected out by mAb 8H3 was further chemo-synthesized, and its binding activity was confirmed by BIAcore biosensor. The result showed that this 7-peptide can bind with mAb 8H3 in a big Ka and Kd form, which means the binding is not stable. These results implicated that conformational dependent neutralization epitope could be partially modeled by short peptide, which provided a feasible route for subunit vaccine development.
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PMID:[Selection of a peptide mimic the neutralization epitope of hepatitis E virus with phage peptide display technology]. 1597 79

Hepatitis E virus (HEV) is a causative agent of acute hepatitis, and it is the sole member of the genus Hepevirus in the family Hepeviridae. The open reading frame 1 (ORF1) protein of HEV encodes nonstructural polyprotein with putative domains for methyltransferase, cysteine protease, helicase and RNA-dependent RNA polymerase. It is not yet known whether ORF1 functions as a single protein with multiple domains or is processed to form separate functional units. On the basis of amino acid conserved motifs, HEV helicase has been grouped into helicase superfamily 1 (SF-1). In order to examine the RNA helicase activity of the NTPase/helicase domain of HEV, the region (amino acids 960 to 1204) was cloned and expressed as histidine-tagged protein in Escherichia coli (HEV Hel) and purified. HEV Hel exhibited NTPase and RNA unwinding activities. Enzyme hydrolyzed all rNTPs efficiently, dATP and dCTP with moderate efficiency, while it showed less hydrolysis of dGTP and dTTP. Enzyme showed unwinding of only RNA duplexes with 5' overhangs showing 5'-to-3' polarity. We also expressed and purified two HEV Hel mutants. Helicase mutant I, with substitution in the nucleotide-binding motif I (GKS to GAS), showed 30% ATPase activity. Helicase mutant II, with substitutions in the Mg(2+) binding motif II (DEAP to AAAP), showed 50% ATPase activity. Both mutants completely lost ability to unwind RNA duplexes with 5' overhangs. These findings represent the first report demonstrating NTPase/RNA helicase activity of the helicase domain of HEV ORF1.
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PMID:NTPase and 5' to 3' RNA duplex-unwinding activities of the hepatitis E virus helicase domain. 2007 63

The presence of therapy-associated hepatitis B virus (HBV) variants is the main drawback of antiviral therapy for HBV infection. Moreover, drug-resistant variants are more insensitive to a second agent and more therapy-associated mutations will be present. To apply better nucleos(t)ide analogues (NA) and reduce the occurrence of resistance, the prevalence and types of drug-resistant mutations in acute hepatitis B patients were investigated in this study. One hundred three HBV DNA-positive patients with symptomatic acute hepatitis B that were observed from 2011 to 2013 were enrolled. Direct polymerase chain reaction sequencing was used firstly to screen HBV reverse-transcriptase domain to detect HBV mutants. Five lamivudine-resistant variants were identified. Clonal sequencing was performed for 5 resistance-positive samples and 10 other random samples. Interestingly, all detected samples harbored drug-resistant mutations, although with different percentage. Thirteen harbored lamivudine-related alone (five) or together with other NA related mutations (five with adefovir, one with entecavir, and one with telbivudine), and two of them harbored adefovir-related mutations. Also, mutations associated with four currently used NA were all detected, and the frequency is in accordance with the popularity of NA used in clinical practice. These data suggest that drug-resistant variants are present in patients with acute hepatitis B and NA should be applied more carefully for chronic hepatitis B patients developed from acute hepatitis B.
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PMID:Prevalence and types of drug-resistant variants in Chinese patients with acute hepatitis B. 2603

Hepatitis E Virus (HEV) is the major causative agent of acute hepatitis in developing countries. Its genome has three open reading frames (ORFs)-called as ORF1, ORF2, and ORF3. ORF1 encodes nonstructural polyprotein having multiple domains, namely: Methyltransferase, Y domain, Protease, Macro domain, Helicase, and RNA-dependent RNA polymerase. In the present study, we show that HEV-macro domain specifically interacts with light chain subunit of human ferritin (FTL). In cultured hepatoma cells, HEV-macro domain reduces secretion of ferritin without causing any change in the expression levels of FTL. This inhibitory effect was further enhanced upon Brefeldin-A treatment. The levels of transferrin Receptor 1 or ferroportin, two important proteins in iron metabolism, remained unchanged in HEV-macro domain expressing cells. Similarly, there were no alterations in the levels of cellular labile iron pool and reactive oxygen species, indicating that HEV-macro domain does not influence cellular iron homeostasis/metabolism. As ferritin is an acute-phase protein, secreted in higher level in infected persons and HEV-macro domain has the property of reducing synthesis of inflammatory cytokines, we propose that by directly binding to FTL, macro domain prevents ferritin from entering into circulation and helps in further attenuation of the host immune response.
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PMID:Hepatitis E virus ORF1 encoded macro domain protein interacts with light chain subunit of human ferritin and inhibits its secretion. 2717 Mar 77

Hepatitis E virus (HEV) is the most common cause of acute viral hepatitis in China. Recently, a shift in molecular epidemiology from hepatitis E genotype 1 (HEV-1) to hepatitis E genotype 4 (HEV-4) has been observed in Northern China, marking a switch from human-to-human transmission to zoonosis. However, similar data from cities in Southern China are lacking. This observational study of human hepatitis E cases in Shenzhen, a metropolitan city in the Pearl River Delta region, aimed to describe the clinical features and molecular epidemiology of hepatitis E in Southern China. Over a 55-month period, we identified 20 patients with acute hepatitis E. Most were middle-aged men, and 50% of patients had concomitant liver disease, of whom 70% were identified to have non-alcoholic fatty liver disease; such patients had a trend toward higher liver enzymes. Quantitative real-time RT-PCR using archived serum samples showed that 12 patients had hepatitis E viremia at presentation. Sequencing of the RNA-dependent RNA polymerase gene was performed for five of these patients, and phylogenetic analysis revealed that these five HEV isolates belonged to subgenotype 4b and were clustered with swine HEV isolates from Southern China. Combined with other studies showing similar findings, this suggests that the molecular epidemiology of hepatitis E in China is evolving toward low-level endemicity driven by foodborne transmission from seafood or pork products. The importance of concomitant liver disease, in particular non-alcoholic fatty liver disease, as a risk factor for severe hepatitis E requires further study.
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PMID:Clinical characteristics and molecular epidemiology of hepatitis E in Shenzhen, China: a shift toward foodborne transmission of hepatitis E virus infection. 2925 25

Hepatitis E virus (HEV) is one of the most common causes of acute hepatitis and jaundice in the world. Current understanding of the molecular virology and pathogenesis of hepatitis E is incomplete, due particularly to the limited availability of functional tools. Here, we report the development of tagged HEV genomes as a novel tool to investigate the viral life cycle. A selectable subgenomic HEV replicon was subjected to random 15-nucleotide sequence insertion using transposon-based technology. Viable insertions in the open reading frame 1 (ORF1) protein were selected in a hepatoblastoma cell line. Functional insertion sites were identified downstream of the methyltransferase domain, in the hypervariable region (HVR), and between the helicase and RNA-dependent RNA polymerase domains. HEV genomes harboring a hemagglutinin (HA) epitope tag or a small luciferase (NanoLuc) in the HVR were found to be fully functional and to allow the production of infectious virus. NanoLuc allowed quantitative monitoring of HEV infection and replication by luciferase assay. The use of HA-tagged replicons and full-length genomes allowed localization of putative sites of HEV RNA replication by the simultaneous detection of viral RNA by fluorescence in situ hybridization and of ORF1 protein by immunofluorescence. Candidate HEV replication complexes were found in cytoplasmic dot-like structures which partially overlapped ORF2 and ORF3 proteins as well as exosomal markers. Hence, tagged HEV genomes yield new insights into the viral life cycle and should allow further investigation of the structure and composition of the viral replication complex.IMPORTANCE Hepatitis E virus (HEV) infection is an important cause of acute hepatitis and may lead to chronic infection in immunocompromised patients. Knowledge of the viral life cycle is incomplete due to the limited availability of functional tools. In particular, low levels of expression of the ORF1 protein or limited sensitivity of currently available antibodies or both limit our understanding of the viral replicase. Here, we report the successful establishment of subgenomic HEV replicons and full-length genomes harboring an epitope tag or a functional reporter in the ORF1 protein. These novel tools should allow further characterization of the HEV replication complex and to improve our understanding of the viral life cycle.
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PMID:Recombinant Hepatitis E Viruses Harboring Tags in the ORF1 Protein. 3131 97