Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.7 (DNA polymerase)
 17,007 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Recent studies have shown that the kinetics of hepatitis B virus (HBV) clearance during antiviral therapy are slow, requiring long-term therapy to control viral replication. It has also been shown that emergence of resistant mutants is accelerated by high HBV replication and hepatocyte turnover, which are common features in patients with chronic HBV infection. It is therefore important to continue research on novel antiviral agents to design optimal combination strategies. The duck and woodchuck models of hepadnavirus infection are currently the best models for the investigation of the inhibitory effect of nucleoside analogues on wild-type and lamivudine-resistant mutants. Our studies revealed that these mutants have a decreased priming and elongation activity, and remain sensitive to novel nucleoside analogues. Tissue culture experiments with transient transfection of wild-type and mutant viruses also confirmed these data. Infection studies in primary hepatocytes and in animal models gave insight into the pathobiology of lamivudine-resistant mutants, as well as into the kinetics of wild-type virus clearance during antiviral therapy. Furthermore, it appears that novel strategies inducing a specific anti-HBV immune response by a DNA vaccine approach may induce viral clearance. Altogether, these results suggest that: (i) lamivudine-resistant mutants are likely to be cross-resistant to other L-cytidine analogues; (ii) antiviral therapy using a single reverse transcriptase (RT) inhibitor is likely to fail to eradicate viral covalently closed circular DNA; and (iii) new nucleoside analogues with unique mode of action (inhibition of priming or elongation of RT, or DNA polymerase activity) and activity against lamivudine-resistant strains are emerging. Combination of these new anti-HBV agents with DNA based immunization may prove useful to eradicate viral infection.
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PMID:Evaluation of novel strategies to combat hepatitis B virus targetting wild-type and drug-resistant mutants in experimental models. 1159 80

Infection of shrimp cells with white spot syndrome virus (WSSV) results in an increase in ribonucleotide reductase (RR) expression at the RNA level. In this article we further express and characterize the induction of a novel ribonucleotide reductase after WSSV infection of shrimp cells. A baculovirus/insect system was used to express the two recombinant protein subunits RR1 and RR2, and a DNA polymerase coupled RR activity assay showed a marked increase in ribonucleotide reductase activity when cell extracts containing recombinant RR1 and RR2 were combined. The same assay revealed that RR activity increased as infection advanced in the gills of experimentally infected shrimp. An increase in RR expression was also detected at the protein level in WSSV-infected shrimp cells. An immunocytochemistry assay by confocal laser scanning microscopy showed that in hemocytes collected from WSSV-infected shrimp, both of the subunit proteins (RR1 and RR2) were concentrated mainly around the nucleus, but only RR1 was detected inside it. All of these results suggest that WSSV RR is functionally involved during WSSV infection.
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PMID:Ribonucleotide reductase of shrimp white spot syndrome virus (WSSV): expression and enzymatic activity in a baculovirus/insect cell system and WSSV-infected shrimp. 1250 69

The DNA polymerase (DNApol) and major capsid protein (MCP) genes were used as models to study promoter activity in Chilo iridescent virus (CIV). Infection of Bombyx mori SPC-BM-36 cells in the presence of inhibitors of DNA or protein synthesis showed that DNApol, as well as helicase, is an immediate-early gene and confirmed that the major capsid protein (MCP) is a late gene. Transcription of DNApol initiated 35 nt upstream and that of MCP 14 nt upstream of the translational start site. In a luciferase reporter gene assay both promoters were active only when cells were infected with CIV. For DNApol sequences between position -27 and -6, relative to the transcriptional start site, were essential for promoter activity. Furthermore, mutation of a G within the sequence TTGTTTT located just upstream of the DNApol transcription initiation site reduced the promoter activity by 25%. Sequences crucial for MCP promoter activity are located between positions -53 and -29.
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PMID:Promoter analysis of the Chilo iridescent virus DNA polymerase and major capsid protein genes. 1469 70

We designed, optimized, and extensively tested several sensitive and specific real-time PCR assays for rapid detection of both smallpox and pan-orthopox virus DNAs. The assays are based on TaqMan 3'-minor groove binder chemistry and were performed on both the rapid-cycling Roche LightCycler and the Cepheid Smart Cycler platforms. The hemagglutinin (HA) J7R, B9R, and B10R genes were used as targets for the variola virus-specific assays, and the HA and DNA polymerase-E9L genes were used as targets for the pan-orthopox virus assays. The five orthopox virus assays were tested against a panel of orthopox virus DNAs (both genomic and cloned) at the U.S. Army Medical Research Institute of Infectious Diseases (USAMRIID). The results indicated that each assay was capable of detecting both the appropriate cloned gene and genomic DNA. The assays showed no cross-reactivity to the 78 DNAs in the USAMRIID bacterial cross-reactivity panel. The limit of detection (LOD) of each assay was determined to be between 12 and 25 copies of target DNA. The assays were also run against a blind panel of DNAs at the Centers for Disease Control and Prevention (CDC) on both the LightCycler and the Smart Cycler. The panel consisted of eight different variola virus isolates, five non-variola virus orthopox virus isolates, two varicella-zoster virus isolates, and one herpes simplex virus isolate. Each sample was tested in triplicate at 2.5 ng, 25 pg, 250 fg, and 2.5 fg, which represent 1.24 x 10(7), 1.24 x 10(5), 1.24 x 10(3), and 1.24 x 10(1) genome equivalents, respectively. The results indicated that each of the five assays was 100% specific (no false positives) when tested against both the USAMRIID panels and the CDC blind panel. With the CDC blind panel, the LightCycler was capable of detecting 96.2% of the orthopox virus DNAs and 93.8% of the variola virus DNAs. The Smart Cycler was capable of detecting 92.3% of the orthopox virus DNAs and between 75 and 93.8% of the variola virus DNAs. However, all five assays had nearly 100% sensitivity on both machines with samples above the LOD (>12 gene copies). These real-time PCR assays represent a battery of tests to screen for and confirm the presence of variola virus DNA. The early detection of a smallpox outbreak is crucial whether the incident is an act of bioterrorism or an accidental occurrence.
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PMID:Smallpox and pan-orthopox virus detection by real-time 3'-minor groove binder TaqMan assays on the roche LightCycler and the Cepheid smart Cycler platforms. 1476 23

The multicapsid nucleopolyhedroviruses (NPVs) of Spodoptera exigua (SeMNPV), Spodoptera frugiperda (SfMNPV), and Spodoptera littoralis (SpliNPV) are genetically similar (78 % similarity) but differ in their degree of host specificity. Infection by each of the three NPVs in these three Spodoptera host species was determined by oral inoculation of larvae with occlusion bodies (OBs) or intrahaemocoelic injection with occlusion derived virions (ODVs). RT-PCR analysis of total RNA from inoculated insects, targeted at immediate early (ie-0), early (egt, DNA polymerase), late (chitinase) and very late genes (polyhedrin), indicated that each of the NPVs initiated an infection in all three host species tested. SpliMNPV produced a fatal NPV disease in both heterologous hosts, S. frugiperda and S. exigua, by oral inoculation or injection. SfMNPV was lethal to heterologous hosts, S. exigua and S. littoralis, but infected larvae did not melt and disintegrate, and progeny OBs were not observed. SeMNPV was able to replicate in heterologous hosts and all genes required for replication were present in the genome, as the virus primary infection cycle was observed. However, gene expression was significantly lower in heterologous hosts. SeMNPV pathogenesis in S. frugiperda and S. littoralis was blocked at the haemocoel transmission stage and very nearly cleared. SeMNPV mixtures with SpliMNPV or SfMNPV did not extend the host range of SeMNPV; in all cases, only the homologous virus was observed to proliferate. It is concluded that entry and the primary virus infection cycle are not the only, or the major determinants, for SeMNPV infection of heterologous Spodoptera species.
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PMID:Virus entry or the primary infection cycle are not the principal determinants of host specificity of Spodoptera spp. nucleopolyhedroviruses. 1544 46

Human herpesvirus 6 (HHV-6), which is present in more than 90% of the human, is known to cause infectious diseases in immuno-compromised patients, e.g., transplant patients. To clarify the possible role of the pattern of expression of HHV-6 genes in various types of HHV-6B infection, we sought to determine whether or not viral DNA microarray could be used for detailed characterization of viral transcription using a HHV-6B DNA microarray that contains 97 known open reading frames of HHV-6B. A subset of genes are preferentially expressed in persistent infection: U16 (IE-B, transactivator, US22 gene family), U18 (IE-B, homolog to HCMV IE glycoprotein), U20 (glycoprotein), U27 (DNA polymerase processivity transactivator), U82 (gL, gH accessory protein), U83 (chemokine), U85 (OX-2 homology, glycoprotein), U90 (IE-A), and U94 (transactivator), respectively. Although the function of each HHV-6B is not fully understood, our study suggests that comprehensive analysis of HHV-6B transcription is useful not only to clarify the pathogenesis of the virus but also to develop new strategies for anti-viral drugs.
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PMID:Transcriptional profiling of human herpesvirus type B (HHV-6B) in an adult T cell leukemia cell line as in vitro model for persistent infection. 1572 Dec 66

Infection of Wi-38 cells with herpes simplex virus induced an elevated DNA polymerase activity which had many biochemical properties different from normal cell DNA polymerase. Phosphonoacetic acid specifically inhibited the virus-induced DNA polymerase as compared to the normal WI-38 cell DNA polymerase. The compound did not appear to inhibit enzyme activity by interacting with the DNA primer.
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PMID:Inhibition of DNA polymerase from herpes simplex virus-infected wi-38 cells by phosphonoacetic Acid. 1678 61

Infection with hepatitis B virus (HBV) is extremely widespread - it infects two billion people out of the six billion world population. It is estimated that between 350 and 400 million people are chronically infected with HBV. Chronic HBV infection leads to development of complications, such as cirrhosis and hepatocellular carcinoma (HCC), which arise in 15-40% of patients. HBV-related liver disease and its complications result in approximately one million deaths each year. The ultimate goals of chronic hepatitis B (CHB) therapy are decreases in the incidence of cirrhosis, end-stage liver disease and HCC. The following six medications are currently approved by the U.S. Food and Drug Administration for the treatment of CHB: interferon (INF)-alpha2b, pegylated INF-alpha2a, lamivudine, adefovir dipivoxil, entecavir and, recently, telbivudine. Interferon therapy has many contraindications and commonly causes multiple intolerable adverse effects. Lamivudine therapy leads to increased development of resistant mutations with each year of use. Entecavir, a new guanosine nucleoside analogue with specific activity against HBV DNA polymerase, represents a third agent within the nucleoside/nucleotide HBV polymerase inhibitor class. It has distinct advantages over lamivudine and adefovir dipivoxil: it has a three-step mechanism of action, is the most potent inhibitor of HBV DNA polymerase, is not associated with any major adverse effects and has a limited potential for resistance. In clinical trials, entecavir was superior to lamivudine in all primary endpoints in both nucleoside-naive and lamivudine-refractory hepatitis B e antigen (HBeAg)-positive and HBeAg-negative patients. Preliminary data support entecavir efficacy in patients with cirrhosis and HIV/HBV coinfected patients. No resistance occurred after two years of entecavir therapy in nucleoside-naive patients. Up to 9% resistance developed in patients with documented prior lamivudine resistance during 96 weeks of entecavir therapy. Currently, entecavir should be considered a first- or second-line treatment option for the management of HBeAg-positive or -negative nucleoside-naive or lamivudine-refractory CHB patients.
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PMID:Entecavir: a new nucleoside analogue for the treatment of chronic hepatitis B. 1746 Jul 84

Infection with equid herpesvirus type 1 (EHV-1) leads to respiratory disease, abortion, and neurologic disorders in horses. Molecular epidemiology studies have demonstrated that a single nucleotide polymorphism resulting in an amino acid variation of the EHV-1 DNA polymerase (N752/D752) is significantly associated with the neuropathogenic potential of naturally occurring strains. To test the hypothesis that this single amino acid exchange by itself influences neuropathogenicity, we generated recombinant viruses with differing polymerase sequences. Here we show that the N752 mutant virus caused no neurologic signs in the natural host, while the D752 virus was able to cause inflammation of the central nervous system and ataxia. Neurologic disease induced by the D752 virus was concomitant with significantly increased levels of viremia (p = 0.01), but the magnitude of virus shedding from the nasal mucosa was similar between the N752 and D752 viruses. Both viruses replicated with similar kinetics in fibroblasts and epithelial cells, but exhibited differences in leukocyte tropism. Last, we observed a significant increase (p < 0.001) in sensitivity of the N752 mutant to aphidicolin, a drug targeting the viral polymerase. Our results demonstrate that a single amino acid variation in a herpesvirus enzyme can influence neuropathogenic potential without having a major effect on virus shedding from infected animals, which is important for horizontal spread in a population. This observation is very interesting from an evolutionary standpoint and is consistent with data indicating that the N752 DNA pol genotype is predominant in the EHV-1 population, suggesting that decreased viral pathogenicity in the natural host might not be at the expense of less efficient inter-individual transmission.
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PMID:A point mutation in a herpesvirus polymerase determines neuropathogenicity. 1799

Nucleic acid amplification technologies are used in the field of molecular biology and recombinant DNA technologies. These techniques are used as leading methods in detecting and analyzing a small quantity of nucleic acids. The polymerase chain reaction (PCR) is the most widely used method for DNA amplification for detection and identification of infectious diseases, genetic disorders and other research purposes. However, it requires a thermocycling machine to separate two DNA strands and then amplify the required fragment. Novel developments in molecular biology of DNA synthesis in vivo demonstrate the possibility of amplifying DNA in isothermal conditions without the need of a thermocycling apparatus. DNA polymerase replicates DNA with the aid of various accessory proteins. Recent identification of these proteins has enabled development of new in vitro isothermal DNA amplification methods, mimicking these in vivo mechanisms. There are several types of isothermal nucleic acid amplification methods such as transcription mediated amplification, nucleic acid sequence-based amplification, signal mediated amplification of RNA technology, strand displacement amplification, rolling circle amplification, loop-mediated isothermal amplification of DNA, isothermal multiple displacement amplification, helicase-dependent amplification, single primer isothermal amplification, and circular helicase-dependent amplification. In this article, we review these isothermal nucleic acid amplification technologies and their applications in molecular biological studies.
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PMID:Nucleic acid isothermal amplification technologies: a review. 1826 8


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