Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.7.6 (RNA polymerase)
34,946 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Phosphonoacetate is a highly specific inhibitor of herpes simplex virus-induced DNA polymerase. Sensitivity of herpesvirus type 1 or type 2 induced DNA polymerase to the drug was similar. However, DNA polymerases from other sources such as the host cells (Wi-38), Micrococcus luteus, and hepatitis B virus were highly resistant. In addition, Escherichia coli RNA polymerase and reverse transcriptase of Rous sarcoma virus were also insensitive to the drug. Enzyme kinetic studies showed that inhibition was noncompetitive with respect to deoxyribonucleotide triphosphates. The Ki value was about 0.45 muM. The apparent Km values for dTTP, dATP, dCTP, and dGTP were 0.71, 0.75, 0.42, and 0.39 muM, respectively. The base composition of template has no profound effect on the extent of inhibition. The drug caused uncompetititve inhibition with respect to template which indicated that phosphonoacetate did not bind directly to template DNA. Results are presented which suggest that phosphonoacetate did not affect the formation of the enzyme-DNA complex but probably inhibited the elongation step of DNA polymerase reaction.
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PMID:Mode of inhibition of herpes simplex virus DNA polymerase by phosphonoacetate. 5 71

The hybridization properties of the herpes simplex virus type 1 (HSV) genome have been analysed. The DNA has a kinetic complexity of 1 X 10(-8). E. cole RNA polymerase was found to initiate synthesis at about 70 sites on the HSV DNA. The in vitro RNA product from this reaction was complementary to about 80% of the HSV genome. The RNA-DNA hybridization rate constant (Kh) was determined using conditions of both RNA excess and DNA excess. Using this rate constant one can analyse the content of HSV sequences in any RNA population.
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PMID:Annealing and hybridization properties of herpes simplex virus type 1 DNA. 18 38

Analysis of nuclei isolated from herpes simplex virus (HSV)-infected cells by electrophoresis in polyacrylamide gels showed that only virion-associated DNA molecules migrated into the gels. The viral DNA molecules, which do not migrate in the gels, are the precursors for the mature viral DNA. Centrifugation of deoxycholate-treated infected nuclei in sucrose gradients revealed that most of the viral DNA co-sedimented with the cellular DNA. The DNA-dependent RNA polymerase activity also co-sedimented with the viral and cellular DNA. Incubation of nuclei isolated from HSV-infected cells, under in vitro conditions that support DNA synthesis, resulted in the synthesis of viral molecules of low molecular weight. Under the same conditions, nuclei from Burkitt lymphoblasts did not synthesize DNA. The nature of the association between cellular DNA and EBV DNA was studied by hybridization with EBV cRNA. EBV-specific sequences were found in the lymphoblast DNA; they banded at a density of 1.707 g/cm3. Some of the viral mRNA transcribed in HSV-infected nuclei is symmetrically transcribed from HSV DNA. The symmetrical portions of the RNA are removed, since poly (A)-containing mRNA molecules lack homologous sequences. Most of the RNA synthesized in HSV-infected nuclei can be released after incubation of the nuclei in vitro in the presence of ATP. The released RNA consists of poly(A)plus and poly (A)-minus molecules. The mechanism of RNA transport from the nuclei still remains to be studied.
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PMID:Nucleic acid biosynthesis in nuclei of cell infected with herpesviruses (HSV and EBV). 19 Jan 26

In herpes simplex virus type 1 (HSV-1)-infected HEp-2 cells, amanitin added before or at various times after infection always reduced viral multiplication. Also, the three waves of transcription of HSV-1 DNA, which led to the synthesis of alpha, beta-, and gamma-polypeptides, were all sensitive to amanitin in HEp-2 cells, and the amanitin-sensitive RNA polymerase activities of isolated nuclei were equally sensitive to the inhibitor before and during the infection. On the contrary, HSV-1 DNA transcription was totally unaffected by amanitin in AR1/9-5B cells, a mutant subline of CHO cells that possesses an amanitin-resistant RNA polymerase B. Together, these results strongly suggest that HSV-1 DNA utilizes for its transcription a polymerase undistinguishable from host cell RNA polymerase B with respect to its sensitivity to amanitin.
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PMID:Evidence that herpes simplex virus DNA is transcribed by cellular RNA polymerase B. 19 58

We have determined the orientation of 4 immediate early (IE) mRNA's on the herpes simplex virus type 1 genome by mapping cDNA's complementary to the 3'-termini of messages. These IE mRNA's are transcribed by a pre-existing cell RNA polymerase, and we propose a model which allows their synthesis from a circular template using a single virus promoter region. The promoter region, which is located in the two repetitive DNA regions which flank the short unique region of the virus genome, may serve to initiate bidirectional transcription of these IE mRNA's.
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PMID:Orientation of herpes simplex virus type 1 immediate early mRNA's. 22 44

Infection with herpes simplex virus (HSV) results in an increase in the transcription of the endogenous Alu repeated sequence by RNA polymerase III. This effect is also observed in uninfected cells stably transformed with a plasmid expressing the HSV immediate-early protein ICP27 or in cells transfected with the gene encoding this protein. Both uninfected cells expressing ICP27 and cells infected with virus producing functional ICP27 display increased activity of the cellular transcription factor TFIIIC when compared with untreated cells. This increase is not observed, however, in cells infected with a mutant strain of virus which does not produce ICP27. Hence ICP27 induces elevated Alu transcription by activating transcription factor TFIIIC, which is the limiting factor for such transcription. This is the first report of increased activity of a cellular transcription factor during HSV infection, when most cellular gene activity is inhibited.
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PMID:The herpes simplex virus immediate-early protein ICP27 stimulates the transcription of cellular Alu repeated sequences by increasing the activity of transcription factor TFIIIC. 132 Mar 73

We examine the RNA polymerase II-dependent transcription directed by several promoters in extracts prepared from distinct developmental stages of Xenopus laevis. RNA polymerase II accurately initiates transcription from the cytomegalovirus, herpes simplex virus thymidine kinase, and Xenopus heat-shock protein (hsp) 70 promoters. The efficiency of transcription of these different promoters is dependent on whether extracts from oocytes, eggs, or somatic cells are used and on the temperature of incubation. In contrast to the viral promoters, the hsp 70 promoter is more active at heat shock temperatures in oocyte and egg extracts (31 degrees-34 degrees C) than at physiological temperatures for Xenopus (20 degrees-25 degrees C). These in vitro transcription extracts should be useful in examining the molecular mechanisms responsible for differential gene expression during Xenopus development.
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PMID:Characterization of RNA polymerase II-dependent transcription in Xenopus extracts. 151 44

The ICP18.5 gene (UL28) of herpes simplex virus type 1 is a member of a well-conserved gene family among herpesviruses and is thought to play a role in localization of viral glycoproteins. We have cloned, sequenced, and expressed the entire pseudorabies virus (PRV) ICP18.5 open reading frame in Escherichia coli as a Cro-ICP18.5 fusion protein. Rabbit antiserum against Cro-ICP18.5 immunoprecipitated a 79-kDa protein from PRV-infected cells as well as a 79-kDa protein from in vitro translation of a T7 RNA polymerase transcript of the ICP18.5 gene. ICP18.5 could be detected in infected cells by 2 h postinfection. Analysis by indirect immunofluorescence demonstrated that ICP18.5 became associated with the nucleus. Subcellular fractionation confirmed that ICP18.5 synthesized during a pulse-chase experiment appeared in the nuclear fraction with time and was stable for at least 2.5 h after synthesis. Pulse-chase analysis revealed that ICP18.5 was synthesized as a monomer during a 2-min pulse labeling but formed faster sedimenting complexes which were sensitive to sodium dodecyl sulfate (SDS) treatment. The majority of ICP18.5 appeared in complexes with an antigenically unrelated 70-kDa protein. Immunoblot analysis of total infected-cell extracts using polyvalent anti-ICP18.5 serum demonstrated that a 74-kDa cellular protein in addition to the 79-kDa ICP18.5 was detected. This cellular protein was present at similar levels in uninfected cells and in PRV-infected cells at least 12 h into the infectious cycle.
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PMID:Overexpression in bacterial and identification in infected cells of the pseudorabies virus protein homologous to herpes simplex virus type 1 ICP18.5. 164 90

The herpes simplex virus type 1 (HSV-1) gene encoding the ribonucleotide reductase (RR) small subunit (R2) was cloned as an unfused and intact open reading frame into a T7 RNA polymerase expression system in Escherichia coli. The expressed product was recovered from bacteria in soluble form and constituted 7% of the soluble protein. Protein purification yielded 3.5 mg of 95% pure R2 per litre of bacterial culture. The correct composition of the purified protein was verified by amino acid analysis and N-terminal sequencing. The isoelectric point of the protein was 5.3. Atomic emission spectroscopy indicated that the iron content of the E. coli-expressed R2 was 0.2 to 0.5 atoms of iron per R2 protomer as compared with a theoretical maximum value of 2. The E. coli-expressed HSV-1 R2 existed as a combination of a stable dimer and monomer. Combination of the E. coli-expressed R2 with the E. coli-expressed large subunit (R1) gave an active holoenzyme. Thus, the T7 expression system provides a rich source of enzymically active HSV-1 RR.
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PMID:Purification and characterization of the herpes simplex virus type 1 ribonucleotide reductase small subunit following expression in Escherichia coli. 164 78

Activator proteins that control transcription initiation by RNA polymerase II usually have two domains: one binds to DNA, and the other activates transcription. A particularly potent acidic activation domain at the C terminus of the herpes simplex virus protein VP16 binds directly and selectively to the human and yeast TATA box-binding factor TFIID. We have now investigated the biological significance of this in vitro interaction by using mutant forms of VP16. For changes at the critical phenylalanine residue at position 442 of VP16 there was a good correlation between transactivation activity in vivo and the binding of VP16 to TFIID in vitro. In contrast, mutants with reduced negative charge were more defective for binding than for activation.
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PMID:Reduced binding of TFIID to transcriptionally compromised mutants of VP16. 164 2


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