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)

The biochemical properties of four acyclovir-resistant mutants are described. Two of these mutants, PAAr5 and BWr, specified nucleotidyl transferase (DNA polymerase) activities which were less sensitive to inhibition by acyclovir triphosphate than their wild-type counterparts. Another mutant, IUdRr, exhibited reduced ability to phosphorylate acyclovir. The fourth mutant, ACGr4, both induced an altered DNA polymerase and failed to phosphorylate appreciable amounts of acyclovir. BWr, a new acyclovir-resistant mutant derived from the Patton strain of herpes simplex virus type 1, induced a DNA polymerase resistant to inhibition by acyclovir triphosphate, but, unlike the polymerases induced by PAAr5 and ACGr4, still sensitive to phosphonoacetic acid. Resistance of BWr to acyclovir mapped close to the PAAr locus and was separable from mutations in the herpes simplex virus thymidine kinase gene by recombination analysis.
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PMID:Acyclovir-resistant mutants of herpes simplex virus type 1 express altered DNA polymerase or reduced acyclovir phosphorylating activities. 627 27

A group of 43 phosphonoacetic acid (PAA)-resistant mutants of herpes simplex virus type 1 was isolated after the mutagenesis of infected cells with nitrosoguanidine. One of these mutants, designated PAA1rts1, was found to be temperature sensitive (ts), that is, unable to replicate at 39.5 degrees C, the nonpermissive temperature. Recombination analysis of PAA1rts1 indicated that the PAA1r mutation and the ts1 mutation are loosely linked and are located on two separate genes. PAA1rts1 showed a defect in viral DNA synthesis at 39.5 degrees C, which presumably can be attributed to the production of a PAA-resistant and thermolabile DNA polymerase. PAA1rts1 was also defective in the shutoff of host DNA synthesis at the restrictive temperature.
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PMID:Isolation and preliminary characterization of a phosphonoacetic acid-resistant and temperature-sensitive mutant of herpes simplex virus type 1. 628 39

Mutants of herpes simplex virus type 1 resistant to the antiviral drug 9-beta-D-arabinofuranosyladenine (araA) have been isolated and characterized. AraA-resistant mutants can be isolated readily and appear at an appreciable frequency in low-passage stocks of wild-type virus. Of 13 newly isolated mutants, at least 11 were also resistant to phosphonoacetic acid (PAA). Of four previously described PAA-resistant mutants, two exhibited substantial araA resistance. The araA resistance phenotype of one of these mutants, PAAr5, has been mapped to the HpaI-B fragment of herpes simplex virus DNA by marker transfer, and araA resistance behaved in marker transfer experiments as if it were closely linked to PAA resistance, a recognized marker for the viral DNA polymerase locus. PAAr5 induced viral DNA polymerase activity which was much less susceptible to inhibition by the triphosphate derivative of araA than was wild-type DNA polymerase. These genetic and biochemical data indicate that the herpes simplex virus DNA polymerase gene is a locus which, when mutated, can confer resistance to araA and thus that the herpes simplex virus DNA polymerase is a target for this antiviral drug.
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PMID:Mutations in the herpes simplex virus DNA polymerase gene can confer resistance to 9-beta-D-arabinofuranosyladenine. 628 81

A virus-specified thymidine kinase appears to be a general requirement for herpes virus susceptibility to the antiviral effect of acyclovir. Surprisingly, mouse cytomegalovirus (MCMV), which does not encode for a thymidine kinase, is exquisitely sensitive to the drug both in vitro and in vivo. The drug is active against the virus in the absence of a cellular thymidine kinase and the antiviral activity is not diminished in the presence of excess thymidine or a variety of nucleosides and deoxynucleosides. Thus, a thymidine phosphorylation pathway is not required for the drug's activation of this infection. The enzyme system responsible for phosphorylation of the drug has not been identified. Mouse cytomegalovirus mutants resistant to the drug have been isolated, indicating that the antiMCMV effect results from selective inhibition of viral replication rather than indirectly through toxicity to the host cell. Eight resistant mutants appear to be in the same complementation group and seven of the mutants demonstrate coresistance to phosphonoacetic acid, a marker for the DNA polymerase locus of herpes viruses. The evidence to date indicates that the MCMV DNA polymerases is the final site of action of the drug. Investigations of the antiMCMV activity of acyclovir should provide insights into the antiviral effects of this drug and other nucleoside analogs in other herpes virus infections in which the virus does not code for a thymidine kinase (for example, human cytomegalovirus and Epstein-Barr virus).
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PMID:Acyclovir in mouse cytomegalovirus infections. 628 1

Five independently derived variants of a herpes simplex virus type I (HSV-1) strain were plaque purified from a virus population passaged in 1 mM phosphonoformic acid (PFA). The DNA polymerase induced by the parent and PFA-resistant viruses were purified and characterized. No differences were observed among the enzymes with respect to their chromatographic properties, specific activities, or polypeptides resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The variant enzymes exhibited levels of PFA resistance which ranged from 15- to 25-fold. Resistance to PFA was always associated with a similar degree of resistance to its congener phosphonoacetic acid, but cross-resistance to beta-phenylphosphonoacetic acid was only seen with two of the five variant enzymes. PFA and pyrophosphate were mutually competitive in PPi exchange reactions, but in DNA synthetic reactions the levels of resistance to PFA and PPi were not equal. The apparent affinities of the enzymes for Mg2+ did parallel their affinities for PFA. Km values of dNTPs were about 2-fold higher than the parent virus enzyme for all of the variant enzymes except one which was 4-fold higher. The processivity of polymerization was apparently unaffected by the enzyme changes related to PFA resistance although one variant enzyme had a lower value. Resistance among the variant enzymes to the triphosphates of 9-(2-hydroxyethoxymethyl)guanine and 2',3'-dideoxyguanosine was directly related to the level of resistance to PFA. The data presented here indicated that (i) PFA resistance may result from several types of active site alterations, since the PFA-resistant enzymes were of three kinetically distinct types. Also, additional enzyme alterations, probably unrelated to PFA resistance, were detected in one enzyme. (ii) PFA and PPi possess some different binding determinants within the active center of herpes simplex virus type I DNA polymerase. (iii) PFA and the triphosphates of 9-(2-hydroxyethoxymethyl)guanine and 2',3'-dideoxyguanosine may have a common ultimate inhibitory mechanism.
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PMID:Characterization of the DNA polymerases induced by a group of herpes simplex virus type I variants selected for growth in the presence of phosphonoformic acid. 628 45

The physical map limits of DNA sequences within the DNA polymerase locus of herpes simplex virus (HSV) type 1 that define resistance mutations to adenine arabinoside (ara-A) and E-5-(2-bromovinyl)-2'-deoxyuridine (BVDU) were determined. Intertypic recombinants between HSV types 1 and 2, generated by marker rescue of temperature-sensitive mutations, had genome structures determined by restriction endonuclease analysis and were used to show that the resistance mutation for ara-Ar-1 is closely linked to the resistance mutations for phosphonoacetic acid (paar-1) and acycloguanosine (acvr-1) within a region of 2.6 kilobase pairs (kbp) in the HSV DNA polymerase locus. The resistance mutation for bvdur-1 is defined by a 2.9-kbp region that overlaps with the region defining resistance to the other three drugs but that is transferred separately. The DNA polymerase locus also contains a 2.2-kbp region that maps adjacent to and to the left of the region defining the bvdur-1 mutation which can be transferred separately and defines a region determining the HSV-1-specific sensitivity to BVDU in a manner analogous to that to acyclovir.
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PMID:Resistance of herpes simplex virus to adenine arabinoside and E-5-(2-bromovinyl)-2'-deoxyuridine: a physical analysis. 628 86

Acyclovir (ACV) has been shown to inhibit the replication of herpes simplex virus (HSV) in vitro. We examined a wide variety of HSV clinical isolates for the presence of naturally occurring ACV-resistant (ACVr) variants. Although the ACV doses that inhibited 50% of these isolates were within the range of doses inhibiting 50% of the ACV-susceptible wild-type strains, we successfully isolated variants resistant to high ACV concentrations (25 to 75 microM) from each virion population even in the absence of prior drug exposure. Furthermore, we demonstrated, by fluctuation analysis of two encephalitis strains, that the ACVr variants were clonally distributed in the virus populations before exposure to ACV and did not result from rapid adaptation to ACV. All variants isolated after a single exposure to a high dose of ACV were true ACVr variants, as demonstrated by their plating efficiencies in the presence of ACV. We found that 36 and 50% of the ACVr variants of the two strains examined in detail displayed plating efficiencies in phosphonoacetic acid of greater than 0.1, possibly indicating that many of the ACVr variants contained alterations in the DNA polymerase gene locus. Because the distribution of ACVr variants in natural populations is relatively high (10(-4), these results suggest that selection of ACVr strains during ACV therapy is possible.
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PMID:Herpes simplex virus variants restraint to high concentrations of acyclovir exist in clinical isolates. 628 42

Fourteen mutants known or likely to contain mutations in the herpes simplex virus DNA polymerase gene were examined for their sensitivity to aphidicolin in plaque reduction assays. Eleven of these exhibited some degree of hypersensitivity to the drug; altered aphidicolin-sensitivity correlated with altered sensitivity to the pyrophosphate analog, phosphonoacetic acid. The DNA polymerase specified by one of these mutants, PAAr5, required roughly seven-fold less aphidicolin to inhibit its activity by 50% than did polymerase specified by its parental strain. Mutations responsible for the aphidicolin-hypersensitivity phenotype of PAAr5 were mapped to an 0.8 kbp region in the herpes simplex virus DNA polymerase locus. These data taken together indicate that 1) mutations in the herpes simplex virus DNA polymerase gene can confer altered sensitivity to aphidicolin, 2) that the HSV polymerase is sensitive to aphidicolin in vivo, and 3) that amino acid alterations which affect aphidicolin binding may affect the pyrophosphate exchange-release site as well, suggesting that aphidicolin binds in close proximity to this site.
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PMID:Mutations in the herpes simplex virus DNA polymerase gene conferring hypersensitivity to aphidicolin. 630 78

A DNA-relaxing enzyme was found to copurify along with herpes simplex virus type I (HSV-1)-induced DNA polymerase throughout a multistep purification scheme. Both the enzymes had similar sedimentation velocity, required high ionic strength for optimal enzymatic activities and showed time dependence of reaction. The DNA-relaxing enzyme however, differed from the HSV-1 DNA polymerase in its requirement for higher Mg2+ concentration, rATP and much broader pH dependence. Furthermore, phosphonoacetic acid, a potent inhibitor of HSV-1 DNA polymerase did not influence the DNA-relaxing activity even at a much higher concentration. On the other hand, the DNA-relaxing enzyme associated with the DNA polymerase may be specified by HSV-1 since IgG fraction of rabbit antisera against the virus-infected cells but not against the mock-infected cells strongly inhibited both the enzymatic activities. Thus, HSV-1-induced DNA polymerase which is known to be associated with a 3' to 5' exonuclease may also be associated with yet another enzymatic activity involved in DNA metabolism.
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PMID:A DNA topoisomerase activity copurifies with the DNA polymerase induced by herpes simplex virus. 630 34

The stimulation of host cell DNA synthesis was studied in permissive human embryonic lung (HEL) cells and in nonpermissive rabbit kidney (RK) cells infected with human cytomegalovirus (HCMV). Host cell DNA synthesis was induced by HCMV infection in resting cells of both types. In permissive cultures the stimulation of cellular DNA synthesis was detectable mainly in those cells which had not become productively infected and in which virus antigens were not detectable. In abortively infected RK cells, on the other hand, stimulation of host cell DNA synthesis and the expression of virus antigens were detected in the same cells. Infection of actively growing permissive HEL cells resulted in a shutdown of cellular DNA synthesis beginning approximately 10 hr postinfection. Shutdown of cellular DNA synthesis also occurred when the infected cells were treated with phosphonoacetic acid and was thus classified as an "early" virus function. In actively growing, abortively infected RK cells, on the other hand, host cell DNA synthesis was not affected, indicating that the early virus function(s) responsible for inhibition of cellular DNA synthesis was not expressed in these cells. Virus-encoded DNA polymerase activity, another early virus gene function, was also not detected in these abortively infected cultures. In RK cells the cellular DNA synthesized as a result of infection was capable of undergoing at least one further round of replication, indicating that the HCMV gene expression which occurred in abortively infected RK cells was not lethal for these cells.
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PMID:Correlation between stimulation of host cell DNA synthesis by human cytomegalovirus and lack of expression of a subset of early virus genes. 631 75


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