共查询到20条相似文献,搜索用时 0 毫秒
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A single-stranded DNA-dependent ATPase from monkey kidney tissue culture cells (CV-1) has been found associated with SV40 chromatin. This ATPase activity is distinguishable from the ATPase activity of T-antigen by the following properties: the Km for ATP, elution from phosphocellulose, and stimulation of the ATPase activity by single-stranded DNA but not by double-stranded DNA. The ATPase has been isolated and characterized from the nuclei of uninfected cells. ATP hydrolysis is dependent on single-stranded DNA and a divalent cation. The km values for ATP and single-stranded DNA are 0.024 mM and 0.09 microgram/ml, respectively. The affinity of the ATPase for single-stranded DNA is sufficiently high that the enzyme co-sediments with single-stranded DNA in glycerol gradients. The binding of single-stranded DNA is independent of ATP and MgCl2; however, ATP hydrolysis increases the exchange of enzyme between different DNA molecules. Form I (superhelical) SV40 DNA is also a substrate for ATPase binding, but relaxed Form I, Form II (nicked circular), and double-stranded linear SV40 DNAs are not substrates. Because the DNA helix within chromatin is not under the same kind of tortional strain as Form I DNA, we hypothesize that the ATPase is bound to the single-stranded regions of replication forks in the SV40 chromatin. 相似文献
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S Wickner 《The Journal of biological chemistry》1984,259(22):14038-14043
The product of bacteriophage P22 gene 12 is known from genetic experiments to be essential for phage DNA replication. The P22 12 protein has been purified to near homogeneity from Escherichia coli lysogenic for lambda-P22 hybrid phage containing the replication genes of P22. The protein has a subunit molecular weight of 46,000. The purified protein contains ATPase activity that is stimulated by single-stranded DNA. The ATPase is poorly stimulated by double-stranded DNA. All four ribonucleoside triphosphates are hydrolyzed; none of the deoxynucleoside triphosphates are hydrolyzed. In addition, the P22 12 protein binds to single-stranded DNA in the presence of ATP. Studies of oligonucleotide synthesis by P22 12 protein in conjunction with E. coli dnaG primase are presented in the succeeding paper (Wickner, S. (1984) J. Biol. Chem. 259, 14044-14047). 相似文献
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Identification of the vaccinia virus gene encoding nucleoside triphosphate phosphohydrolase I, a DNA-dependent ATPase. 总被引:4,自引:2,他引:2
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Vaccinia virus encapsidates a DNA-dependent ATPase known as nucleoside triphosphate phosphohydrolase I (NPH I). A bacteriophage lambda gt11 expression library of poxvirus DNA was screened with antibodies specific for NPH I. Positive clones were used to probe restriction fragments of vaccinia virus genomic DNA to locate the NPH I gene. The identity of the open reading frame (ORF) was confirmed by placing it downstream of a bacteriophage T7 promoter, transcribing the ORF in vitro, and translating the RNA in a reticulocyte lysate. A polypeptide of the correct molecular weight, which was recognized by anti-NPH I antibody, was synthesized. Inspection of the deduced amino acid sequence of the NPH I ORF revealed consensus ATP-binding sites. 相似文献
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Intrinsic DNA-dependent ATPase activity of reverse gyrase 总被引:4,自引:0,他引:4
Reverse gyrase is a type I DNA topoisomerase that promotes positive supercoiling of closed-circular double-stranded DNA through an ATP-dependent reaction, and it was purified from an archaebacterium, Sulfolobus. When ATP is replaced by UTP, GTP, or CTP, this enzyme just relaxes the negatively supercoiled closed-circular double-stranded DNA. We found that reverse gyrase hydrolyzes ATP through a double-stranded DNA-dependent reaction. The superhelicity of the DNA did not affect the ATPase activity. However, reverse gyrase does not hydrolyze UTP, GTP, or CTP. Therefore, any of the four nucleotide 5'-triphosphates acts as an effector for the topoisomerase activity of reverse gyrase, but only ATP supports the positive supercoiling of closed-circular double-stranded DNA, through the energy released on its hydrolysis. Single-stranded DNA was a much more potent cofactor for the ATPase activity of the enzyme than double-stranded DNA, and it acted as a potent inhibitor for the topoisomerase activity on double-stranded DNA. These results indicate that reverse gyrase has higher affinity to single-stranded DNA than to double-stranded DNA, which suggests a cellular function of the enzyme. 相似文献
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N Moussatché 《Biochimica et biophysica acta》1985,826(2-3):113-120
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Conserved sequences near the early transcription start sites of vaccinia virus. 总被引:2,自引:2,他引:0
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A Vassef 《Nucleic acids research》1987,15(4):1427-1443
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DNA-dependent RNA polymerase subunits encoded within the vaccinia virus genome. 总被引:12,自引:6,他引:6
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Antiserum to a multisubunit DNA-dependent RNA polymerase from vaccinia virions was prepared to carry out genetic studies. This antiserum selectively inhibited the activity of the viral polymerase but had no effect on calf thymus RNA polymerase II. The specificity of the antiserum was further demonstrated by immunoprecipitation of RNA polymerase subunits from dissociated virus particles. The presence in vaccinia virus-infected cells of mRNA that encodes the polymerase subunits was determined by in vitro translation. Immunoprecipitable polypeptides with Mrs of about 135,000, 128,000, 36,000, 34,000, 31,000, 23,000, 21,000, 20,000, and 17,000 were made when early mRNA was added to reticulocyte extracts. The subunits were encoded within the vaccinia virus genome, as demonstrated by translation of early mRNA that hybridized to vaccinia virus DNA. The locations of the subunit genes were determined initially by hybridization of RNA to a series of overlapping 40-kilobase-pair DNA fragments that were cloned in a cosmid vector. Further mapping was achieved with cloned HindIII restriction fragments. Results of these studies indicated that RNA polymerase subunit genes are transcribed early in infection and are distributed within the highly conserved central portion of the poxvirus genome in HindIII fragments E, J, H, D, and A. 相似文献
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Cytoplasmic transcription system encoded by vaccinia virus 总被引:17,自引:0,他引:17
B Moss B Y Ahn B Amegadzie P D Gershon J G Keck 《The Journal of biological chemistry》1991,266(3):1355-1358
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Structure of vaccinia virus early promoters 总被引:24,自引:0,他引:24
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Eryilmaz J Ceschini S Ryan J Geddes S Waters TR Barrett TE 《Journal of molecular biology》2006,357(1):62-72
The UvrABC pathway is a ubiquitously occurring mechanism targeted towards the repair of bulky base damage. Key to this process is UvrB, a DNA-dependent limited helicase that acts as a lesion recognition element whilst part of a tracking complex involving UvrA, and as a DNA-binding platform required for the presentation of damage to UvrC for subsequent processing. We have been able to determine the structure of a ternary complex involving UvrB* (a C-terminal truncation of full-length UvrB), a polythymine trinucleotide and ADP. This structure has highlighted the roles of key conserved residues in DNA binding distinct from those of the beta-hairpin, where most of the attention in previous studies has been focussed. We are also the first to report the structural basis underlying conformational re-modelling of the beta-hairpin that is absolutely required for DNA binding and how this event results in an ATPase primed for catalysis. Our data provide the first insights at the molecular level into the transformation of UvrB into an active helicase. 相似文献
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Purification and characterization of a new DNA-dependent ATPase with helicase activity from Escherichia coli 总被引:13,自引:0,他引:13
A previously unreported single-stranded DNA-dependent nucleoside 5'-triphosphatase with DNA unwinding activity has been purified from extracts of Escherichia coli lacking the F factor. Fractions of the purified enzyme contain a major polypeptide of Mr = 75,000 which contains the active site(s) for both ATP hydrolysis and helicase activity. This is consistent with the results of gel filtration chromatography which indicate a native molecular mass of 75 kDa. The 75-kDa helicase has a preference for ATP (dATP) as a substrate in the hydrolysis reaction and requires the presence of a single-stranded DNA cofactor. The helicase reaction catalyzed by the enzyme has been characterized using an in vitro strand displacement assay. The 75-kDa helicase displaces a 71-nucleotide DNA fragment in an enzyme concentration-dependent and time-dependent reaction. The helicase reaction depends on the presence of a hydrolyzable nucleoside 5'-triphosphate (NTP) suggesting that NTP hydrolysis is required for the unwinding activity. In addition, the enzyme can displace a 343-nucleotide DNA fragment albeit less efficiently. The direction of the unwinding reaction is 3' to 5' with respect to the strand of DNA on which the enzyme is bound. The molecular size of this helicase and the direction of the unwinding reaction are similar to both helicase II and Rep protein. However, the 75-kDa helicase has been shown to be distinct from both helicase II and Rep protein using immunological, physical, and genetic criteria. The discovery of a new helicase brings the total number of helicases found in E. coli cell extracts (lacking F factor) to five. 相似文献
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Growth inhibition by vaccinia virus growth factor 总被引:1,自引:0,他引:1
Vaccinia virus growth factor (VGF), a highly glycosylated 77-residue epidermal growth factor (EGF)-like polypeptide encoded in vaccinia poxvirus, is reported to play an important role in stimulating growth of uninfected cells to facilitate virus infection. We have chemically synthesized the unglycosylated forms of VGF and VGF19-69, a shortened VGF analog consisting of 51 residues and comprising the EGF-homologous region (position 19-69) of VGF. Both synthetic forms of VGFs were purified to homogeneity and vigorously characterized by various criteria, including the Cf-252 ion fission fragment mass spectrometry, amino acid sequencing, and enzymatic digestion to confirm the disulfide linkages. Synthetic VGFs exhibited high affinity binding to the EGF receptors in A431, NRK 49F, NRK clone 3, and NIH 3T3 cells, but, unlike the glycosylated form, showed contrasting mitogenic activities in various cells in vitro. Synthetic VGFs showed low levels of mitogenic and colonogenic activities in NRK clone 49F cells and NIH 3T3 cells, full agonist activities in human keratinocytes and Swiss 3T3 cells, and partial agonist activities in NRK clone 3 cells. Our results suggest that the unglycosylated form of VGF is an EGF antagonist to selected cells and that the production of unglycosylated form of VGF by the cytolytic vaccinia virus may serve as a mechanism whereby inhibition of growth and metabolism of selected host cells may be used to facilitate the propagation of the virus infection. 相似文献