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1.
A highly specific endonuclease activity, presumably involving one or both of the products of the ØX174 gene A, has been isolated from ØX174-infected by DNA-cellulose chromatography. The enzyme is not present in uninfected cells and binds extremely tightly to DNA-cellulose. It extensively degrades ØX174 viral DNA but does not degrade the circular or linear forms of single stranded viral DNA of either M13, an unrelated filamentous phage, or G4, a ØX-type phage. 相似文献
2.
W T Hsu E J Lin P P Fu R G Harvey S B Weiss 《Biochemical and biophysical research communications》1979,88(1):251-257
The ØX174 DNA assay system developed earlier is utilized to determine the comparative reactivities with nucleic acid of the diolepoxide metabolites of a series of polycyclic aromatic hydrocarbons varying in carcinogenic potency. The infectious ØX174 viral DNA is exposed to the hydrocarbon derivative for 10 min., then infectivity of the treated DNA is assayed by incubation with spheroplasts, counting plaque formation on agar plates. The bay region diolepoxides of benzo[a]-pyrene, chrysene, and dibenz[a,h]anthracene, implicated as the ultimately active carcinogenic metabolites of the parent hydrocarbons, exhibit potent viral inhibitory activity. On the other hand no correlation is evident between viral inhibitory activity and either the location of the diolepoxide function in a bay region or the theoretically calculated β-delocalization energies (ΔEdeloc.) of the carbonium ion arising from opening the epoxide ring. The significance of these findings with respect to theories of carcinogenesis is discussed. 相似文献
3.
The synthesis of β-galactosidase (EC 3.2.1.23: β-D-galactoside galactohydrolase) in is repressed as a result of infection with single-stranded DNA phage ØX174. An mutant in ØX174 cistron A, which codes for two proteins, does not inhibit the enzyme synthesis while mutants in all other genes do cause repression. A mutant near the amino-terminal end of cistron A, which produces the small 35,000 molecular weight cistron A polypeptide, also inhibits the synthesis of β-galactosidase. Inhibition is also observed in an mutant which does not support the replication of replicative-form DNA. Exogenous nucleotide bases and cyclic 3′,5′-adenosine monophosphate (cyclic AMP) do not have any effect on the degree of repression. 相似文献
4.
Selective inhibition of in vitro DNA synthesis dependent on phiX174 compared with fd DNA. I. Protein requirements for selective inhibition. 总被引:8,自引:0,他引:8
Crude extracts of Escherichia coli selectively convert fd viral DNA and not phiX174 DNA to duplex DNA via a complex series of reactions one of which involves RNA polymerase. Reactions leading to formation of fd duplex-replicative (RFII) structures have been reconstituted with purified proteins from E. coli. Maximal synthesis requires the combined action of E. coli binding protein, DNA elongation factor I, DNA elongation factor II preparations (which are a mixture of dna Z and DNA elongation factor III), DNA polymerase III, DNA-dependent RNA polymerase, Mg2+, dATP, dGTP, dCTP, dTTP, and ATP, GTP, CTP, and UTP. In contrast to crude extracts of E. coli, purified protein fractions do not distinguish between fd DNA and phiX174 DNA in duplex DNA formation. The addition of crude fractions of E. coli to the purified components listed above selectively permits fd RFII formation and prevents phiX RFII formation. This selective inhibition was used as an assay to isolate proteins essential for this phenomenon; they include RNase H, discriminatory factor alpha, and discriminatory factor beta. 相似文献
5.
At 25°C rifampicin strongly stimulates the synthesis of the dinucleotide pppA-U catalyzed by the DNA-dependent RNA polymerase from . If the antibiotic is added to the enzyme during the synthesis of RNA the stimulatory effect on the dinucleotide synthesis is distinctly retarded as is its inhibitory action on RNA synthesis. It is proposed that this lag period is due to a retardation of the binding of rifampicin to RNA polymerase which is required for its action. Because of this slower binding rifampicin — although an inhibitor of RNA chain elongation — mimics the action of an inhibitor of RNA chain initiation. 相似文献
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7.
B K Tye P O Nyman I R Lehman 《Biochemical and biophysical research communications》1978,82(2):434-441
Each of the stages in the replication of ØX174 DNA , e. g., conversion of circular single stranded parental DNA to the duplex replicative form (SS → RF), replication of the closed circular duplex form (RF → RF), and synthesis of circular single stranded progeny DNA (RF →SS), may be affected by a reduced level of dUTPase. Thus, in enzyme preparations from mutant strains defective in dUTPase (?), the complementary strand synthesized in the SS → RF reaction is abnormally short (7–8S vs. 14S), and the extent of RF replication is decreased 10-fold. Preferential removal of dUTPase during fractionation of enzyme preparations from wild type (+) cells may produce comparable effects. In particular, the single stranded circular DNA synthesized in the RF → SS reaction by a set of highly purified enzymes is rapidly degraded upon incubation with the less pure enzymes required for its conversion to RF. All of these effects are plausibly accounted for by the incorporation into DNA of uracil from dUTP, possibly present as a contaminant in one or more components of the reaction, followed by excision of the uracil and phosphodiester bond cleavage at the resulting apyrimidinic site. 相似文献
8.
Neomycin inhibits DNA dependent DNA and RNA synthesis catalyzed by DNA polymerase I and RNA polymerase from . The effect of the antibiotic is more pronounced towards DNA synthesis. The inhibition of DNA synthesis is competitive with template DNA, does not reverse with excess deoxynucleoside triphosphate, Mg2+ or enzyme DNA polymerase I. Neomycin does not reduce the number of potential 3′ -OH end or primer. It seems to shorten the size of the newly formed polynucleotide. 相似文献
9.
D Twardzik J Simonds M Oskarsson F Portugal 《Biochemical and biophysical research communications》1973,52(3):1108-1114
High molecular weight RNA isolated from the oncogenic type C murine leukemia virus, AKR-MuLV, stimulates the incorporation of radioactive amino acids into protein in an cell-free system. Analysis of the translational products by SDS polyacrylamide gel electrophoresis demonstrated the synthesis of at least three proteins corresponding in molecular weight to several authentic viral proteins. Positive immunoprecipitation tests also confirm the translational product as AKR-MuLV related. Although at least 18 proteins were found on analysis of disrupted murine leukemia virions, only three were synthesized in response to AKR-MuLV RNA in the cell-free system. 相似文献
10.
The effect of depurination of polynucleotide templates on the fidelity of DNA synthesis in vitro has been determined. The fidelity of DNA synthesis with Escherichia coli DNA polymerase I, avian myeloblastosis virus DNA polymerase and human placenta DNA polymerase-β is decreased as a result of depurination of the poly[d(A-T)], poly[d(G-C)]and poly[d(A)]templates. The error rate with poly[d(A-T)]increased from to using E. coli Pol I, and from to using the myeloblastosis virus DNA polymerase. Depurination of poly[d(A)]increased the error rate from to using E. coli Pol I, and from to using the DNA polymerase-β from human placenta. Depurination of poly[d(G-C)]resulted in an increase in the error rate with E. coli Pol I from to , and with the virus DNA polymerase from to . This misincorporation is shown to be directly proportional to the extent of depurination. Deletion experiments and alkaline sucrose gradient analyses suggest that the incorporation of complementary and non-complementary nucleotides is dependent on polymerization, and occurs in the same newly synthesized product. Kinetic studies and nearest-neighbor analyses indicate that the incorporation of non-complementary nucleotides occurs randomly as single-base substitutions. The nearest-neighbor studies also suggest that any of the four deoxynucleotides can be incorporated opposite apurinic sites. The number of each nucleotide incorporated relative to the number of apurinic sites was determined to be for dGTP, for dCTP, for dATP and for dTTP with both the poly[d(A-T)] and poly[d(A)] templates. The frequencies of misincorporation relative to the number of apurinic sites with the poly[d(G-C)]template were for dATP, for dTTP, for dGTP and for dCTP. Hydrolysis at the apurinic sites by alkali treatment reversed the effects of depurination on fidelity. The error rates with the depurinated templates were reduced to within 2% of those obtained prior to depurination, providing additional evidence that the misincorporation after depurination results from apurinic sites on the template. These results suggest a possible relationship between depurination of DNA and errors in DNA replication and/or repair. 相似文献
11.
George Boguslawski William S. Oetting Dean A. Stetler 《Biochemical and biophysical research communications》1977,78(2):684-689
An enzyme, ribonucleotide polymerase, isolated from the yeast phase of a fungus, has been found to stimulate the incorporation of dTMP in the reaction catalysed by DNA polymerase from and . The stimulation is dependent on the amount of ribonucleotide polymerase added. The data indicate that protein-protein interaction is responsible for the increase in DNA synthesis. It is suggested that ribonucleotide polymerase may be involved in supplying short RNA primers for DNA polymerase. 相似文献
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D Ohmori 《Biochemical and biophysical research communications》1976,72(2):566-574
Mutants of C were selected for resistance towards a set of cell wall LPS core specific bacteriophages, including øX174. Increasingly deficient LPS's from wt and mutant C were tested for inactivation of øX174, and the core oligosaccharides were subjected to structural analysis by methylation/g.l.c./m.s. Loss of the terminal galactose in the following basic structure of the C wt core was found to lead to adsorption resistance towards øX174: 相似文献
14.
DNA polymerase activities in cell-free lysates of unfertilized eggs, larvae and immature ovaries of were compared to purified DNA polymerase I using several natural and synthetic templates. The templates were tested as the native and denatured forms of normal and DNase I treated molecules. Although the polymerases tended to prefer DNase I treated Xenopus DNA over the other templates tested, so did the polymerase I. In general, the template preferences of the polymerases studied depended in complex ways on both the form and the species of origin of the template. 相似文献
15.
S Friedman 《Biochemical and biophysical research communications》1979,89(4):1328-1333
5-Azacytidine, when added to growing K12, causes a decrease in DNA methylation assayed . This decrease is greater when DNA is used as substrate than when calf thymus DNA is used. The decrease in activity is not due to the inhibition of protein synthesis caused by this drug, since neither chloramphenicol nor rifampin causes a decrease in enzyme activity. The effect is specific for the DNA(cytosine-5)methylase; the methylation of adenine is not affected. The concentration of drug that inhibits the DNA methylase by 50% is the same concentration that inhibits cell growth by 50%. 相似文献
16.
K Schwerzmann P Gazzotti E Carafoli 《Biochemical and biophysical research communications》1976,69(3):812-815
We have determined that Co2+, Ni2+ or Zn2+ may substitute for Mg2+ during DNA synthesis with DNA polymerase I, sea urchin nuclear DNA polymerase and the DNA polymerase from avian myeloblastosis virus (AMV). In addition, the frequency of non-complementary nucleotide incorporation using AMV DNA polymerase was increased using Co2+ or Mn2+ as the metal activator. These results suggest that the fidelity of DNA synthesis may be influenced by the metal activator used during catalysis. 相似文献
17.
Excision of thymine dimers by proteolytic and amber fragments of E. coli DNA polymerase I 总被引:11,自引:0,他引:11
Excision of thymine dimers from specifically incised ultraviolet irradiated DNA by DNA polymerase I is stimulated by concurrent DNA synthesis. The 36,000 molecular-weight “small fragment” obtained by limited proteolysis of DNA polymerase I, which retains only the 5′ → 3′ exonuclease activity, also excises thymine dimers, but at one-tenth the rate of the intact enzyme. However, the rate of excision is increased by addition of the “large” 76,000-molecular weight fragment. With the further addition of the 4 deoxynucleoside triphosphates, permitting DNA synthesis to occur, excision approaches rates observed with the intact enzyme. The same result was obtained with a fragment of DNA polymerase I with 5′ → 3′ exonuclease activity that is present uniquely in polymerase I mutants. 相似文献
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20.
Stimulation of T7 DNA polymerase by a new phage-coded protein 总被引:10,自引:0,他引:10
Eberhard Scherzinger Frank Litfin Erich Jost 《Molecular & general genetics : MGG》1973,123(3):247-262
Summary A bacteriophage-induced DNA-binding protein was purified from T7 infected E. coli. The protein has a molecular weight of about 25000, as judged by SDS-polyacrylamide gel electrophoresis. The purified protein binds to single-stranded but not to native T7 DNA. Like the T4 gene-32 protein and the 22000-dalton unwinding protein of E. coli, the T7 25000 protein lowers the melting temperature of poly d(A-T). Using partially single-stranded T7 DNA as template-primer, the protein stimulates in vitro DNA synthesis by T7 DNA polymerase about five-fold. It was also found that the DNA-unwinding protein of E. coli stimulates T7 DNA polymerase to approximately the same extent. However, neither of the unwinding proteins stimulate DNA polymerase I of E. coli. 相似文献