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:2.7.7.7 (DNA polymerase)
17,007 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

4'-(9-Acridinylamino)methanesulphon-m-anisidide (AMSA) (NSC 141549), an acridine derivative with activity against a variety of laboratory tumors in vivo, is presently undergoing Phase 1 clinical evaluation. The interaction of AMSA with DNA and its effects on nucleic acid-polymerizing enzymes were examined in an attempt to define the site of cytotoxicity of AMSA. Binding of AMSA to DNA, as demonstrated by equilibrium dialysis and spectrophotometric methods, appears to be similar to other aminoacridines, in that two types of binding sites (type 1 and type 2) were observed. Fluorescence studies and thermal denaturation studies gave strong evidence that AMSA type 1 binding was by intercalation into DNA. The binding of AMSA to DNA was without marked base-pair specificity. Furthermore, the effect of AMSA on nucleic acid-polymerizing enzyme activities (mouse embryo DNA polymerase alpha, avian myeloblastosis virus reverse transcriptase, and Escherichia coli RNA polymerase) was studied. Inhibition of enzyme activity by AMSA appeared to be independent of DNA base sequence. The relatively high concentrations of AMSA required for inhibition of these enzymes as compared to the concentrations of AMSA necessary for cytotoxicity in vitro suggest that the interaction with DNA alone might not fully explain its antitumor activity.
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PMID:Interaction of 4'-(9-acridinylamino)methanesulfon-m-anisidide with DNA and inhibition of oncornavirus reverse transcriptase and cellular nucleic acid polymerases. 7 12

Ether-permeabilized (nucleotide-permeable) Escherichia coli cells respond to alkylating and arylalkylating carcinogens with DNA excision repair, as assessed by their stimulation of DNA repair synthesis. In the present work, we have investigated whether DNA repair synthesis in ether-treated E. coli cells can serve as a general indicator to monitor the DNA-binding of carcinogens, mutagens and antitumor agents. Therefore, a standard assay was developed and comparative analyses were performed on 11 ultimate carcinogens, 10 proximate carcinogens, 2 tumor promoters, 6 mutagens, and 12 antitumor agents. All ultimate carcinogens (alkylating, acylating, arylalkylating agents) and mutagens (e.g., hydrogeen peroxide, acridine derivatives) caused DNA excision repair in wild type cells as measured by [3H] dTMP incorporation and simultaneously inhibited replicative DNA synthesis to various extents. Control experiments with the mutant cells uvrA and uvrB were performed to determine whether the pyrimidine-dimer-specific UV-endonuclease was involved in the removal of DNA damage. This was found to be true for the ultimate carcinogens (Ac)2 ONFln, mitomycin C, and for very reactive alkylating carcinogens. None of the ultimate carcinogens induced repair polymerization in mutant cells lacking the 5'-3' exonucleolytic activity of DNA polymerase I. Proximate carcinogens, such as Me2NNO, 4-nitroquinoline-1-oxide and aflatoxins, did not induce excision repair in the standard assay, probably because of the inability of E. coli to perform the activation steps necessary for covalent DNA-binding. However, Me2NNO, when pretreated with Udenfriend's hydroxylating mixture, gave rise to a low level of repair polymerization in ether-treated cells. Intercalating mutagens, such as quinacrine and ethidum bromide, inhibited replicative DNA synthesis. However, they were not found to be repair-inducers. THE TUMOR PROMOters TPA and phorbol-12,13-didecanoate did not cause excision repair, even when applied at high concentrations, nor did they inhibit repair synthesis stimulated by MeNOUr or (Ac)2 ONFln. The antitumor agents may be classified into two groups on the basis of the influence they exert on DNA synthesis: members of the first group (involving BCNU and bleomycin) stimulate repair polymerization and, in addition, inhibit DNA replication. These compounds are known to bind covalently to DNA. The second group of drugs (including adriamycin and cis-Pt(II)diammine complexes) inhibits DNA replication without stimulating repair synthesis. The predominant DNA-interaction of these compounds is known to be a non-covalent (i.e., intercalative, electrostatic) binding. Our experiments show that the ether-permeabilized E. coli cell can be successfully used to test ultimate carcinogens, mutagens and antitumor agents for repair-inducing and replication-inhibiting activity. The standard test might be extended to pre- and proximate carcinogens, provided these can be suitably activated.
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PMID:The nucleotide-permeable Escherichia coli cell, a sensitive DNA repair indicator for carcinogens, mutagens, and antitumor agents binding covalently to DNA. 15 98

Five mutants of Salmonella typhimurium strain LT2 trp DI (ColEI)+, initiallly detected because they released little or no colicin when tested on solid medium, proved to be sensitive to ultraviotet light (u.v.). Further testing indicated that one of the mutants was deficient in genetic recombination and was probably a recA-type mutant, while three of the others were deficient in DNA polymerase activity and appeared to be typical polA mutants. The fifth mutant was less sensitive than the others to methyl methanesulphonate, showed reduced proficiency in genetic recombination, and was of approximately normal u.v. mutability. This mutant may be a counterpart of the class known as uvrD in Escherichia coli. All five mutants degraded significantly more of their DNA following exposure to u.v. than did the wild-type strain. The recA-type mutant and the possible uvrD mutant also degraded significantly more of their DNA spontaneously than did the wild-type. Treatment with visible light and acridine orange (photodynamic treatment) cause no significant degradation of DNA in the wild-type strain, a highly significant increase in the extent of DNA degradation in a polA mutant, and a decrease in the extent of degradation in the recA-type mutant.
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PMID:DNA degradation in wild-type and repair-deficient strains of salmonella typhimurium exposed to ultraviolet light of photodynamic treatment. 33 Aug 21

DNA polymerase gamma from purified nuclei of EMT-6 cells (mice) seems to be identical to the mitochondrial DNA polymerase from the same source following several criteria. These two enzyme activities are strongly inhibited by ethidium bromide and acriflavin, while proflavin, acridine orange, daunomycin and chloroquine inhibition is less pronounced. In the case of DNA polymerases alpha and beta very little inhibition by ethidium bromide was observed. Intercalation of this dye in a poly dA-dT 12-18 template-primer was studied spectrophotometrically under conditions similar to those in the in vitro DNA polymerase assay. The polymerase assay. The inhibition by this drug of the mitochondrial DNA polymerase gamma activity was shown to be competitive at varying concentrations of TTP while the inhibition was of the non-competitive type at different concentrations of poly dA-dT 12-18. We conclude that the drug, most probably in the intercalated form, is able to interact with the active site (s) of mitochondrial DNA polymerase.
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PMID:The inhibition of mitochondrial DNA polymerase gamma from animal cells by intercalating drugs. 67 50

Treatment of growing cultures of Mycobacterium smegmatis with alkylating agents (methyl methanesulphonate, ethyl methanesulphonate, nitrogen mustard, or mitomycin C) or with ultraviolet light resulted in enhanced specific activities of a DNA polymerase and of an ATP-dependent deoxyribonuclease. Similar results had previously been obtained with hydroxyurea and with iron limitation. The three of these treatments which were tested (methyl methanesulphonate, mitomycin C and hydroxyurea) produced strand breaks or alkali-labile regions in the DNA of this organism. The increased enzyme activities could be prevented by simultaneous treatment with inhibitors of protein synthesis. In contrast, treatment of the cultures with intercalating agents (ethidium bromide, acridine orange, or proflavine), 5-fluorouracil, caffeine, or nalidixic acid, inhibited DNA synthesis without increasing the enzyme activities. These treatments did not produce strand breaks in the DNA of this organism. The results support the hypothesis that, in M. smegmatis, damage to DNA induces increased synthesis of enzymes associated with DNA repair.
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PMID:Increased DNA polymerase and ATP-dependent deoxyribonuclease activities following DNA damages in mycobacterium smegmatis. 84 85

The effect of intercalating compounds such as 9-aminoacridine, quinacrine (atebrin), proflavine and daunomycin on the activity of DNA polymerase I(EC 2.7.7.7) was studied in vitro and compared with the binding of these acridines to native DNA. The enzyme kinetics were followed at various concentrations of DNA 3'-OH primer end groups and constant concentrations of deoxynucleosidetriphosphates as well as under the opposite conditions. The Km values for the DNA 3'-OH end groups were 16--38 nM and for the deoxynucleosidetriphosphates 2--5 micrometer, depending on the buffer and pH used in the enzymatic assay. All acridine derivates inhibit the DNA polymerase; at variable DNA concentrations a competitive inhibition was observed, where the Ki values ranged between 0.87 and 8.5 micrometer. At variable concentrations of deoxynucleosidetriphosphates and constant DNA concentration a non-competitive inhibition was observed. On denatured 3'-OH DNA as well as on poly(dA) - (dT)10 as substrate no inhibition by 9-aminoacridine was observed. 5'--3' exonuclease activity of DNA polymerase is inhibited by 9-aminoacridine but 3'--5' exonuclease activity on denatured DNA is not influenced by this intercalating compound. The affinity of the acridines to DNA was determined spectrophotometrically under conditions similar to those in the enzymatic assay and the computed frequency of intercalation was related to the inhibition of enzymatic activity. The mechanism of inhibition is explained by a disturbance of the structure of the double helical DNA due to the interaction of the bound acridine derivates.
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PMID:Action of intercalating agents on the activity of DNA polymerase I. 92 1

The sequence specificity of DNA damage caused by cis-diamminedichloroplatinum(II) (cisplatin) and four analogues in human (HeLa) cells was studied using Taq DNA polymerase and a linear amplification system. The primer extension is inhibited by the drug-DNA adducts, and hence the sites of these lesions can be analyzed on DNA sequencing gels. The repetitive alphoid DNA was used as the target DNA in human cells. A comparison was made between adduct formation in human cells and in purified DNA. The sequence-specific position and relative intensity of damage was similar in both systems for cisplatin, dichloro(ethylenediammine)platinum(II) (PtenCl2), and N-[3-N-(ethylenediamino)propyl]acridine-4-carboxamidedichloropl atinum(II) (4AcC3PtenCl2). However, no DNA damage could be detected in cells for trans-diamminedichloroplatinum(II) (transPt) or N-[3-N-(ethylenediamino)propyl]acridine-2-carboxamide-dichloroplat inum(II) (2AcC3PtenCl2) despite the ability of these latter analogues to damage purified DNA. Cisplatin, PtenCl2, and 4AcC3PtenCl2, which significantly damaged DNA inside cells, also show antitumor activity in mouse models. However, transPt and 2AcC3PtenCl2, which did not detectably damage DNA inside cells, did not show such antitumor activity. This correlation between intracellular DNA damaging ability and in vivo antitumor activity indicates the potential use of the human cells/Taq DNA polymerase/linear amplification technique as a convenient method for screening new cisplatin analogues for useful chemotherapeutic activity.
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PMID:An investigation of the sequence-specific interaction of cis-diamminedichloroplatinum(II) and four analogues, including two acridine-tethered complexes, with DNA inside human cells. 144 14

A series of reagents containing 3- or 4-nitrobenzamido ligands tethered to 9-aminoacridine via variable-length linkers have been prepared and their properties as photochemical DNA cleavers (photonucleases) examined. When irradiated with approximately 300-nm light, where the nitrobenzamido ligand can absorb, they cleave DNA in an oxygen-independent reaction presumably involving oxygen transfer from the nitro group to the deoxyribose units of the DNA backbone (Nielsen et al., 1988b). This reaction is pH independent and only slightly affected by the linker length, and the DNA fragments are not substrates for DNA polymerase. When approximately 420-nm light is used, were only the 9-aminoacridinyl ligands absorb, the DNA cleavage is also oxygen-independent but pH dependent, requires DNA saturation with the reagent (base pair:reagent less than or equal to 2), and is most efficient with the longer linkers. The cleavage is specific for guanine residues and results in 5'-phosphate termini and heterogeneous (more than four products) 3'-termini. One of the products is presumably 3'-hydroxy since DNA photocleaved with nitrobenzamido acridine reagents and 420-nm radiation are substrates for DNA polymerase in a nick translation assay as well as for the Klenow fragment. An electron-transfer mechanism is suggested.
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PMID:Photolytic cleavage of DNA by nitrobenzamido ligands linked to 9-aminoacridines gives DNA polymerase substrates in a wavelength-dependent reaction. 165 88

The purpose of the present study was to examine the distribution pattern of electron-dense acridine orange (AO) chromatin interaction products in rat glioma C6 cells at different phases of the cell cycle. For synchronization in the early S-phase the cells in logarithmic growth were treated with 3 micrograms/ml aphidicolin, a specific inhibitor of DNA polymerase alpha and then cultured in normal medium. For synchronization in the M-phase the cells cultured with aphidicolin and then returned to normal medium were treated with 0.05 micrograms/ml colcemid. Histoautoradiographic analysis of the C6 cells using the pulse chase method demonstrated approximately 16 h of cell cycle time and about 6.5 h of S-phase. Ultracytochemically, AO chromatin interaction products were found in all phases of the cell cycle except for the mitotic phase, namely in G1, S, and G2. The highest percentage of AO chromatin interaction products was observed in the early S-phase and the lowest in the G2 phase. The mean number of AO chromatin interaction products per nuclear area increased in the course of S-phase parallel with an increase of 3H-uridine uptake during the S-phase. The results show a characteristic distribution pattern of AO label specific for each of the four stages of the cell cycle, however, the significance of the coincident RNA synthetic activity remains to be elucidated.
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PMID:Distribution pattern of acridine orange chromatin interaction products in rat glioma C6 cells at different phases of the cell cycle. 169 Apr 12

Various kinds of DNA damage block the 3' to 5' exonuclease action of both E. coli exonuclease III and T4 DNA polymerase. This study shows that a variety of DNA damage likewise inhibits DNA digestion by lambda exonuclease, a 5' to 3' exonuclease. The processive degradation of DNA by the enzyme is blocked if the substrate DNA is treated with ultraviolet irradiation, anthramycin, distamycin, or benzo[a]-pyrene diol epoxide. Furthermore, as with the 3' to 5' exonucleases, the enzyme stops at discrete sites which are different for different DNA damaging agents. On the other hand, digestion of treated DNA by lambda exonuclease is only transiently inhibited at guanine residues alkylated with the acridine mustard ICR-170. The enzyme does not bypass benzo[a]-pyrene diol epoxide or anthramycin lesions even after extensive incubation. While both benzo[a]-pyrene diol epoxide and ICR-170 alkylate the guanine N-7 position, only benzo[a]-pyrene diol epoxide also reacts with the guanine N-2 position in the minor groove of DNA. Anthramycin and distamycin bind exclusively to sites in the minor groove of DNA. Thus lambda exonuclease may be particularly sensitive to obstructions in the minor groove of DNA; alternatively, the enzyme may be blocked by some local helix distortion caused by these adducts, but not by alkylation at guanine N-7 sites.
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PMID:Lesion selectivity in blockage of lambda exonuclease by DNA damage. 169 29


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