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:3.1.30.2 (endonuclease)
18,621 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The cytotoxic action of the antitumor antibiotic mitomycin C occurs primarily at the level of DNA. Using highly sensitive fluorescence assays which depend on the enhancement of ethidium fluorescence only when it intercalates duplex regions of DNA, three aspects of mitomycin C action on DNA have been studied: (a) cross-linking events, (b) alkylation without necessarily cross-linking, and (c) strand breakage. Cross-linking of DNA is determined by the return of fluorescence after a heat denaturation step at alkaline pH's. Under these conditions denatured DNA gives no fluorescence. The cross-linking was independently confirmed by S1-endonuclease (EC 3.1.4.-) digestion. At relatively high concentrations of mitomycin the suppression of ethidium fluorescence enhancement was shown not to be due to depurination but rather to alkylation, as a result of losses in potential intercalation sites. A linear relationship exists between binding ratio for mitomycin and loss of fluorescence. The proportional decrease in fluorescence with pH strongly suggests that the alkylation is due to the aziridine moiety of the antibiotic under these conditions. A parallel increase in the rate and overall efficiency of covalent cross-linking of DNA with lower pH suggests that the cross-linking event, to which the primary cytotoxic action has been linked, occurs sequentially with alkylation by aziridine and then by carbamate. Mitomycin C, reduced chemically, was shown to induce single strand cleavage as well as monoaklylation and covalent cross-linking in PM2 covalently closed circular DNA. The inhibition of this cleavage by superoxide dismutase (EC 1.15.1.1) and catalase (EC 1.11.1.6), and by free radical scavengers suggests that the degradation of DNA observed to accompany the cytotoxic action of mitomycin C is largely due to the free radical O2. In contrast to the behavior of the antibiotic streptonigrin, mitomycin C does not inactivate the protective enzymes superoxide dismutase or catalase. Lastly, mitomycin C is able to cross-link DNA in the absence of reduction at pH 4. This is consistent with the postulated cross-linking mechansims.
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PMID:Studies related to antitumor antibiotics. Part V. Reactions of mitomycin C with DNA examined by ethidium fluorescence assay. 0 1

307 cultures of staphylococci and micrococci were isolated from clinical sources and were subjected to 8 different physiological tests for the recognition of pathogentic strains. These tests were as follows; catalase, oxidase, fermentation and oxidation of glucose and mannitol, v.p. and coagulase activity, heat resistance endonuclease, and lipase production. From the results obtained it can be concluded that for the recognition of pathogenicity of staphylococci, coagulase, heat resistance endonuclease and fermentation of mannitol should be carried out.
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PMID:Evaluation of 8 physiological characteristics of clinically isolated Staphylococci and Micrococci. 80 76

Cigarette smoke can cause DNA single strand breaks in cultured human lung cells (T. Nakayama et al., Nature, 314 (1985) 462-464) but the mechanisms behind this DNA damage have not been clearly elucidated. In the present study we have investigated the possibility that one of the major constituents in cigarette smoke, hydroquinone, may be important for mediating smoke-induced DNA damage in the human epithelial lung cell line, A 549, and the mechanisms behind this damage. Cells were exposed to cigarette smoke, hydrogen peroxide, or hydroquinone, in the absence and presence of different inhibitors, and the resulting DNA damage was assessed either as DNA single strand break formation or formation of the oxidative DNA adduct, 8-hydroxydeoxyguanosine. It was found that (i) exposure to cigarette smoke, hydrogen peroxide or hydroquinone causes a rapid decrease in the intracellular thiol level and a considerable DNA single strand break formation, (ii) the formation of DNA single strand breaks in cells exposed to cigarette smoke is inhibited by catalase, dimethylthiourea, and o-phenantroline, suggesting that hydroxyl radicals generated from iron-catalyzed hydrogen peroxide dissociation are involved in the DNA damage, (iii) hydroquinone causes considerable DNA strand break formation that is blocked by aurintricarboxylic acid, an inhibitor of endonuclease activation, and by BAPTA, an intracellular calcium chelator, (iv) addition of hydroquinone to a smoke condensate greatly enhances its ability to cause DNA single strand breaks, and (v) smoke, but not hydroquinone, causes formation of 8-hydroxydeoxyguanosine, a DNA damage product induced by the action of hydroxyl radicals on the DNA base, deoxyguanosine. These findings suggest that the ability of cigarette smoke to cause DNA single strand breaks in cultured lung cells is due to mechanisms involving hydroxyl radical attack on DNA and endonuclease activation. They also suggest that hydroquinone is an important contributor to the DNA damaging effect of cigarette smoke on human lung cells.
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PMID:Cigarette smoke-induced DNA damage in cultured human lung cells: role of hydroxyl radicals and endonuclease activation. 130 85

Vaccinia virus-dependent CAT expression was observed in virus-infected cells cotransfected with a promoterless CAT gene. Restriction endonuclease resection of the CAT plasmid indicated that expression was due to recognition by vaccinia virus RNA polymerase of sequences within the CAT gene itself, probably located within the 5' untranslated region of the gene. This observation is relevant to the design of reverse-genetic systems which use CAT as a reporter gene to detect replication of negative-strand RNA virus pseudogenomes.
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PMID:Tn9 CAT gene contains a promoter for vaccinia virus transcription: implications for reverse-genetic techniques. 131 41

Nine strains of Helicobacter pylori have been isolated exhibiting spontaneous mutations with a loss of catalase activity. Growth characteristics in vitro were unaffected by the mutation showing that catalase is not essential for growth of Helicobacter pylori. Parent strains and mutants could not be distinguished morphologically from each other when compared by electron microscopy. Restriction endonuclease digestion with HindIII, separated in an 0.7% agarose gel in TBE buffer, showed each pair to be highly related to each other. SDS-PAGE separation of proteins from four mutants and parent strains showed that all mutants lacked a 57 kDa protein. The partial N-terminal sequence of this protein shows homology with maize catalase and may be the subunit of the Helicobacter pylori catalase tetramer. It is concluded that catalase negative mutants of Helicobacter pylori occur spontaneously in vitro, but have not yet been observed in vivo. The paucity of such catalase negative strains in clinical specimens could mean that catalase is a virulence factor in vivo that puts mutants at a selective disadvantage.
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PMID:Catalase negative mutants of Helicobacter pylori. 152 35

Enkephalins are opiate peptides found in a variety of tissues including brain and pituitary. In brain, they function as neurotransmitters, neuromodulators and neurohormones. Recent studies show that proenkephalin mRNA is expressed early in development both in mammals and the amphibian, suggesting that enkephalins may play a unique role in embryogenesis. In order to characterize factors which regulate the onset and patterning of expression of this gene in adult and developing frog embryos, the proenkephalin A gene was cloned from Xenopus laevis. The clones have been characterized by DNA sequencing and restriction endonuclease mapping. The gene is made up of three exons which span approximately 12 kb. Exon I encodes the 5' untranslated region of the mRNA. Exon II contains the signal peptide and the N terminus of the mature protein. Biologically active opioid peptides are generated from exon III. Comparison to mammalian proenkephalin genomic sequence indicated that nucleotide sequences of the 5' flanking region, noncoding exon I and exon II were not well conserved but exon III was highly conserved. Primer extension and RNase protection assay analyses of the RNA transcripts revealed two major 5' ends. The putative TATA box, CAAT box, CRE and Pit 1 elements have been identified on this gene by sequence homology to published consensus sequences. To assay for sequences that could potentially regulate Xenopus proenkephalin expression, we transfected constructs that contained upstream genomic sequences linked to the CAT reporter gene into various eukaryotic cell lines. The expression of the fusion gene constructs were detected and could be induced 10- to 30-fold upon treatment with forskolin.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Characterization of Xenopus laevis proenkephalin gene. 172 92

In order to understand the regulation of basic fibroblast growth factor (bFGF) gene expression, we have cloned and characterized the human bFGF gene and its regulatory elements. Using restriction endonuclease digestion, we have mapped the entire gene and sequenced all intron/exon boundaries to confirm authenticity and to determine organization. The data show that intron 1 is at least 16 kb long while intron 2 is 16 kb long. The human bFGF gene, including its three exons, is therefore at least 36 kb long. There are five GC boxes which may represent SP-1 binding sites and one potential AP-1 binding site within the core promoter region. Primer extension analysis indicates the presence of one bFGF-RNA transcription start site. We used a standard bacterial CAT gene expression system to identify the DNA sequence containing the functional bFGF gene promoter. Deletion analysis suggests the presence of two negative regulatory elements; one in the non-transcribed 5'-promoter region and the other within transcribed (but non-translated) sequences 3' of the promoter core.
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PMID:Functional characterization of the human basic fibroblast growth factor gene promoter. 176 64

A mutant strain of Helicobacter pylori with weak urease activity was created by using N-methyl-N'-nitro-N-nitrosoguanidine. The urease activity of the mutant (0.036 +/- 0.009 nmol of urea per micrograms of bacterial protein per min) was 0.4% of that of the parental strain (8.20 +/- 2.30 nmol of urea per micrograms of bacterial protein per min). The mutant was otherwise indistinguishable from the parental strain. Both demonstrated prominent catalase and oxidase activities, and both produced vacuolating cytotoxin. Restriction endonuclease and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) patterns and ultrastructure were identical for the two strains. The mutant was fully motile, as evaluated by spreading in soft agar and by direct microscopic examination. Growth rate and colony size and morphology were identical for the mutant and parental strains. Seventeen gnotobiotic piglets were challenged with either the mutant or the parental strain and sacrificed 3 or 21 days after challenge. Gastric tissue was examined histologically and cultured for H. pylori. Of seven piglets challenged with the parental strain, all became infected. H. pylori was not recovered from any of 10 piglets challenged with the urease-negative strain. Lymphofollicular gastritis was present in all seven piglets challenged with the parental strain but in none of the piglets challenged with the urease-negative strain. These results suggest that prominent urease activity is essential for colonization by H. pylori.
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PMID:Essential role of urease in pathogenesis of gastritis induced by Helicobacter pylori in gnotobiotic piglets. 205 Apr 11

Exposure of the plasmid pBR 322 to the aerobic xanthine oxidase reaction introduced single strand scissions and endonuclease III-sensitive sites. The latter may be residues of thymine glycol. Both forms of DNA damage were completely prevented by superoxide dismutase or catalase, whereas bovine serum albumin was much less effective. Mannitol and benzoate, added as scavengers of HO., and desferrioxamine or diethylene triamine pentaacetate, added to sequester Fe(III), also protected. These results indicate a metal-catalyzed interaction of O2- with H2O2, which produces HO. which, in turn, causes DNA strand scission and oxidation of thymine residues to thymine glycol. Plasmid isolated from aerobically-incubated cells contained more strand scissions and endonuclease III-sensitive sites than did plasmid from anaerobically-incubated cells, and a low molecular weight scavenger of O2- prevented the damage seen with the aerobic cells. Genetic defects in AP endonucleases rendered E. coli more susceptible to the dioxygen-dependent lethality of plumbagin, which mediates O2- production. Similarly, plasmid DNA, within the endonuclease-deficient cells, exhibited more strand scissions and endonuclease III-sensitive sites upon aerobic exposure to plumbagin than did endonuclease-sufficient cells, and a low molecular weight scavenger of O2- was protective. These results are consistent with the conclusions that strand scissions and formation of endonuclease III-sensitive sites are among the consequences of exposure of DNA to O2- plus H2O2, both in vitro and in vivo.
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PMID:Formation of endonuclease III-sensitive sites as a consequence of oxygen radical attack on DNA. 254 64

We developed a rapid extraction method to analyze the chromosomal DNA of 84 isolates of Campylobacter pylori (formerly Campylobacter pyloridis) from 70 individuals. Only three of the nine endonucleases tested gave satisfactory digestions: HindIII, EcoRI, and SacI. The latter two produced mostly larger bands, whereas HindIII produced smaller bands, which allowed clearer comparisons between isolates. The isolates from 69 Australian subjects and one from England each had a unique profile. Isolates from a husband and wife were different, as were those from a brother and sister. In pairs of isolates from 11 individuals, the second isolate was markedly different in six subjects. The profile changes were not associated with changes in antibiotic susceptibility or with loss of catalase or urease activity, which can occur during storage of C. pylori. These restriction endonuclease profiles suggest considerable subspecies variation in C. pylori. Plasmid bands were found in undigested DNA from 40 of the 84 C. pylori isolates.
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PMID:Restriction endonuclease analysis of the genome of Campylobacter pylori with a rapid extraction method: evidence for considerable genomic variation. 283 Mar 41


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