Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.1.30.2 (endonuclease)
 18,621 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Octadeoxynucleotides based on the recognition sequence of the restriction endonuclease NotI were synthesized containing unmodified nucleotides and nucleotides with methyl and bromide additions at the C5 position of the pyrimidine ring of deoxycytosine. On annealing to single-stranded DNA bearing one NotI site, thin-layer chromatography (TLC) of the different oligonucleotides was used quantitatively to determine differences in dissociation temperature (Td) and binding equilibrium. Buffers used in filter hybridization experiments could be used in this TLC system. In addition, actual hybridizations were carried out to filter-bound DNA with and without a NotI site. The incorporation of 5-methyldeoxycytosine and 5-bromodeoxycytosine led to a significant increase in stability of homoduplex formation during hybridization, due to a shift in the binding equilibrium and an increase of the Td, thereby improving discrimination considerably. Some implications of the results for several techniques involving oligomer hybridization are discussed.
 ...
PMID:Effect of 5-bromo- and 5-methyldeoxycytosine on duplex stability and discrimination of the NotI octadeoxynucleotide. Quantitative measurements using thin-layer chromatography. 239 67

The binding of various damaged DNAs to the single-strand binding protein coded for by gene 32 from bacteriophage T4, on the one hand, and of oligopeptides containing tryptophan and lysine residues, on the other hand, is described. These molecules exhibit a higher affinity for modified DNA than for native DNA in so far as modification results in a local destabilization of the double-stranded structure of the nucleic acid. Stacking interactions between aromatic amino acids and nucleic acid bases appear to play a crucial role in the recognition of destabilized regions induced by chemical agents (carcinogens and antitumor drugs). These interactions confer to the peptide lysyl-tryptophyl-lysine an endonucleolytic activity specific for apurinic sites. From results obtained with such oligopeptides a model for the active sites of Ap-endonucleases is proposed which could account for the strategy used by the denV endonuclease from phage T4 during the first step of excision repair of pyrimidine dimers in DNA. The effect of the overall conformation of modified DNA on repair efficiency is discussed.
 ...
PMID:Recognition of damaged regions in DNA by oligopeptides and proteins. 241 93

The "UV endonuclease" isolated either from M. luteus or bacteriophage T4 infected E. coli (the denV gene product) consists of two enzymatic activities on a single polypeptide chain: a pyrimidine dimer-DNA glycosylase and an AP endonuclease. The repair of pyrimidine dimers by this enzyme is initiated by the cleavage of the N-glycosylic bond of the 5' pyrimidine of the dimer that leaves the cyclobutane dimer still attached to the DNA through the N-glycosylic bond of the 3' pyrimidine of the dimer. This reaction results in the formation of an apyrimidinic site in the DNA. The second step in this repair pathway is the endonucleolytic cleavage of the DNA 3' to the AP site by the associated AP endonuclease. As a result, the nicked DNA contains DNA damage on both sides of the incision site: an apyrimidinic moiety on the 3' end and a thymine-thymidylate dimer on the 5' end. The enzymes prefer double stranded DNA over single stranded DNA, and thymine over cytosine at the 5' position of the dimer. The AP endonuclease activity prefers the AP site created by the pyrimidine dimer-DNA glycosylase on UV irradiated DNA over either apurinic or apyrimidinic DNA. This repair mechanism appears to be operative in vivo since DNA intermediates containing thymine-thymidylate dimer sites have been detected in UV irradiated T4 infected E. coli and in UV irradiated M. luteus. The cloned denV gene partially complements the UV repair deficient uvr A, B, C strains of E. coli.
 ...
PMID:The repair of pyrimidine dimers via a DNA-glycosylase mechanism. 242 65

We have developed a new assay which allows us to monitor the rates of repair of potentially lethal damage in u.v. (254 nm)-irradiated normal human skin fibroblasts. Using this assay we have shown that, in non-dividing cells, the majority of biologically effective excision repair is completed within 4 h following irradiation with low fluences of u.v. (1.5-6.0 J/m2). During this time, non-dividing cells removed only approximately 20% of the pyrimidine dimers induced in DNA by a u.v. fluence of 3.0 J/m2 as measured by the loss of u.v.-endonuclease-sensitive sites under identical repair conditions. The rates of repair of potentially lethal damage were also found to be independent of u.v. fluence over the range 1.5-6.0 J/m2 in non-dividing cells. In contrast, in cells irradiated in exponential growth with 1.5 J/m2, the rate of biologically effective repair was comparable with that observed in non-dividing cells but the efficiency of the repair process declined progressively with increase in u.v. fluence from 1.5 to 6.0 J/m2. Our data support the concept that the biological recovery of u.v.-irradiated cells depends on the preferential repair of damage in functionally important domains in the genome.
 ...
PMID:Rapidly occurring DNA excision repair events determine the biological expression of u.v.-induced damage in human cells. 244 87

The UV endonucleases [endodeoxyribonuclease (pyrimidine dimer), EC 3.1.25.1] from Micrococcus luteus and bacteriophage T4 possess two catalytic activities specific for the site of cyclobutane pyrimidine dimers in UV-irradiated DNA: a DNA glycosylase that cleaves the 5'-glycosyl bond of the dimerized pyrimidines and an apurinic/apyrimidinic (AP) endonuclease that thereupon incises the phosphodiester bond 3' to the resulting apyrimidinic site. We have explored the potential use of methoxyamine, a chemical that reacts at neutral pH with AP sites in DNA, as a selective inhibitor of the AP endonuclease activities residing in the M. luteus and T4 enzymes. The presence of 50 mM methoxyamine during incubation of UV- (4 kJ/m2, 254 nm) treated, [3H]thymine-labeled poly(dA).poly(dT) with either enzyme preparation was found to protect completely the irradiated copolymer from endonucleolytic attack at dimer sites, as assayed by yield of acid-soluble radioactivity. In contrast, the dimer-DNA glycosylase activity of each enzyme remained fully functional, as monitored retrospectively by release of free thymine after either photochemical- (5 kJ/m2, 254 nm) or photoenzymic- (Escherichia coli photolyase plus visible light) induced reversal of pyrimidine dimers in the UV-damaged substrate. Our data demonstrate that the inhibition of the strand-incision reaction arises because of chemical modification of the AP sites and is not due to inactivation of the enzyme by methoxyamine. Our results, combined with earlier findings for 5'-acting AP endonucleases, strongly suggest that methoxyamine is a highly specific inhibitor of virtually all AP endonucleases, irrespective of their modes of action, and may therefore prove useful in a wide variety of DNA repair studies.
 ...
PMID:Selective inhibition by methoxyamine of the apurinic/apyrimidinic endonuclease activity associated with pyrimidine dimer-DNA glycosylases from Micrococcus luteus and bacteriophage T4. 244 60

A novel endonuclease has been isolated from extracts of spinach leaves (Spinacia oleracea). The enzyme has been purified by a series of column chromatography steps and has a molecular size of approximately 43,000 daltons. The spinach endonuclease cleaved double stranded DNA damaged by ultraviolet light or cis-diamminedichloroplatinum (II) primarily at sites of adenine when end-labelled DNA fragments of defined sequence were employed as substrates. The nature of the structural distortion contained in damaged, duplex DNA appears to be an important determinant for endonuclease cleavage. DNA helical distortions produced by UV light-induced (6-4) pyrimidine-pyrimidone photoproducts, but not cyclobutane pyrimidine dimers are recognized by the enzyme. The DNA cleavage products generated by the enzyme contain 3'-hydroxyl and 5'-phosphoryl termini. Single stranded DNA and RNA are hydrolyzed by the spinach endonuclease. This enzyme, which we call nuclease SP, is similar in several respects to other single-strand-specific nucleases such as N. crassa and mung bean nucleases and may function in DNA repair and/or recombination events in spinach cells. Nuclease SP should be a useful tool for the analysis of (6-4) photoproducts occurring in duplex DNA.
 ...
PMID:Nuclease SP: a novel enzyme from spinach that incises damaged duplex DNA preferentially at sites of adenine. 245 4

The role of exonuclease III and endonuclease IV in the repair of pyrimidine dimers in bacteriophage T4-infected Escherichia coli was examined. UV-irradiated T4 showed reduced survival when plated on an xth nfo double mutant but showed wild-type survival on either single mutant. T4 denV phage were equally sensitive when plated on wild-type E. coli or an xth nfo double mutant, suggesting that these endonucleases function in the same repair pathway as T4 pyrimidine dimer-DNA glycosylase. A uvrA mutant of E. coli in which the repair of pyrimidine dimers was dependent on the T4 denV gene carried on a plasmid was constructed. Neither an xth nor an nfo derivative of this strain was more sensitive than the parental strain to UV irradiation. We were unable to construct a uvrA xth nfo triple mutant. In addition, T4, which turns off the host UvrABC excision nuclease, showed reduced plating efficiency on an xth nfo double mutant.
 ...
PMID:Role of exonuclease III and endonuclease IV in repair of pyrimidine dimers initiated by bacteriophage T4 pyrimidine dimer-DNA glycosylase. 246 48

Bacteriophage-T4 UV endonuclease nicks the C(3')-O-P bond 3' to AP (apurinic or apyrimidinic) sites by a beta-elimination reaction. The breakage of this bond is sometimes followed by the nicking of the C(5')-O-P bond 5' to the AP site, leaving a 3'-phosphate end; delta-elimination is proposed as a mechanism to explain this second reaction. The AP site formed when this enzyme acts on a pyrimidine dimer in a polynucleotide chain undergoes the same nicking reactions. Micrococcus luteus UV endonuclease also nicks the C(3')-O-P bond 3' to AP sites by a beta-elimination reaction. No subsequent delta-elimination was observed, but this might be due to the presence of 2-mercaptoethanol in the enzyme preparation.
 ...
PMID:Bacteriophage-T4 and Micrococcus luteus UV endonucleases are not endonucleases but beta-elimination and sometimes beta delta-elimination catalysts. 247 12

Micrococcus luteus UV endonuclease incises DNA at the sites of ultraviolet (UV) light-induced pyrimidine dimers. The mechanism of incision has been previously shown to be a glycosylic bond cleavage at the 5'-pyrimidine of the dimer followed by an apyrimidine endonuclease activity which cleaves the phosphodiester backbone between the pyrimidines. The process by which M. luteus UV endonuclease locates pyrimidine dimers within a population of UV-irradiated plasmids was shown to occur, in vitro, by a processive or "sliding" mechanism on non-target DNA as opposed to a distributive or "random hit" mechanism. Form I plasmid DNA containing 25 dimers per molecule was incubated with M. luteus UV endonuclease in time course reactions. The three topological forms of plasmid DNA generated were analyzed by agarose gel electrophoresis. When the enzyme encounters a pyrimidine dimer, it is significantly more likely to make only the glycosylase cleavage as opposed to making both the glycosylic and phosphodiester bond cleavages. Thus, plasmids are accumulated with many alkaline-labile sites relative to single-stranded breaks. In addition, reactions were performed at both pH 8.0 and pH 6.0, in the absence of NaCl, as well as 25,100, and 250 mM NaCl. The efficiency of the DNA scanning reaction was shown to be dependent on both the ionic strength and pH of the reaction. At low ionic strengths, the reaction was shown to proceed by a processive mechanism and shifted to a distributive mechanism as the ionic strength of the reaction increased. Processivity at pH 8.0 is shown to be more sensitive to increases in ionic strength than reactions performed at pH 6.0.
 ...
PMID:Modulation of the DNA scanning activity of the Micrococcus luteus UV endonuclease. 247 71

Although the biological role of many bacterial repair genes is known, there is still an interest in evaluating the capacity of repair pyrimidine dimers in some strains. For this purpose, we have developed a rapid assay. Cells bearing a plasmid are UV irradiated and incubated to allow recovery. The plasmid DNA is extracted, purified and treated with UV endonuclease from Micrococcus luteus that specifically produces single strand breaks at the site of pyrimidine dimers. The amount of open circular and covalently closed circular forms of the plasmid DNA after treatment and post-incubation provides an estimate of the repair capability of the host strain. The wild type strain and the uvrA mutant of Escherichia coli were used to adjust the assay. The lexA mutant of E. coli has been tested and its repair capability is equivalent to that of wild-type strain. The assay has been extended to Streptococcus pneumoniae, which is naturally deficient in photoreactivation and SOS-like functions. This strain is efficient in the repair of pyrimidine dimers, formed after UV irradiation.
 ...
PMID:Excision-repair capacity of UV-irradiated strains of Escherichia coli and Streptococcus pneumoniae, estimated by plasmid recovery. 249 85


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>