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:4.1.99.3 (PRE)
1,923 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The ultraviolet-light induction of DNA damage has been measured in the epidermis of hairless mice with the use of damage-specific endonucleases from Micrococcus luteus. The rates of induction of endonuclease-sensitive sites in HRS/J/Anl and Skh:hairless-1 mice were 6.1 +/- 0.5 X 10(-11) and 6.5 +/- 0.8 X 10(-11)/dalton/J/sq m from a FS40 fluorescent sun lamp (280 to 400 nm), respectively. Enzymatic photoreactivation with yeast photoreactivating enzyme showed that approximately 80% of the endonuclease-sensitive sites were cycloburyl pyrimidine dimers. In both strains of mice the pyrimidine dimers remained in high-molecular-weight DNA for 24 hr after irradiation. These data show that mouse epithelial cells in vivo have little or no capacity for the excision repair of pyrimidine dimers.
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PMID:Induction and persistence of pyrimidine dimers in the epidermal DNA of two strains of hairless mice. 88 73

The action of an endonuclease from Micrococcus luteus that operates on UV damage in DNA overlaps with that of DNA photolyase from yeast: homo- and heterocyclobutane dipyrimidines in DNA are substrates for both enzymes, but pyrimidine adducts or the "spore photoproduct" in DNA are not. As expected from this overlap, the action of the two enzymes is mutually interfering: single-strand nicks introduced by the endonuclease effectively preclude photoreactivation; conversely, formation of a photolyase-cyclobutane dipyrimidine complex can prevent nicking by the endonuclease.
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PMID:Substrate specificity of Micrococcus luteus UV endonuclease and its overlap with DNA photolyase activity. 119 Nov 74

A DNA-binding protein specific for ultraviolet irradiated DNA has been purified extensively from human placenta. The binding preparation is free of exonuclease, polymerase, endonuclease, and N-glycosidase activity. The binding activity is salt dependent and is specific for double-stranded irradiated DNA. DNA from which the pyrimidine dimers have been monomerized by the action of photolyase (photoreactivating enzyme) remains an effective substrate for the binding protein, suggesting that the protein recognizes photoproducts other than pyrimidine dimers. This is supported by the finding that DNA irradiated under conditions which introduce only pyrimidine dimers is not a substrate for the binding protein. Examination of three of the xeroderma pigmentosum complementation groups has revealed no deficiency in this binding activity.
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PMID:A DNA binding protein from human placenta specific for ultraviolet damaged DNA. 127 48

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.
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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

To extend our knowledge of the excision repair system in mammalian cells we have focussed on the isolation of genes and proteins involved in this process. For the purification and characterization of human repair proteins the microneedle injection assay technique is utilized. This system is based on the transient correction of the excision repair defect of xeroderma pigmentosum (XP) fibroblasts (scored as increase of ultraviolet (u.v.)-induced unscheduled DNA synthesis (UDS) upon microinjection of crude extracts from complementing XP or normal cells. Specific correction is observed in fibroblasts of all (9) excision-deficient XP complementation groups. The XP-A and G correcting factors were found to be proteins and several purification steps (including (NH4)2SO4 fractionation, chromatography of phosphocellulose, heparin and u.v.-irradiated DNA-cellulose) have been worked out for the XP-A correcting protein. The microinjection system was also used for the introduction of (partially) purified repair enzymes of lower organisms. Micrococcus luteus endonuclease and bacteriophage T4 endonuclease V were able to correct all XP complementation groups tested, in marked contrast to the more sophisticated Escherichia coli uvrABC complex injected with uvrD. Photoreversal of dimers could be registered after introduction of the yeast photoreactivating enzyme in repair-competent, XP-variant, XP-C and XP-I fibroblasts (monitored as decrease of (residual) UDS). Remarkably, no effect was noticed in XP-A, D, E and H, suggesting that something prevents dimers in these cells from being monomerized by the injected enzyme. Using DNA-mediated gene transfer we have cloned a human gene (designated ERCC-1) that compensates for the excision defect of the u.v. and mitomycin C-sensitive Chinese hamster ovary cell (CHO) mutant 43-3B (complementation group 2). Characterization of this gene and its cDNA revealed the following features: (1) ERCC-1 corrects the full spectrum of repair deficiencies in mutants of complementation group 2. No correction is observed in mutants of the other CHO complementation groups. (2) The ERCC-1 gene has a size of 15 X 10(3) base-pairs (bp) and consists of 10 exons, one of which appears to be differentially spliced. (3) It encodes two largely identical mRNAs, which differ in the presence or absence of a 72 bp coding exon, situated in the 3' half of the mRNA. Only the cDNA of the large transcript is able to confer repair proficiency to 43-3B cells. No effect of u.v. treatment is found at the level of ERCC-1 transcription in HeLa cells.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Characterization of genes and proteins involved in excision repair of human cells. 282 Oct 19

Cyclobutyl pyrimidine dimers, measured as sites recognized by the dimer-specific ultraviolet (UV) endonuclease from Micrococcus luteus, were produced in DNA of human skin exposed in situ to UVA (320-400 nm) radiation. The dimer yields produced by a broadband UVA source, by broadband UVA filtered to remove all light of wavelength less than 340 nm, and by narrow band radiation centered at 365 nm were similar, indicating that UVA radiation, and not stray shorter wavelength radiation, was responsible for dimer production. The identity of the UVA-induced DNA lesions was confirmed as pyrimidine dimers by photoreactivation of approximately 100% of the endonuclease-sensitive sites in vitro with the 40,000 dalton Escherichia coli photoreactivating enzyme.
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PMID:Production of pyrimidine dimers in DNA of human skin exposed in situ to UVA radiation. 355 69

We have measured UVB (280-320 nm)-induced DNA damage in skin of individuals with different sensitivities to UVB irradiation as measured by minimal erythema dose (MED). The DNA damage was susceptible to cleavage by Micrococcus luteus UV endonuclease, which recognizes pyrimidine dimers in DNA. An alkaline agarose gel electrophoresis method was used to quantitate the number of M. luteus UV endonuclease-sensitive sites in nonradioactive DNA from skin biopsies of 7 individuals irradiated with UVB (0-180 mJ X cm-2). The production of sites correlated well with MED (correlation coefficient = 0.78). The slope of the dose response curve for the most UVB-sensitive individual (MED = 24 mJ X cm-2) and for the least UVB-sensitive individual (MED = 146 mJ X cm-2) were 11.5 X 10(-4) and 2.6 X 10(-4) sites per 1000 bases per mJ X cm-2, respectively. The UVB-induced DNA damage was determined to be pyrimidine dimers by its susceptibility to cleavage by M. luteus UV endonuclease and its photoreactivability by Escherichia coli photoreactivating enzyme.
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PMID:Higher pyrimidine dimer yields in skin of normal humans with higher UVB sensitivity. 375 38

Cultured human and embryonic chick fibroblasts possess different enzyme-mediated processes to repair cyclobutyl pyrimidine dimers induced in their deoxyribonucleic acid (DNA) by ultraviolet (UV) radiation. While dimers are corrected in human cells by excision repair, a photoenzymatic repair process exists in embryonic chick cells for the removal of these potentially deleterious UV photoproducts. We have utilized a sensitive enzymatic assay to monitor the disappearance, i.e. repair, of dimer-containing sites in fused populations of human and chick cells primarily consisting of multinucleate human/chick heterokaryons. Fused cultures were constructed such that UV photoproducts were present only in chick DNA when evaluating excision repair and only in human DNA when evaluating photoenzymatic repair. Based on the kinetics of site removal observed in these cultures we are led to conclude the following: Within heterokaryons per se the photoreactivating enzyme derived from chick nuclei and at least one excision-repair enzyme (presumably a UV endonuclease) derived from human nuclei act on UV-damaged DNA in foreign nuclei with an efficiency equal to that displayed toward their own nuclear DNA. Hence, after cell fusion these chick and human repair enzymes are apparently able to diffuse into foreign nuclei and once therein competently attack UV-irradiated DNA independently of its origin. In harmony with the situation in nonfused parental cultures, in heterokaryons the chick photoenzymatic repair process rapidly removed all dimer-containing sites from human DNA including the residual fraction normally acted upon slowly by the human excision-repair process.
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PMID:DNA repair in human/embryonic chick heterokaryons. Ability of each species to aid the other in the removal of ultraviolet-induced damage. 447 28

Chromosomeless "minicells" are formed by misplaced cell fissions near the polar extremities of an Escherichia coli K-12 mutant strain. Resistance (R)-factor deoxyribonucleic acid (DNA) can be introduced into minicells by segregation from an R(+) (R64-11) derivative of the original mutant. We have assessed the ability of R(+) minicells to correct defects produced in their plasmid DNA by ultraviolet (UV) and gamma radiations. Minicells harboring plasmid DNA, in comparison with their repair-proficient minicell-producing parents, possess (i) an equal competence to rejoin single-strand breaks induced in DNA by gamma rays, (ii) a reduced capacity for the photoenzymatic repair of UV-induced pyrimidine dimers, and (iii) a total inability to excise dimers, apparently owing to a deficiency in UV-specific endonuclease activity responsible for mediating the initial incision step in excision repair. Assuming that the DNA repair properties of R(+) minicells reflect the concentration of repair enzymes located in the plasmid-containing polar caps of entire cells, these findings suggest that: (i) the enzymes responsible for rejoining single-strand breaks are distributed throughout the cell; (ii) photoreactivating enzyme molecules tend to be concentrated near bacterial DNA and to a lesser extent near plasmid DNA; and (iii) UV-specific endonuclease molecules are primarily confined to the central region of the E. coli cell and, thus, seldom segregate with R-factor DNA into minicells.
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PMID:Photoreactivatiion, excision, and strand-rejoining repair in R factor-containing minicells of Escherichia coli K-12. 455 7

Partially purified extracts of Escherichia coli containing either uvrA+ or a mixture of uvrB+ and uvrC+ gene products were tested for an endonuclease activity on DNA treated with 8-methoxypsoralen plus 360-nm light. Neither of these fractions was active alone. The combined fractions, however, caused extensive strand cleavage of the psoralen-treated DNA. The endonuclease activity was dependent upon addition of ATP and Mg2+ to the reaction mixtures, and hence appeared similar to the UV-endonuclease activity previously shown to be reconstituted from the same fractions. It is concluded that the uvr+ gene products in these fractions interact to cause breakage of both psoralen-treated and UV-irradiated DNA. An examination of the dose-dependence relationship of the break formation in psoralen-treated DNA revealed that the enzyme acts upon psoralen mono-adducts. By varying the experimental conditions to increase the ratio of interstrand cross-links to mono-adducts it was found that the enzyme also acts upon cross-links, but with lower efficiency than for mono-adducts. Further studies of break formation in UV-irradiated DNA showed that elimination of pyrimidine dimers by treatment with photoreactivating enzyme in the light resulted in a loss of endonuclease-sensitive sites. This shows directly that pyrimidine dimers are the lesions recognized by the complemented uvr+ gene products in UV-irradiated DNA. For comparison, another endonuclease acting at pyrimidine dimers in DNA, the Micrococcus luteus UV-endonuclease, was also tested with psoralen-treated DNA, but no activity was observed. This and other data indicate that the repair endonuclease encoded by the uvr+ genes in E. coli is basically different from the other dimer-specific endonucleases previously characterized.
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PMID:Strand cleavage at psoralen adducts and pyrimidine dimers in DNA caused by interaction between semi-purified uvr+ gene products from Escherichia coli. 626 55


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