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)

Mitomycin C and certain analogues alkylate DNA with their C-1 position and cross-link it by a second alkylation involving C-10. We now show that monoalkylation by C-10 (carbamate group) can occur for mitosene analogues that have no reactive C-1 functionality. Sodium dithionite reduction of 2,7-diaminomitosene or 2,7-diamino-1-hydroxymitosene in the presence of calf thymus DNA resulted in alkylation of the DNA to the extent of one molecule per 14 and 11 bases, respectively, although no covalent binding was observed on catalytic reduction. Reduction of each of these mitosenes by sodium dithionite in the presence of 2'-deoxyguanosine gave monoalkylation on the 2-amino group of this nucleotide. The 2,7-diaminomitosenes inhibited L-1210 leukemia cell colony formation in vitro at concentrations 3-4-fold greater (less potent) than mitomycin C. DNA single-strand breaks were also produced by each mitosene, but these lesions did not correlate with cytotoxicity and were less prominent than breaks produced by another monofunctional alkylating agent, methyl methanesulfonate. Mitosene-induced DNA strand breaks are probably due to excission-repair endonuclease activity and not from oxygen free radicals produced by redox cycling of the quinone moiety. There was no evidence of DNA-DNA cross-links by either 2,7-diaminomitosene.
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PMID:Alkylation of DNA by C-10 of 2,7-diaminomitosene. 229 22

The self-complementing dodecamer 5'-CGCGAATTCGCG-3' and its complexes with the antibiotic netropsin and the restriction endonuclease EcoRI provide substrates of known three-dimensional structure to study the stereochemistry and mechanism of the artificial nuclease of 1,10-phenanthroline-copper ion [(OP)2Cu+]. Analysis of the reaction products with the 5'-32P dodecamer on 20% sequencing gels has demonstrated the presence of 3'-phosphoglycolate ends in addition to 3'-phosphomonoester ends expected from previous studies. A reaction intermediate, which is a precursor to 3'-phosphomonoester termini, has been trapped; in contrast, no comparable species for the 5'-phosphomonoester termini can be detected when 3'-labeled DNAs are utilized as substrates. The reactive oxidative species formed by the coreactants (OP)2Cu+ and hydrogen peroxide is distinguishable in its chemistry from the hydroxyl radicals produced by cobalt-60 gamma-irradiation. The freely diffusible hydroxyl radicals generated by cobalt-60 irradiation produce equivalent amounts of 3'-phosphomonoester and 3'-phosphoglycolate termini whereas the 3'-phosphomonoesters are the preferred product of (OP)2Cu+ and H2O2. On the basis of the structures of the products obtained, the principal site of attack of the coordination complex is on the C-1 of the deoxyribose within the minor groove. This conclusion is supported by the footprinting of netropsin binding to the dodecamer. Crystallographic results have demonstrated that netropsin binds to the minor groove at the central AATT residue. A clear protection of attack by the coordination complex at the deoxyriboses associated with A-5, T-6, T-7, and C-9 is fully consistent with attack from the minor groove without intercalation during the course of the cleavage reaction.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Nuclease activity of 1,10-phenanthroline-copper ion: reaction with CGCGAATTCGCG and its complexes with netropsin and EcoRI. 302 54

Detergent-lysed BS-C-1, HeLa, and mouse L cells incorporate ADP-ribose from NAD+ into two classes of macromolecules. Metabolically stable products, which appear to be a variety of proteins to which are attached one or a few ADP-ribose residues, predominate when the cellular DNA remains intact. In addition, ghost cells have a potentially much greater capacity to synthesize poly(ADP-ribose), which is completely dependent upon the introduction of strand breaks into their DNA. The initial rate of poly(ADP-ribose) synthesis increases linearly with prior x-ray dose or with the concentration of endonuclease added and, once synthesized, the polymer is rapidly degraded with a half-life of 10 min or less. It appears that sites on the DNA capable of supporting a certain amount of poly(ADP-ribose) synthesis are created as a result of x-irradiation or nucleolytic cleavage and are rapidly eliminated, or "repaired," during subsequent incubation. The sites accumulate if cells are irradiated at 0 degree C; further incubation of the lysed cells with NAD+ at 35 degrees C results in both a burst of poly(ADP-ribose) synthesis and the elimination of the sites. NAD+ enhances the elimination of x-ray-induced sites. Thus, the synthesis of poly(ADP-ribose) may be required for the repair of DNA strand breaks.
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PMID:ADP-ribosylation in mammalian cell ghosts. Dependence of poly(ADP-ribose) synthesis on strand breakage in DNA. 743 Jan 32

The recognition of 'regular' and 'oxidized' sites of base loss (AP sites) in DNA by various AP endonucleases was compared. Model substrates with regular AP sites (resulting from mere hydrolysis of the glycosylic bond) were produced by damaging bacteriophage PM2 DNA by exposure to low pH; those with AP sites oxidized at the C-4'- and C-1'-position of the sugar moiety by exposure to Fe(III)-bleomycin in the presence of H2O2 and to Cu(II)-phenanthroline in the presence of H2O2 and ethanol, respectively. The results confirmed that AP sites-together with single-strand breaks-are indeed the predominant type of DNA modification in all three cases. For the recognition of 4'-oxidized AP sites, a 400-fold higher concentration of Escherichia coli exonuclease III and between 5-fold and 50-fold higher concentrations of bacteriophage T4 endonuclease V, E. coli endonuclease III and E. coli FPG protein were required than for the recognition of regular AP sites. In contrast, the recognition of 4'-oxidized AP sites by E. coli endonuclease IV was effected by 4-fold lower concentrations than needed for regular AP sites. 1'-oxidized AP sites (generated by activated Cu(II)-phenanthroline) were recognized by endonuclease IV and exonuclease III only slightly (3-fold and 13-fold, respectively) less efficiently than regular AP sites. In contrast, there was virtually no recognition of 1'-oxidized AP sites by the enzymes which cleave at the 3' side of AP sites (T4 endonuclease V, endonuclease III and FPG protein). The described differences were exploited for the analysis of the DNA damage induced by hydroxyl radicals, generated by ionizing radiation or Fe(III)-nitrilotriacetate in the presence of H2O2. The results indicate that both regular and 1'-oxidized AP sites represent only minor fractions of the AP sites induced by hydroxyl radicals.
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PMID:Recognition of oxidized abasic sites by repair endonucleases. 751 77

A number of repair endonuclease, viz. endonuclease III, formamidopyrimidine-DNA glycosylase (FPG protein), endonuclease IV, exonuclease III and UV endonuclease, is used to simultaneously quantify various types of DNA modifications, which were induced by agents that generate reactive oxygen species. Under cell-free conditions, two types of DNA damage profiles are obtained. The profiles induced by chemically generated singlet oxygen and by various photosensitizers (acridine orange, methylene blue, riboflavin, hematoporphyrin) plus light are dominated by base modifications sensitive to FPG protein, while 5,6-dihydropyrimidines (recognized by endonuclease III), sites of base loss (AP sites, recognized by endonuclease IV and exonuclease III) and strand breaks are minor lesions. In contrast, the DNA damage profile induced by hydroxyl radicals (gamma-rays) consists of approx. equal levels of base modifications. AP sites and strand breaks. The damage profiles induced by Fe(III)-EDTA in the presence of superoxide and by Fe(III)-nitrilotriacetate in the presence of H2O2 do not differ from that by hydroxyl radicals. The damage profile induced by Cu(II)-phenanthroline deviates by high levels of AP sites that are recognized by endonuclease IV and exonuclease III-but not by those AP endonucleases which cleave at the 3' site-and probably represent AP sites oxidized at C-1'. The damage induced by Fe(III)-bleomycin plus H2O2 deviates by an increased level of double strand breaks and the absence of endonuclease-sensitive base modifications. Cellular DNA damage profiles are obtained from bacteria, cultured mammalian cells and mammalian mitochondria after exposure to acridine orange plus visible light. A comparison with the cell-free profiles reveals that the damage in all three systems is not induced indirectly by hydroxyl radicals or an activation of cellular nucleases, but by the same mechanism that is responsible for the cell-free DNA damage.
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PMID:Use of repair endonucleases to characterize DNA damage induced by reactive oxygen species in cellular and cell-free systems. 838 92

Oxidative damage to DNA deoxyribose generates oxidized abasic sites (OAS) that may constitute one-third of ionizing radiation damage. The antitumor drug bleomycin produces exclusively OAS in the form of C-4-keto-C-1-aldehydes in unbroken DNA strands and 3'-phosphoglycolate esters terminating strand breaks. We investigated whether two human DNA repair enzymes can mediate OAS excision in vitro: Ape1 protein (the main human abasic endonuclease (also called Hap1, Apex, or Ref1)) and DNA polymerase beta, which carries out both the abasic excision and the resynthesis steps. We used a duplex oligonucleotide substrate with one main target for bleomycin-induced damage. Ape1 catalyzed effective incision at the C-4-keto-C-1-aldehyde sites at a rate that may be only a few-fold lower than incision of hydrolytic abasic sites at the same location. Consistent with several previous studies, Ape1 hydrolyzed 3'-phosphoglycolates 25-fold more slowly than C-4-keto-C-1-aldehydes. DNA polymerase beta excised the 5'-terminal OAS formed by Ape1 incision at a rate similar to its removal of unmodified abasic residues. Polymerase beta-mediated excision of 5'-terminal OAS was stimulated by Ape1 as it is for unmodified abasic sites. Escherichia coli Fpg (MutM) protein also excised 5'-terminal OAS, but in our hands, the RecJ protein did not. These observations help define mammalian pathways of OAS repair, point to interactions that might coordinate functional steps, and suggest that still unknown factors may contribute to removal of 3'-phosphoglycolate esters.
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PMID:Excision of C-4'-oxidized deoxyribose lesions from double-stranded DNA by human apurinic/apyrimidinic endonuclease (Ape1 protein) and DNA polymerase beta. 978 84

Oxidized abasic residues in DNA constitute a major class of radiation and oxidative damage. Free radical attack on the nucleotidyl C-1' carbon yields 2-deoxyribonolactone (dL) as a significant lesion. Although dL residues are efficiently incised by the main human abasic endonuclease enzyme Ape1, we show here that subsequent excision by human DNA polymerase beta is impaired at dL compared with unmodified abasic sites. This inhibition is accompanied by accumulation of a protein-DNA cross-link not observed in reactions of polymerase beta with unmodified abasic sites, although a similar form can be trapped by reduction with sodium borohydride. The formation of the stably cross-linked species with dL depends on the polymerase lysine 72 residue, which forms a Schiff base with the C-1 aldehyde during excision of an unmodified abasic site. In the case of a dL residue, attack on the lactone C-1 by lysine 72 proceeds more slowly and evidently produces an amide linkage, which resists further processing. Consequently dL residues may not be readily repaired by "short-patch" base excision repair but instead function as suicide substrates in the formation of protein-DNA cross-links that may require alternative modes of repair.
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PMID:Covalent trapping of human DNA polymerase beta by the oxidative DNA lesion 2-deoxyribonolactone. 1180 79

The medicinal chemistry and structure-activity relationships (SAR) for a novel series of carbamoyl pyridone bicycle (CAB) compounds as influenza Cap-dependent endonuclease (CEN) inhibitors are disclosed. Substituent effects were evaluated at the C (N)-1, N-3, and C-7 positions of the CAB ring system using a docking study. Submicromolar EC₅₀ values were achieved in the cellular assay with C-7-unsubstituted CAB which possessed a benzhydryl group on either the C-1 or the N-1 position. An N-3 substituent was found to be critical for the plasma protein binding effect in vitro, and the CAB-N analogue 2v exhibited reasonable total clearance (CL_tot). More importantly, compound 2v displayed significant efficacy in a mouse model infected with influenza viruses.
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PMID:Synthesis and SAR Study of Carbamoyl Pyridone Bicycle Derivatives as Potent Inhibitors of Influenza Cap-dependent Endonuclease. 3138 63

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