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)

Human cytochrome P450IIE1 (CYP2E) is involved in the metabolic activation of procarcinogens such as N-nitrosodimethylamine, benzene and ethyl carbamate. We screened DNA from 28 individuals for restriction fragment length polymorphisms (RFLPs) is the human P450IIE1 gene and detected an RFLP for the restriction endonuclease DraI. The distribution of the genotypes of this polymorphisms among lung cancer patients (n = 74) differed from that among controls (n = 73) with statistical significance of p < 0.05. In addition, the distribution among patients with cancers of the digestive system (n = 38) was also different from that among controls. Our findings indicate an association between the DraI polymorphism of the IIE1 gene and susceptibility to cancers of the lung and the digestive system.
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PMID:Human cytochrome P450IIE1 gene: DraI polymorphism and susceptibility to cancer. 136 74

Cytochrome P450IIE1 (P450IIE1) is involved in metabolic activation of carcinogenic nitrosamines, aniline and benzene. We detected a restriction fragment length polymorphism of the human P450IIE1 gene with the restriction endonuclease DraI. The population was thus divided into three genotypes, namely, heterozygotes (CD) and two forms of homozygotes (CC and DD). The distribution of these genotypes among lung cancer patients differed from that among controls with statistical significance of P less than 0.05 (chi 2 = 7.01 with 2 degrees of freedom). This result strongly suggests that host susceptibility to lung cancer is associated with the DraI polymorphism of the P450IIE1 gene.
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PMID:Association between restriction fragment length polymorphism of the human cytochrome P450IIE1 gene and susceptibility to lung cancer. 167 75

Plasmid pRO1957 contains a 26.5-kb BamHI restriction endonuclease-cleaved DNA fragment cloned from the chromosome of Pseudomonas pickettii PKO1 that allows P. aeruginosa PAO1c to grow on toluene, benzene, phenol, or m-cresol as the sole carbon source. The genes encoding enzymes for meta cleavage of catechol or 3-methylcatechol, derived from catabolism of these substrates, were subcloned from pRO1957 and were shown to be organized into a single operon with the promoter proximal to tbuE. Deletion and analysis of subclones demonstrated that the order of genes in the meta cleavage operon was tbuEFGKIHJ, which encoded catechol 2,3-dioxygenase, 2-hydroxymuconate semialdehyde hydrolase, 2-hydroxymuconate semialdehyde dehydrogenase, 4-hydroxy-2-oxovalerate aldolase, 4-oxalocrotonate decarboxylase, 4-oxalocrotonate isomerase, and 2-hydroxypent-2,4-dienoate hydratase, respectively. The regulatory gene for the tbuEFGKIHJ operon, designated tbuS, was subcloned into vector plasmid pRO2317 from pRO1957 as a 1.3-kb PstI fragment, designated pRO2345. When tbuS was not present, meta pathway enzyme expression was partially derepressed, but these activity levels could not be fully induced. However, when tbuS was present in trans with tbuEFGKIHJ, meta pathway enzymes were repressed in the absence of an effector and were fully induced when an effector was present. This behavior suggests that the gene product of tbuS acts as both a repressor and an activator. Phenol and m-cresol were inducers of meta pathway enzymatic activity. Catechol, 3-methylcatechol, 4-methylcatechol, o-cresol, and p-cresol were not inducers but could be metabolized by cells previously induced by phenol or m-cresol.
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PMID:Genetic organization and regulation of a meta cleavage pathway for catechols produced from catabolism of toluene, benzene, phenol, and cresols by Pseudomonas pickettii PKO1. 185 61

A 26-kilobase BamHI restriction endonuclease DNA fragment was cloned from Pseudomonas pickettii PKO1, a strain isolated from a soil microcosm that had been amended with benzene, toluene, and xylene. This DNA fragment, cloned into vector plasmid pRO1727 and designated pRO1957, allowed Pseudomonas aeruginosa PAO1c to grow on phenol as the sole source of carbon. Physical and functional restriction endonuclease maps have been derived for the cloned DNA fragment. Two DNA fragments carried in trans and derived from subclones of pRO1957 show phenol hydroxylase activity in cell extracts of P. aeruginosa. Deletion and subcloning analyses of these fragments indicated that the gene encoding phenol hydroxylase is positively regulated. Phenol and m-cresol were shown to be inducers of the enzyme. o-Cresol and p-cresol did not induce enzymatic activity but could be metabolized by cells that had been previously exposed to phenol or m-cresol; moreover, the enzyme exhibited a rather broad substrate specificity and was sensitive to thiol-inhibiting reagents. A novel polypeptide with an estimated molecular mass of 80,000 daltons was detected in extracts of phenol-induced cells of P. aeruginosa carrying plasmid pRO1959.
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PMID:Molecular cloning, characterization, and regulation of a Pseudomonas pickettii PKO1 gene encoding phenol hydroxylase and expression of the gene in Pseudomonas aeruginosa PAO1c. 211 72

In order to study the toluene and o-xylene catabolic genes of Pseudomonas stutzeri OX1, a genomic library was constructed. A 28-kb EcoRI restriction endonuclease DNA fragment, cloned into the vector plasmid pLAFR1 and designated pFB3401, permitted Pseudomonas putida PaW340 to convert toluene and o-xylene into the corresponding meta-ring fission products. Physical and functional endonuclease restriction maps have been derived from the cloned DNA fragment. Further subcloning into and deletion analysis in the Escherichia coli vector pGEM-3Z allowed the genes for the conversion of toluene or o-xylene into the corresponding catechols to be mapped within a 6-kb region of the pFB3401 insert and their direction of transcription to be determined. Following exposure to toluene, E. coli cells carrying this 6-kb region produce a mixture of o-cresol, m-cresol, and p-cresol, which are further converted to 3-methylcatechol and 4-methylcatechol. Similarly, a mixture of 2,3-dimethylphenol and 3,4-dimethylphenol, further converted into dimethylcatechols, was detected after exposure to o-xylene. The enzyme involved in the first step of toluene and o-xylene degradation exhibited a broad substrate specificity, being able to oxidize also benzene, ethylbenzene, m-xylene, p-xylene, styrene, and naphthalene. Deletions of the 6-kb region which affect the ability to convert toluene or o-xylene into the corresponding methylphenols compromise also their further oxidation to methylcatechols. This suggests that a single enzyme system could be involved in both steps of the early stages of toluene and o-xylene catabolism.
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PMID:Cloning of the genes for and characterization of the early stages of toluene and o-xylene catabolism in Pseudomonas stutzeri OX1. 883 26

The major human apurinic/apyrimidinic (AP) endonuclease (class II) is known to cleave DNA 5' adjacent to an AP site, which is probably the most common DNA damage produced hydrolytically or by glycosylase-mediated removal of modified bases. p-Benzoquinone (pBQ), one of the major benzene metabolites, reacts with DNA to form bulky exocyclic adducts. Herein we report that the human AP endonuclease directly catalyzes incision in a defined oligonucleotide containing 3,N4-benzetheno-2'-deoxycytidine (pBQ-dC) without prior generation of an AP site. The enzyme incises the oligonucleotide 5' to the adduct and generates 3'-hydroxyl and 5'-phosphoryl termini but leaves the pBQ-dC on the 5' terminus of the cleavage fragment. The AP function of the enzyme is not involved in this action, as no preexisting AP site is present nor is a DNA glycosylase activity involved. Nicking of the pBQ-dC adduct also leads to the same "dangling base" cleavage when two Escherichia coli enzymes, exonuclease III and endonuclease IV, are used. Our finding of this unusual mode of action used by both human and bacterial AP endonucleases raises important questions regarding the requirements for substrate recognition and catalytic active site(s) for this essential cellular repair enzyme. We believe this to be the first instance of the presence of a bulky carcinogen adduct leading to this unusual mode of action.
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PMID:An unusual mechanism for the major human apurinic/apyrimidinic (AP) endonuclease involving 5' cleavage of DNA containing a benzene-derived exocyclic adduct in the absence of an AP site. 894 4

Chronic exposure of humans to benzene (BZ) causes acute myeloid leukemia (AML). Both BZ and therapy-related secondary AML are characterized by chromosomal translocations that may occur by inappropriate recombinational events. DNA topoisomerase II (topo II) is an essential sulfhydryl (SH)-dependent endonuclease required for replication, recombination, chromosome segregation, and chromosome structure. Topo II cleaves DNA at purine(R)/pyrimidine(Y) repeat sequences that have been shown to be highly recombinogenic in vivo. Certain antineoplastic drugs stabilize topo II-DNA cleavage complexes at RY repeat sequences, which leads to translocations of the type observed in leukemia. Hydroquinone (HQ) is metabolized to p-benzoquinone (BQ) in a peroxidase-mediated reaction in myeloid progenitor cells. BQ interacts wit SH groups of SH-dependent enzymes. Consequently, the aims of this research were to determine whether HQ and BQ are topo II inhibitors. The ability of the compounds to inhibit the activity of topo III was tested using an assay system that depends on the conversion, by homogeneous human topo II, of catenated kinetoplast DNA into open and/or nicked open circular DNA that can be separated from the catenated DNA by electrophoresis in a 1% agarose-ethidium bromide gel. We provide preliminary data that indicate that both HQ and BQ cause a time and concentration (microM)-dependent inhibition of topo II activity. These compounds, which potentially can form adducts with DNA, have no effect on the migration of the supercoiled and open circular forms in the electrophoretic gradient, and BQ-adducted KDNA can be decatenated by topo II. Using a pRYG plasmid DNA with a single RY repeat as a cleavage site, it was determined that BQ does not stimulate the production of linear DNA indicative of an inhibition of topo II religation of strand breaks by stabilization of the covalent topo III-DNA cleavage complex. Rather, BQ most probably inhibits the SH-dependent topo II by binding to an essential SH group. The inhibition of topo II by BQ has implications for the formation of deleterious translocations that may be involved in BZ-induced initiation of leukemogenesis.
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PMID:Inhibition of human DNA topoisomerase II by hydroquinone and p-benzoquinone, reactive metabolites of benzene. 911 3

p-Benzoquinone (p-BQ), a stable metabolite of the human carcinogen benzene, forms two-ring benzetheno exocyclic base adducts with C, A, and G bases in DNA. As a part of a project for studying the biological effect of the p-BQ adducts, we report here on the first biophysical characterization of oligodeoxyribonucleotide duplexes containing a single site-specific p-BQ-C, using thermal denaturation and circular dichroism (CD). We find that the thermal and thermodynamic stabilities of the control duplex are reduced by p-BQ-C. The Tm value decreases by 12.6 degrees C at the duplex concentration of 1.5 microM and the Delta G o by 10.2 kcal/mol. The latter was determined from the concentration dependence of the Tm values. The destabilization has little dependence on the nature of the opposite base. This reduction is higher than that of the single base mismatches studied (-4.9 to -7.9 kcal/mol) and is close to that observed with an adjacent double mismatch-containing duplex (-11.3 kcal/mol). The overall B-conformation of the duplex with a p-BQ-C is, however, only slightly altered, relative to the parent duplex, as shown by CD spectra. The p-BQ-C-containing duplex has been found recently to be a good substrate for the major human AP endonuclease as compared to an abasic site-containing duplex [Hang, B., et al. (1997) Biochemistry 36, 15411-15418]. We now find that the thermodynamic properties and the localized conformational changes of a p-BQ-C-containing duplex are apparently related to those reported for a duplex containing an abasic site.
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PMID:A single cyclic p-benzoquinone adduct can destabilize a DNA oligonucleotide duplex. 954 3

We report here that the newly synthesized DNA adduct, 1,N6-benzetheno-dA (pBQ-dA), in defined oligonucleotides [Chenna and Singer, Chem. Res. Toxicol., 8, 865-874], is a substrate for the major human AP endonuclease, HAP1, and the Escherichia coli AP endonucleases, exonuclease III and endonuclease IV. The mechanism of cleavage is identical to that reported previously for 3,N4-benzetheno-dC (pBQ-dC) and leads to a phosphodiester bond cleavage 5' to the adduct. There are, however, significant differences in the rate of cleavage of this adduct by these enzymes. The two bacterial AP endonucleases are both much more efficient than the human repair enzyme. In addition, using two random oligodeoxynucleotide sequences containing a single pBQ-dA, exonuclease III and endonuclease IV are similarly active, while HAP1 shows a distinct sequence preference of approximately 10-fold in efficiency of cleavage. The repair of this adduct by the three recombinant enzymes is further confirmed by using both active site mutant HAP1 proteins and by E.coli mutant strains lacking exonuclease III and/ or endonuclease IV. This sequence-dependent repair of pBQ-dA by HAP1 may play an important role in modulating benzene-induced carcinogenesis.
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PMID:Differential cleavage of oligonucleotides containing the benzene-derived adduct, 1,N6-benzetheno-dA, by the major human AP endonuclease HAP1 and Escherichia coli exonuclease III and endonuclease IV. 974 26

Two human carcinogens that have been extensively studied are vinyl chloride and benzene. The active metabolites used in this study are chloroacetaldehyde (CAA) and para-benzoquinone (pBQ). Each forms exocyclic adducts between the N1 and N6 of A, the N3 and N4 of C and the N1 and N2 of G. Only CAA has been found to form the N2,3 adduct of G. CAA and pBQ adducts differ structurally in size and in the number of added rings, pBQ adding two rings to the base, while etheno bases have a single five-membered ring. The mechanism of repair of these two types of adducts by human enzymes has been studied in our laboratory with defined oligodeoxynucleotides and a site-specific adduct. The etheno derivatives are repaired by DNA glycosylase activity; two mammalian glycosylases are responsible: alkylpurine-DNA-N-glycosylase (APNG) and mismatch-specific thymine-DNA glycosylase. The former repairs 1,N6-ethenoA (epsilon A) as rapidly as the original substrate, 3-methyladenine, while the latter repairs 3,N4-ethenoC (epsilon C) more efficiently than the G/T mismatch. Our finding that there are separate enzymes for epsilon A and epsilon C has been confirmed by the use of tissue extracts from an APNG knockout mouse. As pBQ is much less efficient than CAA in modifying bases, the biochemical studies required total synthesis of the nucleosides. Furthermore, the pBQ adduct-containing oligomers are cleaved, to various extents by a different class of enzyme: human and bacterial N-5'-alkylpurine (AP) endonucleases. The enzyme incises such oligomers 5' to the adduct and generates 3'-hydroxyl and 5'-phosphoryl termini but leaves the modified base on the 5'-terminus of the 3' cleavage fragment ('dangling base'). Using active-site mutants of the human AP endonuclease, we found that the active site for the primary substrate, abasic (AP) site, is the same as that for the bulky pBQ adducts. There appears to be no clear rationale for the widely differing recognition and repair mechanisms for these exocyclic adducts, as shown for the repair of the same types of modification on different bases (e.g. epsilon A and epsilon C) and for completely unrelated lesions (e.g. AP site and pBQ adducts). Another important variable that affects the rate and extent of repair is the effect of neighbouring bases. In the case of epsilon A, this sequence-dependent repair correlates with the extent of double-strandedness of the substrate, as demonstrated by thermal stability studies.
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PMID:Mammalian enzymatic repair of etheno and para-benzoquinone exocyclic adducts derived from the carcinogens vinyl chloride and benzene. 1062 24


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