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 8-oxoguanine-DNA glycosylase (OGG1) efficiently removes mutagenic 8-oxo-7,8-dihydroguanine (8-oxoGua) and 2,6-diamino-4-hydroxy-5-formamidopyrimidine when paired with cytosine in oxidatively damaged DNA. Excision of 8-oxoGua mispaired with adenine may lead to G-->T transversions. Post-translational modifications such as phosphorylation could affect the cellular distribution and enzymatic activity of OGG1. Mutations and polymorphisms of OGG1 may affect the enzymatic activity and have been associated with increased risk of several cancers. In this study, we used double-stranded oligodeoxynucleotides containing 8-oxoGua:Cyt or 8-oxoGua:Ade pairs, as well as gamma-irradiated calf thymus DNA, to investigate the kinetics and substrate specificity of several known OGG1 polymorphic variants and phosphomimetic Ser-->Glu mutants. Among the polymorphic variants, A288V and S326C displayed opposite-base specificity similar to that of wild-type OGG1, whereas OGG1-D322N was 2.3-fold more specific for the correct opposite base than the wild-type enzyme. All phosphomimetic mutants displayed approximately 1.5-3-fold lower ability to remove 8-oxoGua in both assays, whereas the substrate specificity of the phosphomimetic mutants was similar to that of the wild-type enzyme. OGG1-S326C efficiently excised 8-oxoGua from oligodeoxynucleotides and 2,6-diamino-4-hydroxy-5-formamidopyrimidine from gamma-irradiated DNA, but excised 8-oxoG rather inefficiently from gamma-irradiated DNA. Otherwise, kcat values for 8-oxoGua excision obtained from both types of experiments were similar for all OGG1 variants studied. It is known that the human AP endonuclease APEX1 can stimulate OGG1 activity by increasing its turnover rate. However, when wild-type OGG1 was replaced by one of the phosphomimetic mutants, very little stimulation of 8-oxoGua removal was observed in the presence of APEX1.
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PMID:Substrate specificity and excision kinetics of natural polymorphic variants and phosphomimetic mutants of human 8-oxoguanine-DNA glycosylase. 1967 7

Oxidative DNA damage has been implicated in a number of central nervous system pathologies. The base excision repair (BER) pathway is one of the most important cellular protection mechanisms that respond to oxidative DNA damage. Human apurinic (apyrimidinic) endonuclease/redox effector factor (APE1/Ref-1 or APE1) is an essential enzyme in the BER pathway and is expressed in both mitotic and post-mitotic cells in humans. In neurons, a reduction of APE1 expression increases chemotherapy-induced cytotoxicity, while overexpression of APE1 protects cells against the cytotoxicity. However, given the multiple functions of APE1, knockdown of total APE1 is not completely informative of whether it is the redox or DNA repair activity, or interactions with other proteins. Therefore, the use of selective small molecules that can block each function independent of the other is of great benefit in ascertaining APE1 function in post-mitotic cells. In this study, we chose differentiated SH-SY5Y cells as our post-mitotic cell line model to investigate whether a drug-induced decrease in APE1 DNA repair or redox activity contributes to the growth and survival of post-mitotic cells under oxidative DNA damaging conditions. Here, we demonstrate that overexpression of WT-APE1 or C65-APE1 (repair competent) results in significant increase in cell viability after exposure to H(2)O(2). However, the 177/226-APE1 (repair deficient) did not show a protective effect. This phenomenon was further confirmed by the use of methoxyamine (MX), which blocks the repair activity of APE1 that results in enhanced cell killing and apoptosis in differentiated SH-SY5Y cells and in neuronal cultures after oxidative DNA damaging treatments. Blocking APE1 redox function by a small molecule inhibitor, BQP did not decrease viability of SH-SY5Y cells or neuronal cultures following oxidative DNA damaging treatments. Our results demonstrate that the DNA repair function of APE1 contributes to the survival of nondividing post-mitotic cells following oxidative DNA damage.
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PMID:Role of APE1 in differentiated neuroblastoma SH-SY5Y cells in response to oxidative stress: use of APE1 small molecule inhibitors to delineate APE1 functions. 1972 41

A new assay for measuring glycosylase activity of human 8-oxoguanine-DNA glycosylase is described. The assay measures the amount of released 8-oxoguanine from synthetic oligonucleotides containing modified base in the middle of the sequence. After enzymatic release, the amount of base is quantified by liquid chromatography-mass spectrometry. Chromatographic separation is carried out on a reversed-phase C(18) column using 10% methanol/water. Quantitation of 8-oxoguanine is carried out by negative-ion electrospray on a single quadrupole mass spectrometer operated in selected-ion monitoring mode. The limit of quantitation was 6 nM and the assay was linear from 6 to 1000 nM. The method was evaluated by monitoring the kinetics of base excision of several substrates as well as by measuring stimulation of activity in the presence of APE1 endonuclease. The new assay provides much higher throughput compared to traditional gel-based assays, which is particularly important when large number of samples need to analyzed.
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PMID:A liquid chromatography-mass spectrometric assay for measuring activity of human 8-oxoguanine-DNA glycosylase. 1978 38

Resistance to platinum is a major limitation for the treatment of ovarian cancer. In an effort to overcome the platinum resistance problem in ovarian cancer treatment, we explored the correlation between cisplatin resistance and the human AP endonuclease (APE1 or Ref-1). APE1/Ref-1 is a multifunctional protein that is not only an essential enzyme in base excision repair pathway, but also acts as a major redox-signaling factor that has a wide variety of important cellular functions including transcription factor regulation, oxidative signaling and cell cycle control. In this study, we examined APE1/Ref-1 expression by immunohistochemistry in sections of ovarian cancers from 78 patients who were administered standard adjuvant chemotherapy based on platinum post-operatively. Altered levels and subcellular APE1/Ref-1 expression was found in patients not responding to platinum-based chemotherapy comparing with those who responded to platinum-based chemotherapy. Meanwhile, we detected the APE1/Ref-1 expression in A2780 and CP70 cell lines which have different sensitivity to cisplatin. We found similar altered APE1/Ref-1 expression in them. We hypothesized that the APE1/Ref-1 expression is responsible in part for the cisplatin resistance. To answer this hypothesis, we decreased the APE1/Ref-1 level by silencing RNA targeting technology in A2780 and CP70 cell lines. The A2780 cells treated with APE1-siRNA had IC50 values ranging from 6.70 to 1.74 microM cisplatin compared with 15.81 microM for control A2780 cells. The CP70 cells treated with APE1-siRNA had 1.62-4.63-fold enhancement in cisplatin sensitivity. The apoptosis assays using TUNEL analysis showed that decreased APE1/Ref-1 level resulted in increased apoptosis levels in A2780 and CP70 cell lines compared with the control-treated cells. These data suggest that APE1/Ref-1 levels play an important role in the sensitization of ovarian cancer cells to apoptosis. In vitro studies revealed that it is possible to substantially enhance the cisplatin cytotoxicity by decreasing APE1/Ref-1 level in cisplatin-resistant cell lines.
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PMID:Alterations in the expression of the apurinic/apyrimidinic endonuclease-1/redox factor-1 (APE1/Ref-1) in human ovarian cancer and indentification of the therapeutic potential of APE1/Ref-1 inhibitor. 1978 61

The induction of mutations in mammalian cells exposed to cadmium has been associated with the oxidative stress triggered by the metal. There is increasing evidence that the mutagenic potential of Cd is not restricted to the induction of DNA lesions. Cd has been shown to inactivate several DNA repair enzymes. Here we show that exposure of human cells to sub-lethal concentrations of Cd leads to a time- and concentration-dependent decrease in hOGG1 activity, the major DNA glycosylase activity responsible for the initiation of the base excision repair (BER) of 8-oxoguanine, an abundant and mutagenic form of oxidized guanine. Although there is a slight effect on the level of hOGG1 transcripts, we show that the inhibition of the 8-oxoguanine DNA glycosylase activity is mainly associated with an oxidation of the hOGG1 protein and its disappearance from the soluble fraction of total cell extracts. Confocal microscopy analyses show that in cells exposed to Cd hOGG1-GFP is recruited to discrete structures in the cytoplasm. These structures were identified as stress granules. Removal of Cd from the medium allows the recovery of the DNA glycosylase activity and the presence of hOGG1 in a soluble form. In contrast to hOGG1, we show here that exposure to Cd does not affect the activity of the second enzyme of the pathway, the major AP endonuclease APE1.
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PMID:Inactivation by oxidation and recruitment into stress granules of hOGG1 but not APE1 in human cells exposed to sub-lethal concentrations of cadmium. 1980 Aug 94

Apurinic/apyrimidinic endonuclease 1 (APE1 or Ref-1) is the major enzyme in mammals for processing abasic sites in DNA. These cytotoxic and mutagenic lesions arise via spontaneous rupture of the base-sugar bond or the removal of damaged bases by a DNA glycosylase. APE1 cleaves the DNA backbone 5' to an abasic site, giving a 3'-OH primer for repair synthesis, and mediates other key repair activities. The DNA repair functions are essential for embryogenesis and cell viability. APE1-deficient cells are hypersensitive to DNA-damaging agents, and APE1 is considered an attractive target for inhibitors that could potentially enhance the efficacy of some anti-cancer agents. To enable an important new method for studying the structure, dynamics, catalytic mechanism, and inhibition of APE1, we assigned the chemical shifts (backbone and (13)C(beta)) of APE1 residues 39-318. We also report a protocol for refolding APE1, which was essential for achieving complete exchange of backbone amide sites for the perdeuterated protein.
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PMID:Chemical shift assignments for human apurinic/apyrimidinic endonuclease 1. 1988 78

Redox factor-1 (Ref-1), also known as HAP1, APE or APEX, is a multifunctional protein that regulates gene transcription as well as the response to oxidative stress. By interacting with transcription factors such as AP-1, NF-kappaB and p53, and directly participating in the cleavage of apurininic/apyrimidinic DNA lesions, Ref-1 plays crucial roles in both cell death signaling pathways and DNA repair, respectively. Oxidative stress induced by aggregated beta-amyloid (Abeta) peptide, altered DNA repair and transcriptional activation of cell death pathways have been implicated in the pathophysiology of Alzheimer's disease (AD). Here we show that varying concentrations of Abeta(1-42) differentially regulate Ref-1 expression, Ref-1 function and neuronal survival in vitro. Abeta (5.0 muM) caused a relatively rapid decrease in Ref-1 expression and activity associated with extensive DNA damage and neuronal degeneration. In contrast, Ref-1 induction occurred in cells exposed to Abeta (1.0 muM) without significant neuronal cell death. Abeta-induced attenuation of Ref-1 expression and endonuclease activity, and neuronal cell death were prevented by the anti-oxidant, catalase. Similar differential effects on Ref-1 expression and cell viability were observed in N2A neuroblastoma cells treated with either high or low dose hydrogen peroxide. These findings demonstrate the differential regulation of Ref-1 expression by varying degrees of oxidative stress. Parallels between the Ref-1 response to Abeta and H(2)O(2) suggest similarities between DNA repair pathways activated by different inducers of oxidative stress. In AD brain, colocalization of Ref-1 and Abeta the absence of significant DNA damage are consistent with the cell culture results and suggests that Ref-1 may play a more neuroprotective role under these conditions. Modulation of Ref-1 expression and activity by local variations in Abeta concentration may be an important determinant of neuronal vulnerability to oxidative stress in AD.
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PMID:Differential Expression of Redox Factor-1 Associated with Beta-Amyloid-Mediated Neurotoxicity. 1989 78

Apurinic/apyrimidinic endonuclease/redox factor-1 (APE/Ref-1), as a type of multifunctional protein, plays an essential role in the base excision repair (BER) pathway, which is responsible for the repair of DNA caused by oxidative and alkylation damage. As importantly, APE/Ref-1 also functions as a redox factor maintaining transcription factors in an active reduced state. APE/Ref-1 stimulates the DNA-binding activity of numerous transcription factors that are involved in cancer promotion and progression, such as AP-1 (Fos/Jun), NF-kappaB, HIF-1alpha, p53, and others. Based on the structures and functions of APE1/Ref-1, we will provide an overview of its activities and explore the budding clinical use of this protein as a target in cancer treatment, and propose that APE/Ref-1 has a great potential for application in clinical research.
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PMID:Anticancer clinical utility of the apurinic/apyrimidinic endonuclease/redox factor-1 (APE/Ref-1). 2019 21

Apurinic apyrimidinic endonuclease (APE1)/redox effector factor 1 (Ref-1), which is a multifunction protein involved in both transcriptional regulation of gene expression during adaptive cellular responses to oxidative stress and in the base excision repair pathway of DNA lesions generated as a consequence of oxidant-induced base damage, contributes to the maintenance of genome stability. APE1/Ref-1 is normally localized in the nucleus; cytoplasmic localization observed in several tumors has been correlated with a poor prognosis. Hepatocellular carcinoma (HCC) grading is an essential tool to predict the risk of relapse and patient prognosis, particularly in patients undergoing liver transplantation (OLT). The aim of this study was to identify the role of APE1/Ref-1 in predicting a posttransplant HCC relapse. We studied 48 patients transplanted for HCC to define grading as well as nuclear and cytoplasmic APE1/Ref-1 expression within neoplastic versus nonneoplastic parenchyma. We defined a cutoff of 60% of cytoplasmic APE1/Ref-1 expression to identify positive cases. At a minimum of 1.5-year follow-up after transplantation, 32 patients are alive and 16 patients are deceased after HCC relapse. Among low-grade HCC (grades 1 and 2), 76% of cases are alive; only 34% showed cytoplasmic APE1/Ref-1 immunoreactivity. Among the high-grade cases (grades 3 and 4), 50% were alive with 64% showing cytoplasmic immunoreactivity. Nuclear reactivity was generally similar either in neoplastic or in cirrhotic livers, irrespective of the grade. These data seemed to support the hypothesis of a predictive role of APE1/Ref-1 for HCC risk of relapse, which together with tumor grade by analysis of a pretransplant needle biopsy should aid decision making for OLT.
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PMID:Apurinic apyrimidinic endonuclease/redox effector factor 1 immunoreactivity and grading in hepatocellular carcinoma risk of relapse after liver transplantation. 2053 62

The exact biological function of granzyme A, a granule-associated serine protease belonging to the tryptase family of proteases, is still a matter of debate because conflicting roles have been suggested, such as initiation of caspase-independent apoptosis-like cell death and endogenous modulation of inflammatory processes. In contrast to its well-studied family member, granzyme B, far less is known about the physiological targets of granzyme A. Using an N-terminal peptide-centric proteomics technology, the substrate specificity of human granzyme A was extensively characterized at the level of macromolecular protein substrates. Overall, more than 260 cleavage sites, almost exclusively favoring basic residues at the P1 position, in approximately 200 unique protein substrates, including the well-known in vitro substrates APEX-endonuclease 1 and different histones, were identified. Further substrate characterization was used to delineate physical properties in the substrate specificity profiles, which further highlights important aspects in protease/substrate biology.
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PMID:The substrate specificity profile of human granzyme A. 2053 82


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