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Uracil dna glycosylase

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https://www.readbyqxmd.com/read/29310268/integrating-dna-structure-switch-with-branched-hairpins-for-the-detection-of-uracil-dna-glycosylase-activity-and-inhibitor-screening
#1
Jing Zhu, Qijie Hao, Yi Liu, Zhaohui Guo, Buayxigul Rustam, Wei Jiang
The detection of uracil-DNA glycosylase (UDG) activity is pivotal for its biochemical studies and the development of drugs for UDG-related diseases. Here, we explored an integrated DNA structure switch for high sensitive detection of UDG activity. The DNA structure switch containing two branched hairpins was employed to recognize UDG enzyme and generate fluorescent signal. Under the action of UDG, one branched hairpin was impelled folding into a close conformation after the excision of the single uracil. This reconfigured hairpin could immediately initiate the polymerization/nicking amplification reaction of another branched hairpin accompanying with the release of numerous G-quadruplexes (G4s)...
March 1, 2018: Talanta
https://www.readbyqxmd.com/read/29178619/abasic-site-assisted-inhibition-of-nicking-endonuclease-activity-for-the-sensitive-determination-of-uracil-dna-glycosylase
#2
Jun Ki Ahn, Chang Yeol Lee, Ki Soo Park, Hyun Gyu Park
We herein describe a novel strategy to accurately determine uracil DNA glycosylase (UDG) activity based on the finding that nicking endonuclease-assisted cleavage reaction can be regulated by the presence of abasic site. This strategy utilizes DNA probes rationally designed to contain uracil base at the cleavage site for nicking endonuclease, which is coupled to the isothermal nicking endonuclease amplification reaction (NEAR) method. In the absence of UDG, intact DNA probes generate a large number of double-stranded (ds) DNA products through the NEAR, but the presence of UDG that converts uracil base into abasic site suppresses nicking endonuclease activity and the subsequent NEAR...
November 27, 2017: Biotechnology Journal
https://www.readbyqxmd.com/read/29171849/uracil-removal-inhibited-ligase-reaction-in-combination-with-catalytic-hairpin-assembly-for-the-sensitive-and-specific-detection-of-uracil-dna-glycosylase-activity
#3
Xiaowen Xu, Lei Wang, Yushu Wu, Wei Jiang
Sensitive and specific detection of uracil-DNA glycosylase (UDG) activity is crucial in biomedical study and disease diagnosis. Here, we developed a uracil removal-inhibited ligase reaction in combination with catalytic hairpin assembly (CHA) for the sensitive and specific detection of UDG activity. A hairpin probe is specially designed, which contains two uracil bases in the loop and is extended with toehold and branch-migration domains at the ends of the stem. Two short oligonucleotides are separately hybridized to one-half of the loop of the hairpin probe to form a DNA complex with a nick...
November 24, 2017: Analyst
https://www.readbyqxmd.com/read/29163805/thymine-dna-glycosylase-tdg-is-involved-in-the-pathogenesis-of-intestinal-tumors-with-reduced-apc-expression
#4
Jinfei Xu, Salvatore Cortellino, Rossella Tricarico, Wen-Chi Chang, Gabrielle Scher, Karthik Devarajan, Michael Slifker, Robert Moore, Maria Rosaria Bassi, Elena Caretti, Margie Clapper, Harry Cooper, Alfonso Bellacosa
Thymine DNA Glycosylase (TDG) is a base excision repair enzyme that acts as a thymine and uracil DNA N-glycosylase on G:T and G:U mismatches, thus protecting CpG sites in the genome from mutagenesis by deamination. In addition, TDG has an epigenomic function by removing the novel cytosine derivatives 5-formylcytosine and 5-carboxylcytosine (5caC) generated by Ten-Eleven Translocation (TET) enzymes during active DNA demethylation. We and others previously reported that TDG is essential for mammalian development...
October 27, 2017: Oncotarget
https://www.readbyqxmd.com/read/29137710/loop-mediated-isothermal-amplification-using-self-avoiding-molecular-recognition-systems-and-antarctic-thermal-sensitive-uracil-dna-glycosylase-for-detection-of-nucleic-acid-with-prevention-of-carryover-contamination
#5
Yi Wang, Dongxin Liu, Jianping Deng, Yan Wang, Jianguo Xu, Changyun Ye
Loop-mediated isothermal amplification (LAMP) is the most popular technique to amplify nucleic acid sequence without the use of temperature cycling. However, LAMP is often confounded by false-positive results, arising from interactions between (hetero-dimer) or within (self-dimerization) primers, off-target hybrids and carryover contaminants. Here, we devised a new LAMP technique that is self-avoiding molecular recognition system (SAMRS) components and antarctic thermal sensitive uracil-DNA-glycosylase (AUDG) enzyme-assisted, termed AUDG-SAMRS-LAMP...
December 15, 2017: Analytica Chimica Acta
https://www.readbyqxmd.com/read/29122935/in-vivo-measurements-of-interindividual-differences-in-dna-glycosylases-and-ape1-activities
#6
Isaac A Chaim, Zachary D Nagel, Jennifer J Jordan, Patrizia Mazzucato, Le P Ngo, Leona D Samson
The integrity of our DNA is challenged with at least 100,000 lesions per cell on a daily basis. Failure to repair DNA damage efficiently can lead to cancer, immunodeficiency, and neurodegenerative disease. Base excision repair (BER) recognizes and repairs minimally helix-distorting DNA base lesions induced by both endogenous and exogenous DNA damaging agents. Levels of BER-initiating DNA glycosylases can vary between individuals, suggesting that quantitating and understanding interindividual differences in DNA repair capacity (DRC) may enable us to predict and prevent disease in a personalized manner...
November 9, 2017: Proceedings of the National Academy of Sciences of the United States of America
https://www.readbyqxmd.com/read/29117941/loss-of-uracil-dna-glycosylase-selectively-re-sensitizes-p53-mutant-and-deficient-cells-to-5-fdu
#7
Yan Yan, Yulan Qing, John J Pink, Stanton L Gerson
Thymidylate synthase (TS) inhibitors including fluoropyrimidines [e.g., 5-Fluorouracil (5-FU) and 5-Fluorodeoxyuridine (5-FdU, floxuridine)] and antifolates (e.g., pemetrexed) are widely used against solid tumors. Previously, we reported that shRNA-mediated knockdown (KD) of uracil DNA glycosylase (UDG) sensitized cancer cells to 5-FdU. Since p53 has also been shown as a critical determinant of the sensitivity to TS inhibitors, we further interrogated 5-FdU cytotoxicity after UDG depletion with regard to p53 status...
November 8, 2017: Molecular Cancer Research: MCR
https://www.readbyqxmd.com/read/29099587/combining-h-d-exchange-mass-spectrometry-and-computational-docking-to-derive-the-structure-of-protein-protein-complexes
#8
Victoria A Roberts, Michael E Pique, Simon Hsu, Sheng Li
Protein-protein interactions are essential for biological function, but structures of protein-protein complexes are difficult to obtain experimentally. To derive the protein complex of the DNA-repair enzyme human uracil-DNA-glycosylase (hUNG) with its protein inhibitor (UGI), we combined rigid-body computational docking with hydrogen/deuterium exchange mass spectrometry (DXMS). Computational docking of the unbound protein structures provides a list of possible three-dimensional models of the complex; DXMS identifies solvent-protected protein residues...
November 16, 2017: Biochemistry
https://www.readbyqxmd.com/read/29079575/hiv-1-vpr-protein-directly-loads-helicase-like-transcription-factor-hltf-onto-the-crl4-dcaf1-e3-ubiquitin-ligase
#9
Xiaohong Zhou, Maria Delucia, Caili Hao, Kasia Hrecka, Christina Monnie, Jacek Skowronski, Jinwoo Ahn
The Vpr protein is an accessory virulence factor of HIV-1 that facilitates infection in immune cells. Cellular functions of Vpr are tied to its interaction with DCAF1, a substrate receptor component of the CRL4 E3 ubiquitin ligase. Recent proteomic approaches suggested that Vpr degrades HLTF DNA helicase in a proteasome-dependent manner by redirecting the CRL4-DCAF1 E3 ligase. However, the precise molecular mechanism of Vprdependent HLTF depletion is not known. Here, using in vitro reconstitution assays, we show that Vpr mediates polyubiquitination of HLTF, by directly loading it onto the C-terminal WD40 domain of DCAF1 in complex with the CRL4 E3 ubiquitin ligase...
October 27, 2017: Journal of Biological Chemistry
https://www.readbyqxmd.com/read/29051947/pre-steady-state-kinetic-analysis-of-damage-recognition-by-human-single-strand-selective-monofunctional-uracil-dna-glycosylase-smug1
#10
Alexandra A Kuznetsova, Danila A Iakovlev, Inna V Misovets, Alexander A Ishchenko, Murat K Saparbaev, Nikita A Kuznetsov, Olga S Fedorova
In all organisms, DNA glycosylases initiate base excision repair pathways resulting in removal of aberrant bases from DNA. Human SMUG1 belongs to the superfamily of uracil-DNA glycosylases catalyzing the hydrolysis of the N-glycosidic bond of uridine and uridine lesions bearing oxidized groups at C5: 5-hydroxymethyluridine (5hmU), 5-formyluridine (5fU), and 5-hydroxyuridine (5hoU). An apurinic/apyrimidinic (AP) site formed as the product of an N-glycosylase reaction is tightly bound to hSMUG1, thus inhibiting the downstream action of AP-endonuclease APE1...
November 21, 2017: Molecular BioSystems
https://www.readbyqxmd.com/read/29038818/a-c-hcr-assembly-of-branched-dna-nanostructures-for-amplified-uracil-dna-glycosylase-assays
#11
Jing Wang, Min Pan, Jie Wei, Xiaoqing Liu, Fuan Wang
An autonomous nonenzymatic DNA machine has been successfully engineered based on a two-layered cascaded hybridization chain reaction (C-HCR) circuit, in which the tandem outputs of the upstream HCR-1 unit activate the downstream HCR-2 unit to induce successive repeated hybridizations, generating branched DNA structures and enabling sensitive and selective detection of uracil-DNA glycosylase and its inhibitors.
October 17, 2017: Chemical Communications: Chem Comm
https://www.readbyqxmd.com/read/29037087/a-label-free-technique-for-accurate-detection-of-nucleic-acid-based-self-avoiding-molecular-recognition-systems-supplemented-multiple-cross-displacement-amplification-and-nanoparticles-based-biosensor
#12
Yi Wang, Yan Wang, Hong Wang, Jianguo Xu, Changyun Ye
Here, we devised a novel isothermal technique on the basis of standard multiple cross-displacement amplification (MCDA), which is assisted with self-avoiding molecular recognition system (SAMRS) components and antarctic thermal-sensitive uracil-DNA-glycosylase enzyme (AUDG), termed AUDG-SAMRS-MCDA. To enable product detection on the dipsticks, we firstly developed an analysis strategy, which did not require the labelled primers or probes, and thus, the analysis system avoids the false-positive results arising from undesired hybridization (between two labelled primers, or the labelled probe and primer)...
October 16, 2017: Artificial Cells, Nanomedicine, and Biotechnology
https://www.readbyqxmd.com/read/29036472/measurement-of-nanoscale-dna-translocation-by-uracil-dna-glycosylase-in-human-cells
#13
Alexandre Esadze, Gaddiel Rodriguez, Brian P Weiser, Philip A Cole, James T Stivers
DNA 'sliding' by human repair enzymes is considered to be important for DNA damage detection. Here, we transfected uracil-containing DNA duplexes into human cells and measured the probability that nuclear human uracil DNA glycosylase (hUNG2) excised two uracil lesions spaced 10-80 bp apart in a single encounter without escaping the micro-volume containing the target sites. The two-site transfer probabilities were 100% and 54% at a 10 and 40 bp spacing, but dropped to only 10% at 80 bp. Enzyme trapping experiments suggested that site transfers over 40 bp followed a DNA 'hopping' pathway in human cells, indicating that authentic sliding does not occur even over this short distance...
September 25, 2017: Nucleic Acids Research
https://www.readbyqxmd.com/read/29031631/most-probable-number-with-visual-based-lamp-for-the-quantification-of-reductive-dehalogenase-genes-in-groundwater-samples
#14
Yogendra H Kanitkar, Robert D Stedtfeld, Paul B Hatzinger, Syed A Hashsham, Alison M Cupples
The remediation of chlorinated solvent contaminated sites frequently involves bioaugmentation with mixed cultures containing Dehalococcoides mccartyi. Their activity is then examined by quantifying reductive dehalogenase (RDase) genes. Recently, we described a rapid, low cost approach, based on loop mediated isothermal amplification (LAMP), which allowed for the visual detection of RDase genes from groundwater. In that study, samples were concentrated (without DNA extraction), incubated in a water bath (avoiding the use of a thermal cycler) and amplification was visualized by the addition of SYBR green (post incubation)...
October 12, 2017: Journal of Microbiological Methods
https://www.readbyqxmd.com/read/29031294/a-label-free-and-highly-sensitive-strategy-for-uracil-dna-glycosylase-activity-detection-based-on-stem-loop-primer-mediated-exponential-amplification-spea
#15
Wenfang Du, Junjie Li, Fubing Xiao, Ruqin Yu, Jianhui Jiang
Uracil-DNA glycosylase (UDG) plays essential roles in base excision repair (BER) pathway by eliminating uracil from DNA to sustain the genome integrity. Sensitive detection of UDG activity is of great significance in the study of many fundamental biochemical processes and clinical applications. We develop a label-free method for UDG activity detection using stem-loop primer-mediated exponential amplification (SPEA). In the presence of active UDG, the uracil base in helper hairpin probe (HP) can be excised to generate an abasic site (AP site), which can be cleaved by endonuclease IV (Endo IV) with a blocked primer released...
October 23, 2017: Analytica Chimica Acta
https://www.readbyqxmd.com/read/28977725/an-unconventional-family-1-uracil-dna-glycosylase-in-nitratifractor%C3%A2-salsuginis
#16
Jing Li, Ran Chen, Ye Yang, Zhemin Zhang, Guang-Chen Fang, Wei Xie, Weiguo Cao
The uracil DNA glycosylase superfamily consists of at least six families with a diverse specificity toward DNA base damage. Family 1 uracil N-glycosylase (UNG) exhibits exclusive specificity on uracil-containing DNA. Here, we report a family 1 UNG homolog from Nitratifractor salsuginis with distinct biochemical features that differentiate it from conventional family 1 UNGs. Globally, the crystal structure of N. salsuginisUNG shows a few additional secondary structural elements. Biochemical and enzyme kinetic analysis, coupled with structural determination, molecular modeling, and molecular dynamics simulations, shows that N...
October 4, 2017: FEBS Journal
https://www.readbyqxmd.com/read/28955788/biochemical-characterization-and-novel-inhibitor-identification-of-mycobacterium-tuberculosis-endonuclease-viii-2-rv3297
#17
Kiran Lata, Mohammad Afsar, Ravishankar Ramachandran
Nei2 (Rv3297) is a DNA Base Excision Repair (BER) glycosylase that is essential for survival of Mycobacterium tuberculosis in primates. We show that MtbNei2 is a bifunctional glycosylase that specifically acts on oxidized pyrimidine-containing single-stranded, double-stranded, 5'/3' fork and bubble DNA substrates. MtbNei2 possesses Uracil DNA glycosylase activity unlike E. coli Nei. Mutational studies demonstrate that Pro2 and Glu3 located in the active site are essential for glycosylase activity of MtbNei2...
December 2017: Biochemistry and Biophysics Reports
https://www.readbyqxmd.com/read/28940147/backbone-resonance-assignment-of-the-human-uracil-dna-glycosylase-2
#18
Hesna Kara, Luc Ponchon, Serge Bouaziz
The HIV-1 viral protein R (Vpr) is incorporated into virus particle during budding suggesting that its presence in the mature virion is required in the early steps of the virus life cycle in newly infected cells. Vpr is released into the host cell cytoplasm to participate to the translocation of the preintegration complex (PIC) into the nucleus for integration of the viral DNA into the host genome. Actually, Vpr plays a key role in the activation of the transcription of the HIV-1 long terminal repeat (LTR), mediates cell cycle arrest in G2 to M transition, facilitates apoptosis and controls the fidelity of reverse transcription...
September 22, 2017: Biomolecular NMR Assignments
https://www.readbyqxmd.com/read/28879561/backbone-1-h-13-c-and-15-n-chemical-shift-assignment-of-full-length-human-uracil-dna-glycosylase-ung2
#19
Edith Buchinger, Siv Å Wiik, Anna Kusnierczyk, Renana Rabe, Per A Aas, Bodil Kavli, Geir Slupphaug, Finn L Aachmann
Human uracil N-glycosylase isoform 2-UNG2 consists of an N-terminal intrinsically disordered regulatory domain (UNG2 residues 1-92, 9.3 kDa) and a C-terminal structured catalytic domain (UNG2 residues 93-313, 25.1 kDa). Here, we report the backbone (1)H, (13)C, and (15)N chemical shift assignment as well as secondary structure analysis of the N-and C-terminal domains of UNG2 representing the full-length UNG2 protein.
September 6, 2017: Biomolecular NMR Assignments
https://www.readbyqxmd.com/read/28875174/improved-base-excision-repair-inhibition-and-bacteriophage-mu-gam-protein-yields-c-g-to-t-a-base-editors-with-higher-efficiency-and-product-purity
#20
Alexis C Komor, Kevin T Zhao, Michael S Packer, Nicole M Gaudelli, Amanda L Waterbury, Luke W Koblan, Y Bill Kim, Ahmed H Badran, David R Liu
We recently developed base editing, the programmable conversion of target C:G base pairs to T:A without inducing double-stranded DNA breaks (DSBs) or requiring homology-directed repair using engineered fusions of Cas9 variants and cytidine deaminases. Over the past year, the third-generation base editor (BE3) and related technologies have been successfully used by many researchers in a wide range of organisms. The product distribution of base editing-the frequency with which the target C:G is converted to mixtures of undesired by-products, along with the desired T:A product-varies in a target site-dependent manner...
August 2017: Science Advances
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