The major Arabidopsis thaliana apurinic/apyrimidinic endonuclease, ARP is involved in the plant nucleotide incision repair pathway

Akishev, Zhiger; Taipakova, Sabira; Joldybayeva, Botagoz; Zutterling, Caroline; Smekenov, Izat; Ищенко, А. А.; Zharkov, Dmitry O.; Bissenbaev, Amangeldy K.; Saparbaev, Murat

doi:10.1016/j.dnarep.2016.10.009

Cited by 22 publications

(19 citation statements)

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“…It should be mentioned that the activity of APE1 depends on the concentration of Mg 2+ [ 14 , 34 ]. Typically, the AP-site cleavage activity is tested in the presence of ≥5 mM Mg 2+ [ 14 , 35 , 36 , 37 , 38 ], but the cleavage of DNA bearing a damaged base is tested at a low concentration of Mg 2+ , within the 0.01–0.5 mM range [ 12 , 14 , 35 , 36 , 39 ]. Consequently, to verify the efficiency of APE1 binding to the DHU-containing DNA duplex at various Mg 2+ levels, the kinetics of substrate–enzyme complex formation and of the catalysis were investigated by the stopped-flow method ( Figure 1 a).…”

Section: Resultsmentioning

confidence: 99%

The Role of Active-Site Plasticity in Damaged-Nucleotide Recognition by Human Apurinic/Apyrimidinic Endonuclease APE1

et al. 2020

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Human apurinic/apyrimidinic (AP) endonuclease APE1 hydrolyzes phosphodiester bonds on the 5′ side of an AP-site, and some damaged nucleotides such as 1,N6-ethenoadenosine (εA), α-adenosine (αA), and 5,6-dihydrouridine (DHU). To investigate the mechanism behind the broad substrate specificity of APE1, we analyzed pre-steady-state kinetics of conformational changes in DNA and the enzyme during DNA binding and damage recognition. Molecular dynamics simulations of APE1 complexes with one of damaged DNA duplexes containing εA, αA, DHU, or an F-site (a stable analog of an AP-site) revealed the involvement of residues Asn229, Thr233, and Glu236 in the mechanism of DNA lesion recognition. The results suggested that processing of an AP-site proceeds faster in comparison with nucleotide incision repair substrates because eversion of a small abasic site and its insertion into the active site do not include any unfavorable interactions, whereas the insertion of any target nucleotide containing a damaged base into the APE1 active site is sterically hindered. Destabilization of the α-helix containing Thr233 and Glu236 via a loss of the interaction between these residues increased the plasticity of the damaged-nucleotide binding pocket and the ability to accommodate structurally different damaged nucleotides. Nonetheless, the optimal location of εA or αA in the binding pocket does not correspond to the optimal conformation of catalytic amino acid residues, thereby significantly decreasing the cleavage efficacy for these substrates.

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Section: Resultsmentioning

confidence: 99%

The Role of Active-Site Plasticity in Damaged-Nucleotide Recognition by Human Apurinic/Apyrimidinic Endonuclease APE1

et al. 2020

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“…In addition to the AP site and 3′-end processing activities, certain EEP superfamily enzymes possess the NIR activity that nicks DNA at some sites containing damaged bases. Notably, the NIR activity is distributed unevenly in the phylogeny: it has been reported for human APE1 [ 13 ], Arabidopsis ARP [ 33 ], and Methanothermobacter Mth212 [ 60 ], but is lacking or negligible in ExoIII-like enzymes [ 13 , 26 ], bacterial/archaeal Ape1-like enzymes [[ 49 ] and this work] and plant Ape1L [ 35 ]. Ape2-like enzymes, in view of their low AP endonuclease activity and the predominant 3’→5’-exonuclease activity [ 61 ], are also likely to be free of the NIR activity.…”

Section: Discussionmentioning

confidence: 95%

“…However, Ape1-like enzymes vary at this position, containing Asp, Asn, or small amino acids (Gly/Ala). Despite influencing the metal preference and/or the balance of endonuclease, exonuclease, and 3’-phosphodiesterase activities [ 33 ], both Asn and Asp are likely suitable for the biologically relevant function of EEP superfamily AP endonucleases.…”

Section: Resultsmentioning

confidence: 99%

“…They included the 30mer X-RT where X is tetrahydrofuran (THF, a synthetic analog of an abasic site), α-2’-deoxyadenosine (αdA), 5-hydroxycytosine (5ohC), or 5,6-dihydrouracil (DHU), and complementary 30mer oligonucleotides containing dA, dG, dC, or T opposite the lesion. DNA substrates were constructed as described previously [ 26 , 33 ]. Briefly, the complementary oligonucleotides were annealed; the resulting duplex oligonucleotides are referred to as X•C (G, A, T), respectively, where X is a modified residue.…”

Section: Methodsmentioning

confidence: 99%

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Characterization of biochemical properties of an apurinic/apyrimidinic endonuclease from Helicobacter pylori

et al. 2018

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Apurinic/apyrimidinic (AP) endonucleases play critical roles in the repair of abasic sites and strand breaks in DNA. Complete genome sequences of Helicobacter pylori reveal that this bacterial specie has a single AP endonuclease. An H. pylori homolog of Xth (HpXth) is a member of exonuclease III family, which is represented by Escherichia coli Xth. Currently, it remains unknown whether this single AP endonuclease has DNA repair activities similar to those of its counterpart in E. coli and other bacteria. We report that HpXth possesses efficient AP site cleavage, 3’-repair phosphodiesterase, and 3’-phosphatase activities but not the nucleotide incision repair function. Optimal reaction conditions for HpXth’s AP endonuclease activity are low ionic strength, high Mg2+ concentration, pH in the range 7–8, and temperature 30 °C. The kinetic parameters measured under steady-state conditions showed that HpXth removes the AP site, 3’-blocking sugar-phosphate, and 3’-terminal phosphate in DNA strand breaks with good efficiency (kcat/KM = 1240, 44, and 5,4 μM–1·min–1, respectively), similar to that of E. coli Xth. As expected, the presence of HpXth protein in AP endonuclease—deficient E. coli xth nfo strain significantly reduced the sensitivity to an alkylating agent and H2O2. Mutation of active site residue D144 in HpXth predicted to be essential for catalysis resulted in a complete loss of enzyme activities. Several important structural features of HpXth were uncovered by homology modeling and phylogenetic analysis. Our data show the DNA substrate specificity of H. pylori AP endonuclease and suggest that HpXth counteracts the genotoxic effects of DNA damage generated by endogenous and host-imposed factors.

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“…For detection of APE2's capability to release free phosphate residue, DNA substrate with 3′-32 P terminus was generated according to a previously described method (Akishev et al, 2016). Briefly, a 25-nucleotide primer was 3′-end labeled using 3′-α[ 32 P]-dATP (cordycepin 5′-triphosphate; Perkin-Elmer) to generate a 26-nucleotide up-strand primer, which was then treated with recombinant human tyrosyl-DNA phosphodiesterase 1 (Tdp1).…”

Section: Enzymatic Assaysmentioning

confidence: 99%

APURINIC/APYRIMIDINIC ENDONUCLEASE2 and ZINC FINGER DNA 3′-PHOSPHOESTERASE Play Overlapping Roles in the Maintenance of Epigenome and Genome Stability

Liang

et al. 2018

Plant Cell

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Base excision repair (BER) is essential for active DNA demethylation and DNA damage repair in mammals and plants. Here, we provide genetic and biochemical evidence that APURINIC/APYRIMIDINIC ENDONUCLEASE2 (APE2) plays overlapping roles with ZINC FINGER DNA 3′-PHOSPHOESTERASE (ZDP) in active DNA demethylation and DNA damage repair in Arabidopsis thaliana. Simultaneous mutation of APE2 and ZDP causes DNA hypermethylation at more than 2000 loci, most of which are not hypermethylated in ape2 or zdp single mutants. The zdp and ape2 single mutants exhibit normal development, but the zdp ape2 double mutants display pleiotropic developmental defects and are supersensitive to the DNA alkylating reagent methyl methanesulfonate. The gradual accumulation of DNA lesions in the zdp ape2 seedlings is accompanied by constitutive activation of the DNA damage response and alteration of the cell cycle. Interestingly, knockout of the key DNA demethylase REPRESSOR OF SILENCING1 reduces the magnitude of DNA lesion accumulation and the DNA damage response in the zdp ape2 mutants, suggesting that a proportion of the DNA damage in the zdp ape2 mutants arises from incomplete active DNA demethylation. Lastly, we find that APE2 has 3′-phosphatase activity and strong 3′-5′ exonuclease activity in vitro. Together, our results suggest that APE2 and ZDP, two BER proteins, play overlapping roles in the maintenance of epigenome and genome stability in plants.

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The major Arabidopsis thaliana apurinic/apyrimidinic endonuclease, ARP is involved in the plant nucleotide incision repair pathway

Cited by 22 publications

References 59 publications

The Role of Active-Site Plasticity in Damaged-Nucleotide Recognition by Human Apurinic/Apyrimidinic Endonuclease APE1

The Role of Active-Site Plasticity in Damaged-Nucleotide Recognition by Human Apurinic/Apyrimidinic Endonuclease APE1

Characterization of biochemical properties of an apurinic/apyrimidinic endonuclease from Helicobacter pylori

APURINIC/APYRIMIDINIC ENDONUCLEASE2 and ZINC FINGER DNA 3′-PHOSPHOESTERASE Play Overlapping Roles in the Maintenance of Epigenome and Genome Stability

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