The Experts below are selected from a list of 4161 Experts worldwide ranked by ideXlab platform
Olga I. Lavrik - One of the best experts on this subject based on the ideXlab platform.
-
Quantitative parameters of the 3′–5′ exonuclease reaction of human APurinic/APyrimidinic endonuclease 1 with nicked DNA containing dYMP or a modified dCMP analogue
Russian Journal of Bioorganic Chemistry, 2020Co-Authors: N S Dyrkheeva, Svetlana N. Khodyreva, Olga I. LavrikAbstract:Human APurinic/APyrimidinic (AP) endonuclease 1 (APE1) is a multifunctional enzyme. In addition to its main AP endonuclease activity, that incises DNA 5′ to the AP-Site, it possesses other weak enzymatic activities. One of them is 3′–5′ exonuclease activity, which is most effectively exhibited for DNA duplexes containing modified or mismatched nucleotides at the 3′-end of the primer chain. There is a presumption that APE1 can correct the DNA synthesis catalyzed by DNA polymerase β through the base excision repair process. We determined the quantitative parameters of the 3′–5′ exonuclease reaction in dependence on the reaction conditions to reveal the detailed mechanism of this process. The kinetic parameters of APE1 exonuclease excision of mismatched dCMP and dTMP from the 3′ terminus of single-strand DNA and of photoreactive dCMP analogues APplied for photoaffinity modification of proteins and DNA in recombinant systems and cell/nuclear extracts were determined.
-
interaction of parp 2 with AP Site containing dna
Biochimie, 2015Co-Authors: M M Kutuzov, Svetlana N. Khodyreva, Ekaterina S Ilina, Maria V Sukhanova, Olga I. LavrikAbstract:Abstract In eukaryotes the stability of genome is provided by functioning of DNA repair systems. One of the main DNA repair pathways in eukaryotes is the base excision repair (BER). This system requires precise regulation for correct functioning. Two members of the PARP family – PARP-1 and PARP-2, which can be activated by DNA damage – are widely considered as regulators of DNA repair processes, including BER. In contrast to PARP-1, the role of PARP-2 in BER has not been extensively studied yet. Since AP Site is one of the most frequent type of DNA damage and a key intermediate of BER at the stage preceding formation of DNA breaks, in this pAPer we focused on the characterization of PARP-2 interaction with AP Site-containing DNAs. We demonstrated that PARP-2, like PARP-1, can interact with the intact AP Site via Schiff base formation, in spite of crucial difference in the structure of the DNA binding domains of these PARPs. By cross-linking of PARPs to AP DNA, we determined that the N-terminal domains of both PARPs are involved in formation of cross-links with AP DNA. We have also confirmed that DNA binding by PARP-2, in contrast to PARP-1, is not modulated by autoPARylation. PARP-2, like PARP-1, can inhibit the activity of APE1 by binding to AP Site, but, in contrast to PARP-1, this inhibitory influence is hardly regulated by PAR synthesis. At the same time, 5′-dRP lyase activity of both PARPs is comparable, although being much weaker than that of Pol β, which is considered as the main 5′-dRP lyase of the BER process.
-
Effect of multifunctional protein YB-1 on the AP Site cleavage by AP endonuclease 1 and tyrosyl phosphodiesterase 1
Biopolymers & Cell, 2012Co-Authors: P E Pestryakov, Svetlana N. Khodyreva, P. A. Curmi, L. P. Ovchinnikov, Olga I. LavrikAbstract:APurinic/APyrimidinic Sites (AP Sites) which represent one of the most abundantly generated DNA lesions in the cell are generally repaired by base excision repair (BER) pathway. Multifunctional protein YB-1 is known to participate in cellular response to genotoxic stress and was shown to interact with several components of BER – DNA glycosylases NTH1, NEIL2, DNA polymerase and DNA ligase III. Therefore, it is of great interest to investigate the influence of YB-1 on one of the major BER enzymes, responsible for AP Site cleavage, AP endonuclease APE1, and on tyrosyl phosphodiesterase Tdp1, participating in APE1 independent pathway of AP Site repair. Aim. Effect of multifunctional protein YB-1 on the AP Site cleavage by the activities of APE1 and Tdp1 was studied. Methods. Gel-mobility shift assays and enzyme activity tests. Results. YB-1 was shown to inhibit the cleavage of AP Site located in single-stranded DNA by both APE1 and Tdp1. Stimulation of APE1 activity on protruding double-stranded DNA in the presence of YB-1 was observed, whereas no effect on Tdp1-mediated cleavage of AP Site in double-stranded DNA was found. Conclusions. YB-1 can modulate the repair of AP Sites in DNA by both positively stimulating APE1 during the classic BER of AP Sites and avoiding a possible generation of doublestrand breaks, arising from the cleavage of single-stranded portion of DNA substrate already used by different DNA-processing pathwa
-
Tyrosyl-DNA phosphodiesterase 1 initiates repair of APurinic/APyrimidinic Sites.
Biochimie, 2012Co-Authors: N A Lebedeva, Nadejda I Rechkunova, Sherif F. El-khamisy, Olga I. LavrikAbstract:Tyrosyl-DNA phosphodiesterase 1 (Tdp1) catalyzes the hydrolysis of the phosphodiester linkage between the DNA 3′ phosphate and a tyrosine residue as well as a variety of other DNA 3′ damaged termini. Recently we have shown that Tdp1 can liberate the 3′ DNA phosphate termini from APurinic/APyrimidinic (AP) Sites. Here, we found that Tdp1 is more active in the cleavage of the AP Sites inside bubble-DNA structure in comparison to ssDNA containing AP Site. Furthermore, Tdp1 hydrolyzes AP Sites oppoSite to bulky fluorescein adduct faster than AP Sites located in dsDNA. Whilst the Tdp1 H493R (SCAN1) and H263A mutants retain the ability to bind an AP Site-containing DNA, both mutants do not reveal endonuclease activity, further suggesting the specificity of the AP cleavage activity. We suggest that this Tdp1 activity can contribute to the repair of AP Sites particularly in DNA structures containing ssDNA region or AP Sites in the context of clustered DNA lesions.
-
AP Site cleavage activity of tyrosyl dna phosphodiesterase 1
FEBS Letters, 2011Co-Authors: N A Lebedeva, Nadejda I Rechkunova, Olga I. LavrikAbstract:APE-independent base excision repair (BER) pathway plays an important role in the regulation of DNA repair mechanisms. In this study it has been found that recently discovered tyrosyl-DNA phosphodiesterase 1 (Tdp1) catalyzes the AP Site cleavage reaction to generate breaks with the 3′- and 5′-phosphate termini. The removal of the 3′-phosphate is performed by polynucleotide kinase phosphatase (PNKP). Tdp1 is known to interact stably with BER proteins: DNA polymerase beta (Pol β), XRCC1, PARP1 and DNA ligase III. The data suggest a role of Tdp1 in the new APE-independent BER pathway in mammals.
O I Lavrik - One of the best experts on this subject based on the ideXlab platform.
-
AP endonuclease 1 as a key enzyme in repair of APurinic/APyrimidinic Sites
Biochemistry (Moscow), 2016Co-Authors: N S Dyrkheeva, N A Lebedeva, O I LavrikAbstract:Human APurinic/APyrimidinic endonuclease 1 (APE1) is one of the key participants in the DNA base excision repair system. APE1 hydrolyzes DNA adjacent to the 5′-end of an APurinic/APyrimidinic (AP) Site to produce a nick with a 3′-hydroxyl group and a 5′-deoxyribose phosphate moiety. APE1 exhibits 3′-phosphodiesterase, 3′-5′-exonuclease, and 3-phosphatase activities. APE1 was also identified as a redox factor (Ref-1). In this review, data on the role of APE1 in the DNA repair process and in other metabolic processes occurring in cells are analyzed as well as the interaction of this enzyme with DNA and other proteins participating in the repair system.
-
poly adp ribose polymerase 1 stimulates the AP Site cleavage activity of tyrosyl dna phosphodiesterase 1
Bioscience Reports, 2015Co-Authors: N A Lebedeva, Maria V Sukhanova, Inna A Vasileva, Nadejda I Rechkunova, Rashid O Anarbaev, O I LavrikAbstract:The influence of poly(ADP-ribose)polymerase 1 (PARP1) on the APurinic/APyrimidinic (AP)-Site cleavage activity of tyrosyl–DNA phosphodiesterase 1 (TDP1) and interaction of PARP1 and TDP1 were studied. The efficiency of single or clustered AP-Site hydrolysis catalysed by TDP1 was estimated. It was shown that the efficiency of AP-Site cleavage increases in the presence of an additional AP-Site in the oppoSite DNA strand depending on its position. PARP1 stimulates TDP1; the stimulation effect was abolished in the presence of NAD+. The interaction of these two proteins was characterized quantitatively by measuring the dissociation constant for the TDP1–PARP1 complex using fluorescently-labelled proteins. The distance between the N-termini of the proteins within the complex was estimated using FRET. The data obtained suggest that PARP1 and TDP1 bind in an antiparallel orientation; the N-terminus of the former protein interacts with the C-terminal domain of the latter. The functional significance of PARP1 and TDP1 interaction in the process of DNA repair was demonstrated for the first time.
-
the mechanism of human tyrosyl dna phosphodiesterase 1 in the cleavage of AP Site and its synthetic analogs
DNA Repair, 2013Co-Authors: N A Lebedeva, Alexander A Ishchenko, Murat Saparbaev, Nadejda I Rechkunova, O I LavrikAbstract:Abstract The mechanism of hydrolysis of the APurinic/APyrimidinic (AP) Site and its synthetic analogs by using tyrosyl-DNA phosphodiesterase 1 (Tdp1) was analyzed. Tdp1 catalyzes the cleavage of AP Site and the synthetic analog of the AP Site, 3-hydroxy-2(hydroxymethyl)-tetrahydrofuran (THF), in DNA by hydrolysis of the phosphodiester bond between the substituent and 5′ adjacent phosphate. The product of Tdp1 cleavage in the case of the AP Site is unstable and is hydrolyzed with the formation of 3′- and 5′-margin phosphates. The following repair demands the ordered action of polynucleotide kinase phosphorylase, with XRCC1, DNA polymerase β, and DNA ligase. In the case of THF, Tdp1 generates break with the 5′-THF and the 3′-phosphate termini. Tdp1 is also able to effectively cleave non-nucleotide insertions in DNA, decanediol and diethyleneglycol moieties by the same mechanism as in the case of THF cleavage. The efficiency of Tdp1 catalyzed hydrolysis of AP-Site analog correlates with the DNA helix distortion induced by the substituent. The following repair of 5′-THF and other AP-Site analogs can be processed by the long-patch base excision repair pathway.
-
clustered dna lesions containing 5 formyluracil and AP Site repair via the ber system
PLOS ONE, 2013Co-Authors: E A Belousova, Inna A Vasileva, Nina Moor, Timofei S Zatsepin, T S Oretskaya, O I LavrikAbstract:Lesions in the DNA arise under ionizing irradiation conditions or various chemical oxidants as a single damage or as part of a multiply damaged Site within 1–2 helical turns (clustered lesion). Here, we explored the repair opportunity of the APurinic/APyrimidinic Site (AP Site) composed of the clustered lesion with 5-formyluracil (5-foU) by the base excision repair (BER) proteins. We found, that if the AP Site is shifted relative to the 5-foU of the oppoSite strand, it could be repaired primarily via the short-patch BER pathway. In this case, the cleavage efficiency of the AP Site-containing DNA strand catalyzed by human APurinic/APyrimidinic endonuclease 1 (hAPE1) decreased under AP Site excursion to the 3'-side relative to the lesion in the other DNA strand. DNA synthesis catalyzed by DNA polymerase lambda was more accurate in comparison to the one catalyzed by DNA polymerase beta. If the AP Site was located exactly oppoSite 5-foU it was expected to switch the repair to the long-patch BER pathway. In this situation, human processivity factor hPCNA stimulates the process.
N A Lebedeva - One of the best experts on this subject based on the ideXlab platform.
-
AP endonuclease 1 as a key enzyme in repair of APurinic/APyrimidinic Sites
Biochemistry (Moscow), 2016Co-Authors: N S Dyrkheeva, N A Lebedeva, O I LavrikAbstract:Human APurinic/APyrimidinic endonuclease 1 (APE1) is one of the key participants in the DNA base excision repair system. APE1 hydrolyzes DNA adjacent to the 5′-end of an APurinic/APyrimidinic (AP) Site to produce a nick with a 3′-hydroxyl group and a 5′-deoxyribose phosphate moiety. APE1 exhibits 3′-phosphodiesterase, 3′-5′-exonuclease, and 3-phosphatase activities. APE1 was also identified as a redox factor (Ref-1). In this review, data on the role of APE1 in the DNA repair process and in other metabolic processes occurring in cells are analyzed as well as the interaction of this enzyme with DNA and other proteins participating in the repair system.
-
poly adp ribose polymerase 1 stimulates the AP Site cleavage activity of tyrosyl dna phosphodiesterase 1
Bioscience Reports, 2015Co-Authors: N A Lebedeva, Maria V Sukhanova, Inna A Vasileva, Nadejda I Rechkunova, Rashid O Anarbaev, O I LavrikAbstract:The influence of poly(ADP-ribose)polymerase 1 (PARP1) on the APurinic/APyrimidinic (AP)-Site cleavage activity of tyrosyl–DNA phosphodiesterase 1 (TDP1) and interaction of PARP1 and TDP1 were studied. The efficiency of single or clustered AP-Site hydrolysis catalysed by TDP1 was estimated. It was shown that the efficiency of AP-Site cleavage increases in the presence of an additional AP-Site in the oppoSite DNA strand depending on its position. PARP1 stimulates TDP1; the stimulation effect was abolished in the presence of NAD+. The interaction of these two proteins was characterized quantitatively by measuring the dissociation constant for the TDP1–PARP1 complex using fluorescently-labelled proteins. The distance between the N-termini of the proteins within the complex was estimated using FRET. The data obtained suggest that PARP1 and TDP1 bind in an antiparallel orientation; the N-terminus of the former protein interacts with the C-terminal domain of the latter. The functional significance of PARP1 and TDP1 interaction in the process of DNA repair was demonstrated for the first time.
-
the mechanism of human tyrosyl dna phosphodiesterase 1 in the cleavage of AP Site and its synthetic analogs
DNA Repair, 2013Co-Authors: N A Lebedeva, Alexander A Ishchenko, Murat Saparbaev, Nadejda I Rechkunova, O I LavrikAbstract:Abstract The mechanism of hydrolysis of the APurinic/APyrimidinic (AP) Site and its synthetic analogs by using tyrosyl-DNA phosphodiesterase 1 (Tdp1) was analyzed. Tdp1 catalyzes the cleavage of AP Site and the synthetic analog of the AP Site, 3-hydroxy-2(hydroxymethyl)-tetrahydrofuran (THF), in DNA by hydrolysis of the phosphodiester bond between the substituent and 5′ adjacent phosphate. The product of Tdp1 cleavage in the case of the AP Site is unstable and is hydrolyzed with the formation of 3′- and 5′-margin phosphates. The following repair demands the ordered action of polynucleotide kinase phosphorylase, with XRCC1, DNA polymerase β, and DNA ligase. In the case of THF, Tdp1 generates break with the 5′-THF and the 3′-phosphate termini. Tdp1 is also able to effectively cleave non-nucleotide insertions in DNA, decanediol and diethyleneglycol moieties by the same mechanism as in the case of THF cleavage. The efficiency of Tdp1 catalyzed hydrolysis of AP-Site analog correlates with the DNA helix distortion induced by the substituent. The following repair of 5′-THF and other AP-Site analogs can be processed by the long-patch base excision repair pathway.
-
Tyrosyl-DNA phosphodiesterase 1 initiates repair of APurinic/APyrimidinic Sites.
Biochimie, 2012Co-Authors: N A Lebedeva, Nadejda I Rechkunova, Sherif F. El-khamisy, Olga I. LavrikAbstract:Tyrosyl-DNA phosphodiesterase 1 (Tdp1) catalyzes the hydrolysis of the phosphodiester linkage between the DNA 3′ phosphate and a tyrosine residue as well as a variety of other DNA 3′ damaged termini. Recently we have shown that Tdp1 can liberate the 3′ DNA phosphate termini from APurinic/APyrimidinic (AP) Sites. Here, we found that Tdp1 is more active in the cleavage of the AP Sites inside bubble-DNA structure in comparison to ssDNA containing AP Site. Furthermore, Tdp1 hydrolyzes AP Sites oppoSite to bulky fluorescein adduct faster than AP Sites located in dsDNA. Whilst the Tdp1 H493R (SCAN1) and H263A mutants retain the ability to bind an AP Site-containing DNA, both mutants do not reveal endonuclease activity, further suggesting the specificity of the AP cleavage activity. We suggest that this Tdp1 activity can contribute to the repair of AP Sites particularly in DNA structures containing ssDNA region or AP Sites in the context of clustered DNA lesions.
-
AP Site cleavage activity of tyrosyl dna phosphodiesterase 1
FEBS Letters, 2011Co-Authors: N A Lebedeva, Nadejda I Rechkunova, Olga I. LavrikAbstract:APE-independent base excision repair (BER) pathway plays an important role in the regulation of DNA repair mechanisms. In this study it has been found that recently discovered tyrosyl-DNA phosphodiesterase 1 (Tdp1) catalyzes the AP Site cleavage reaction to generate breaks with the 3′- and 5′-phosphate termini. The removal of the 3′-phosphate is performed by polynucleotide kinase phosphatase (PNKP). Tdp1 is known to interact stably with BER proteins: DNA polymerase beta (Pol β), XRCC1, PARP1 and DNA ligase III. The data suggest a role of Tdp1 in the new APE-independent BER pathway in mammals.
Murat Saparbaev - One of the best experts on this subject based on the ideXlab platform.
-
reading targeted dna damage in the active demethylation pathway role of accessory domains of eukaryotic AP endonucleases and thymine dna glycosylases
Journal of Molecular Biology, 2020Co-Authors: Alexander V Popov, Murat Saparbaev, Inga R Grin, Antonina P Dvornikova, Bakhyt T Matkarimov, Regina Groisman, Dmitry O ZharkovAbstract:Abstract Base excision DNA repair (BER) is an important process used by all living organisms to remove nonbulky lesions from DNA. BER is usually initiated by DNA glycosylases that excise a damaged base leaving an APurinic/APyrimidinic (AP) Site, and an AP endonuclease then cuts DNA at the AP Site, and the repair is completed by correct nucleotide insertion, end processing, and nick ligation. It has emerged recently that the BER machinery, in addition to genome protection, is crucial for active epigenetic demethylation in the vertebrates. This pathway is initiated by TET dioxygenases that oxidize the regulatory 5-methylcytosine, and the oxidation products are treated as substrates for BER. T:G mismatch-specific thymine-DNA glycosylase (TDG) and AP endonuclease 1 (APE1) catalyze the first two steps in BER-dependent active demethylation. In addition to the well-structured catalytic domains, these enzymes possess long tails that are structurally uncharacterized but involved in multiple interactions and regulatory functions. In this review, we describe the known roles of the tails in TDG and APE1, discuss the importance of order and disorder in their structure, and consider the evolutionary aspects of these accessory protein regions. We also propose that the tails may be important for the enzymes’ oligomerization on DNA, an aspect of their function that only recently gained attention.
-
pre steady state fluorescence analysis of damaged dna transfer from human dna glycosylases to AP endonuclease APe1
Biochimica et Biophysica Acta, 2014Co-Authors: Alexandra A Kuznetsova, Alexander A Ishchenko, Nikita A Kuznetsov, Murat Saparbaev, Olga S FedorovaAbstract:Abstract Background DNA glycosylases remove the modified, damaged or mismatched bases from the DNA by hydrolyzing the N-glycosidic bonds. Some enzymes can further catalyze the incision of a resulting abasic (APurinic/APyrimidinic, AP) Site through β- or β,δ-elimination mechanisms. In most cases, the incision reaction of the AP-Site is catalyzed by special enzymes called AP-endonucleases. Methods Here, we report the kinetic analysis of the mechanisms of modified DNA transfer from some DNA glycosylases to the AP endonuclease, APE1. The modified DNA contained the tetrahydrofurane residue (F), the analogue of the AP-Site. DNA glycosylases AAG, OGG1, NEIL1, MBD4 cat and UNG from different structural superfamilies were used. Results We found that all DNA glycosylases may utilise direct protein–protein interactions in the transient ternary complex for the transfer of the AP-containing DNA strand to APE1. Conclusions We hypothesize a fast “flip-flop” exchange mechanism of damaged and undamaged DNA strands within this complex for monofunctional DNA glycosylases like MBD4 cat , AAG and UNG. Bifunctional DNA glycosylase NEIL1 creates tightly specific complex with DNA containing F-Site thereby efficiently competing with APE1. Whereas APE1 fast displaces other bifunctional DNA glycosylase OGG1 on F-Site thereby induces its shifts to undamaged DNA regions. General significance Kinetic analysis of the transfer of DNA between human DNA glycosylases and APE1 allows us to elucidate the critical step in the base excision repair pathway.
-
the mechanism of human tyrosyl dna phosphodiesterase 1 in the cleavage of AP Site and its synthetic analogs
DNA Repair, 2013Co-Authors: N A Lebedeva, Alexander A Ishchenko, Murat Saparbaev, Nadejda I Rechkunova, O I LavrikAbstract:Abstract The mechanism of hydrolysis of the APurinic/APyrimidinic (AP) Site and its synthetic analogs by using tyrosyl-DNA phosphodiesterase 1 (Tdp1) was analyzed. Tdp1 catalyzes the cleavage of AP Site and the synthetic analog of the AP Site, 3-hydroxy-2(hydroxymethyl)-tetrahydrofuran (THF), in DNA by hydrolysis of the phosphodiester bond between the substituent and 5′ adjacent phosphate. The product of Tdp1 cleavage in the case of the AP Site is unstable and is hydrolyzed with the formation of 3′- and 5′-margin phosphates. The following repair demands the ordered action of polynucleotide kinase phosphorylase, with XRCC1, DNA polymerase β, and DNA ligase. In the case of THF, Tdp1 generates break with the 5′-THF and the 3′-phosphate termini. Tdp1 is also able to effectively cleave non-nucleotide insertions in DNA, decanediol and diethyleneglycol moieties by the same mechanism as in the case of THF cleavage. The efficiency of Tdp1 catalyzed hydrolysis of AP-Site analog correlates with the DNA helix distortion induced by the substituent. The following repair of 5′-THF and other AP-Site analogs can be processed by the long-patch base excision repair pathway.
-
the hAP1 protein stimulates the turnover of human mismatch specific thymine dna glycosylase to process 3 n4 ethenocytosine residues
Mutation Research, 2001Co-Authors: Cyril V Privezentzev, Murat Saparbaev, Jacques LavalAbstract:Abstract When present in DNA, 3,N 4 -ethenocytosine (eC) residues are potentially mutagenic and carcinogenic in vivo. The enzymatic activity responsible for the repair of the eC residues in human cells is the hTDG protein, the human thymine-DNA-glycosylase that removes thymine in a T/G base pair [Proc. Natl. Acad. Sci., U.S.A., 95 (1998) 8508]. One of the distinctive properties of the hTDG protein is that it remains tightly bound to the AP-Site resulting from its glycosylase activity. In this pAPer we report that the human AP endonuclease, the HAP1 (APe1, APEX Ref-1) protein, stimulates the processing of eC residues by the hTDG protein in vitro, in a dose-dependent manner. This property of HAP1 protein is specific since E.coli Fpg, Nfo and Nth proteins, all endowed with an AP nicking activity, do not show similar features. The results suggest that the HAP1 protein displaces the hTDG protein bound to the AP-Site and therefore increases the turnover of the hTDG protein. However, using a variety of techniques including gel retardation assay, surface plasmon resonance and two-hybrid system, it was not possible to detect evidence for a complex including the substrate, the hTDG and HAP1 proteins.
-
The HAP1 protein stimulates the turnover of human mismatch-specific thymine-DNA-glycosylase to process 3,N(4)-ethenocytosine residues.
Mutation research, 2001Co-Authors: Cyril V Privezentzev, Murat Saparbaev, Jacques LavalAbstract:When present in DNA, 3,N(4)-ethenocytosine (epsilon C) residues are potentially mutagenic and carcinogenic in vivo. The enzymatic activity responsible for the repair of the epsilon C residues in human cells is the hTDG protein, the human thymine-DNA-glycosylase that removes thymine in a T/G base pair [Proc. Natl. Acad. Sci., U.S.A., 95 (1998) 8508]. One of the distinctive properties of the hTDG protein is that it remains tightly bound to the AP-Site resulting from its glycosylase activity. In this pAPer we report that the human AP endonuclease, the HAP1 (APe1, APEX Ref-1) protein, stimulates the processing of epsilon C residues by the hTDG protein in vitro, in a dose-dependent manner. This property of HAP1 protein is specific since E.coli Fpg, Nfo and Nth proteins, all endowed with an AP nicking activity, do not show similar features. The results suggest that the HAP1 protein displaces the hTDG protein bound to the AP-Site and therefore increases the turnover of the hTDG protein. However, using a variety of techniques including gel retardation assay, surface plasmon resonance and two-hybrid system, it was not possible to detect evidence for a complex including the substrate, the hTDG and HAP1 proteins.
Dmitry O Zharkov - One of the best experts on this subject based on the ideXlab platform.
-
reading targeted dna damage in the active demethylation pathway role of accessory domains of eukaryotic AP endonucleases and thymine dna glycosylases
Journal of Molecular Biology, 2020Co-Authors: Alexander V Popov, Murat Saparbaev, Inga R Grin, Antonina P Dvornikova, Bakhyt T Matkarimov, Regina Groisman, Dmitry O ZharkovAbstract:Abstract Base excision DNA repair (BER) is an important process used by all living organisms to remove nonbulky lesions from DNA. BER is usually initiated by DNA glycosylases that excise a damaged base leaving an APurinic/APyrimidinic (AP) Site, and an AP endonuclease then cuts DNA at the AP Site, and the repair is completed by correct nucleotide insertion, end processing, and nick ligation. It has emerged recently that the BER machinery, in addition to genome protection, is crucial for active epigenetic demethylation in the vertebrates. This pathway is initiated by TET dioxygenases that oxidize the regulatory 5-methylcytosine, and the oxidation products are treated as substrates for BER. T:G mismatch-specific thymine-DNA glycosylase (TDG) and AP endonuclease 1 (APE1) catalyze the first two steps in BER-dependent active demethylation. In addition to the well-structured catalytic domains, these enzymes possess long tails that are structurally uncharacterized but involved in multiple interactions and regulatory functions. In this review, we describe the known roles of the tails in TDG and APE1, discuss the importance of order and disorder in their structure, and consider the evolutionary aspects of these accessory protein regions. We also propose that the tails may be important for the enzymes’ oligomerization on DNA, an aspect of their function that only recently gained attention.
-
recognition but no repair of abasic Site in single stranded dna by human ribosomal us3 protein residing within intact 40s subunit
Nucleic Acids Research, 2017Co-Authors: Anastasia S Grosheva, Dmitry O Zharkov, Joachim Stahl, Alexander V Gopanenko, Alexey E Tupikin, Marsel R Kabilov, Dmitri M Graifer, G G KarpovaAbstract:Isolated human ribosomal protein uS3 has extra-ribosomal functions including those related to base excision DNA repair, e.g. AP lyase activity that nicks double-stranded (ds) DNA 3' to the abasic (AP) Site. However, the ability of uS3 residing within ribosome to recognize and cleave damaged DNA has never been addressed. Here, we compare interactions of single-stranded (ss) DNA and dsDNA bearing AP Site with human ribosome-bound uS3 and with the isolated protein, whose interactions with ssDNA were not yet studied. The AP lyase activity of free uS3 was much higher with ssDNA than with dsDNA, whereas ribosome-bound uS3 was completely deprived of this activity. Nevertheless, an exposed peptide of ribosome-bound uS3 located far away from the putative catalytic center previously suggested for isolated uS3 cross-linked to full-length uncleaved ssDNA, but not to dsDNA. In contrast, free uS3 cross-linked mainly to the 5'-part of the damaged DNA strand after its cleavage at the AP Site. ChIP-seq analysis showed preferential uS3 binding to nucleolus-associated chromatin domains. We conclude that free and ribosome-bound uS3 proteins interact with AP Sites differently, exhibiting their non-translational functions in DNA repair in and around the nucleolus and in regulation of DNA damage response in looped DNA structures, respectively.
-
effect of the multifunctional proteins rpa yb 1 and xpc repair factor on AP Site cleavage by dna glycosylase neil1
Journal of Molecular Recognition, 2012Co-Authors: P E Pestryakov, Dmitry O Zharkov, Inga R Grin, Elizaveta E Fomina, Loic Hamon, Irina A Eliseeva, I O Petruseva, Patrick A CurmiAbstract:DNA glycosylases are key enzymes in the first step of base excision DNA repair, recognizing DNA damage and catalyzing the release of damaged nucleobases. Bifunctional DNA glycosylases also possess associated APurinic/APyrimidinic (AP) lyase activity that nick the damaged DNA strand at an abasic (or AP) Site, formed either spontaneously or at the first step of repair. NEIL1 is a bifunctional DNA glycosylase cAPable of processing lesions, including AP Sites, not only in double-stranded but also in single-stranded DNA. Here, we show that proteins participating in DNA damage response, YB-1 and RPA, affect AP Site cleavage by NEIL1. Stimulation of the AP lyase activity of NEIL1 was observed when an AP Site was located in a 60 nt-long double-stranded DNA. Both RPA and YB-1 inhibited AP Site cleavage by NEIL1 when the AP Site was located in single-stranded DNA. Taking into account a direct interaction of YB-1 with the AP Site, located in single-stranded DNA, and the high affinity of both YB-1 and RPA for single-stranded DNA, this behavior is presumably a consequence of a competition with NEIL1 for the DNA substrate. Xeroderma pigmentosum complementation group C protein (XPC), a key protein of another DNA repair pathway, was shown to interact directly with AP Sites but had no effect on AP Site cleavage by NEIL1. Copyright © 2012 John Wiley & Sons, Ltd.
