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Leona D Samson - One of the best experts on this subject based on the ideXlab platform.
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Does the cycad genotoxin MAM implicated in Guam ALS-PDC induce disease-relevant changes in mouse brain that includes olfaction?
Communicative & Integrative Biology, 2011Co-Authors: Glen E Kisby, Eli A. Magun, Michael R Lasarev, Leona D Samson, Mihail S. Iordanov, Valerie S. Palmer, Peter S. SpencerAbstract:Western Pacific amyotrophic lateral sclerosis (ALS) and parkinsonism-dementia complex (PDC), a prototypical neurodegenerative disease (tauopathy) affecting distinct genetic groups with common exposure to neurotoxic chemicals in cycad seed, has many features of Parkinson's and Alzheimer's diseases (AD), including early olfactory dysfunction. Guam ALS-PDC incidence correlates with cycad flour content of cycasin and its aglycone methylazoxymethanol (MAM), which produces persistent DNA damage (O6-methylguanine) in the brains of mice lacking O6-methylguanine methyltransferase (Mgmt-/-). We described in Mgmt-/-mice up to 7 days post-MAM treatment that brain DNA damage was linked to brain gene expression changes found in human neurological disease, cancer, and skin and hair development. This addendum reports 6 months post-MAM treatment- related brain transcriptional changes as well as elevated mitogen activated protein kinases and increased caspase-3 activity, both of which are involved in tau aggregation and neurofibrillary tangle formation typical of ALS-PDC and AD, plus transcriptional changes in olfactory receptors. Does cycasin act as a "slow (geno)toxin" in ALS-PDC?
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recognition and processing of a new repertoire of dna substrates by human 3 methyladenine dna glycosylase aag
Biochemistry, 2009Co-Authors: James C Delaney, Maria Kartalou, Gondichatnahalli M Lingaraju, Ayelet Maorshoshani, John M. Essigmann, Leona D SamsonAbstract:The human 3-methyladenine DNA glycosylase (AAG) recognizes and excises a broad range of purines damaged by alkylation and oxidative damage, including 3-methyladenine, 7-Methylguanine, hypoxanthine (Hx), and 1,N6-ethenoadenine (eA). The crystal structures of AAG bound to eA have provided insights into the structural basis for substrate recognition, base excision, and exclusion of normal purines and pyrimidines from its substrate recognition pocket. In this study, we explore the substrate specificity of full-length and truncated Δ80AAG on a library of oligonucleotides containing structurally diverse base modifications. Substrate binding and base excision kinetics of AAG with 13 damaged oligonucleotides were examined. We found that AAG bound to a wide variety of purine and pyrimidine lesions but excised only a few of them. Single-turnover excision kinetics showed that in addition to the well-known eA and Hx substrates, 1-methylguanine (m1G) was also excised efficiently by AAG. Thus, along with eA and ethanoa...
Ralf Ficner - One of the best experts on this subject based on the ideXlab platform.
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Crystal Structures of the Novel Cytosolic 59-Nucleotidase IIIB Explain Its Preference for m7GMP
2016Co-Authors: Thomas Monecke, Juliane Buschmann, Piotr Neumann, Elmar Wahle, Ralf FicnerAbstract:59-nucleotidases catalyze the hydrolytic dephosphorylation of nucleoside monophosphates. As catabolic enzymes they contribute significantly to the regulation of cellular nucleotide levels; misregulation of nucleotide metabolism and nucleotidase deficiencies are associated with a number of diseases. The seven human 59-nucleotidases differ with respect to substrate specificity and cellular localization. Recently, the novel cytosolic 59-nucleotidase III-like protein, or cN-IIIB, has been characterized in human and Drosophila. cN-IIIB exhibits a strong substrate preference for the modified nucleotide 7-methylguanosine monophosphate but the structural reason for this preference was unknown. Here, we present crystal structures of cN-IIIB from Drosophila melanogaster bound to the reaction products 7-methylguanosine or cytidine. The structural data reveal that the cytosine- and 7-Methylguanine moieties of the products are stacked between two aromatic residues in a coplanar but off-centered position. 7-methylguanosine is specifically bound through p-p interactions and distinguished from unmodified guanosine by additional cation-p coulomb interactions between the aromatic side chains and the positively charged 7-Methylguanine. Notably, the base is further stabilized by T-shaped edge-to-face stacking of an additional tryptophan packing perpendicularly against the purine ring and forming, together with the other aromates, an aromatic slot. The structural data in combination with site-directed mutagenesis experiments reveal the molecular basis for the broad substrate specificity of cN-IIIB but also explain the substrate preference for 7-methylguanosine monophosphate
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Crystal Structures of the Novel Cytosolic 5′-Nucleotidase IIIB Explain Its Preference for m7GMP
PloS one, 2014Co-Authors: Thomas Monecke, Juliane Buschmann, Piotr Neumann, Elmar Wahle, Ralf FicnerAbstract:5′-nucleotidases catalyze the hydrolytic dephosphorylation of nucleoside monophosphates. As catabolic enzymes they contribute significantly to the regulation of cellular nucleotide levels; misregulation of nucleotide metabolism and nucleotidase deficiencies are associated with a number of diseases. The seven human 5′-nucleotidases differ with respect to substrate specificity and cellular localization. Recently, the novel cytosolic 5′-nucleotidase III-like protein, or cN-IIIB, has been characterized in human and Drosophila. cN-IIIB exhibits a strong substrate preference for the modified nucleotide 7-methylguanosine monophosphate but the structural reason for this preference was unknown. Here, we present crystal structures of cN-IIIB from Drosophila melanogaster bound to the reaction products 7-methylguanosine or cytidine. The structural data reveal that the cytosine- and 7-Methylguanine moieties of the products are stacked between two aromatic residues in a coplanar but off-centered position. 7-methylguanosine is specifically bound through π-π interactions and distinguished from unmodified guanosine by additional cation-π coulomb interactions between the aromatic side chains and the positively charged 7-Methylguanine. Notably, the base is further stabilized by T-shaped edge-to-face stacking of an additional tryptophan packing perpendicularly against the purine ring and forming, together with the other aromates, an aromatic slot. The structural data in combination with site-directed mutagenesis experiments reveal the molecular basis for the broad substrate specificity of cN-IIIB but also explain the substrate preference for 7-methylguanosine monophosphate. Analyzing the substrate specificities of cN-IIIB and the main pyrimidine 5′-nucleotidase cN-IIIA by mutagenesis studies, we show that cN-IIIA dephosphorylates the purine m7GMP as well, hence redefining its substrate spectrum. Docking calculations with cN-IIIA and m7GMP as well as biochemical data reveal that Asn69 does not generally exclude the turnover of purine substrates thus correcting previous suggestions.
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Crystal structures of the novel cytosolic 5'-nucleotidase IIIB explain its preference for m7GMP.
Public Library of Science (PLoS), 2024Co-Authors: Thomas Monecke, Juliane Buschmann, Piotr Neumann, Elmar Wahle, Ralf FicnerAbstract:5'-nucleotidases catalyze the hydrolytic dephosphorylation of nucleoside monophosphates. As catabolic enzymes they contribute significantly to the regulation of cellular nucleotide levels; misregulation of nucleotide metabolism and nucleotidase deficiencies are associated with a number of diseases. The seven human 5'-nucleotidases differ with respect to substrate specificity and cellular localization. Recently, the novel cytosolic 5'-nucleotidase III-like protein, or cN-IIIB, has been characterized in human and Drosophila. cN-IIIB exhibits a strong substrate preference for the modified nucleotide 7-methylguanosine monophosphate but the structural reason for this preference was unknown. Here, we present crystal structures of cN-IIIB from Drosophila melanogaster bound to the reaction products 7-methylguanosine or cytidine. The structural data reveal that the cytosine- and 7-Methylguanine moieties of the products are stacked between two aromatic residues in a coplanar but off-centered position. 7-methylguanosine is specifically bound through π-π interactions and distinguished from unmodified guanosine by additional cation-π coulomb interactions between the aromatic side chains and the positively charged 7-Methylguanine. Notably, the base is further stabilized by T-shaped edge-to-face stacking of an additional tryptophan packing perpendicularly against the purine ring and forming, together with the other aromates, an aromatic slot. The structural data in combination with site-directed mutagenesis experiments reveal the molecular basis for the broad substrate specificity of cN-IIIB but also explain the substrate preference for 7-methylguanosine monophosphate. Analyzing the substrate specificities of cN-IIIB and the main pyrimidine 5'-nucleotidase cN-IIIA by mutagenesis studies, we show that cN-IIIA dephosphorylates the purine m7GMP as well, hence redefining its substrate spectrum. Docking calculations with cN-IIIA and m7GMP as well as biochemical data reveal that Asn69 does not generally exclude the turnover of purine substrates thus correcting previous suggestions
John M. Essigmann - One of the best experts on this subject based on the ideXlab platform.
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Recognition and Processing of a New Repertoire of DNA Substrates by Human 3-Methyladenine DNA Glycosylase (AAG)†
2016Co-Authors: Ayelet Maor-shoshani, John M. Essigmann, Leona DAbstract:The human 3-methyladenine DNA glycosylase (AAG) recognizes and excises a broad range of purines damaged by alkylation and oxidative damage, including 3-methyladenine, 7-Methylguanine, hypoxanthine (Hx), and 1,N6-ethenoadenine (εA). The crystal structures of AAG bound to εA have provided insights into the structural basis for substrate recognition, base excision, and exclusion of normal purines and pyrimidines from its substrate recognition pocket. In the present study, we explored the substrate specificity of full-length and truncated Δ80AAG on a library of oligonucleotides containing structurally diverse base modifications. Substrate binding and base excision kinetics of AAG with 13 damaged oligonucleotides were examined. We found that AAG bound to a wide variety of purine and pyrimidine lesions, but excised only few of them. Single-turnover excision kinetics showed that in addition to the well-known εA and Hx substrates, 1-methylguanine (m1G) was also excised efficiently by AAG. Thus, along with εA and ethanoadenine (EA), m1G is another substrate that is shared between AAG and the direct repair protein AlkB. In addition, we found that both the full-length and truncated AAG excised 1,N2-ethenoguanine (1,N2-εG), albeit weakly, from duplex DNA. Uracil was excised from both single- and double-strande
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Removal of N‑Alkyl Modifications from N2‑Alkylguanine and N4‑Alkylcytosine in DNA by the Adaptive Response Protein AlkB
2016Co-Authors: Cintyu Wong, Catherine L. Drennan, John M. EssigmannAbstract:ABSTRACT: The AlkB enzyme is an Fe(II)- and α-ketoglutarate-dependent dioxygenase that repairs DNA alkyl lesions by a direct reversal of damage mechanism as part of the adaptive response in E. coli. The reported substrate scope of AlkB includes simple DNA alkyl adducts, such as 1-methyladenine, 3-methylcytosine, 3-ethylcytosine, 1-methylgua-nine, 3-methylthymine, and N6-methyladenine, as well as more complex DNA adducts, such as 1,N6-ethenoadenine, 3,N4-ethenocytosine, and 1,N6-ethanoadenine. Previous studies have revealed, in a piecemeal way, that AlkB has an impressive repertoire of substrates. The present study makes two additions to this list, showing that alkyl adducts on the N2 position of guanine and N4 position of cytosine are also substrates for AlkB. Using high resolution ESI-TOF mass spectrometry, we show that AlkB has the biochemical capability to repair in vitro N2-methylguanine, N2-ethylguanine, N2-furan-2-yl-methylguanine, N2-tetrahydrofuran-2-yl-methylguanine, and N4-methylcytosine in ssDNA but not in dsDNA. When viewed together with previous work, the experimental data herein demonstrate that AlkB is abl
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a chemical genetics analysis of the roles of bypass polymerase dinb and dna repair protein alkb in processing n2 alkylguanine lesions in vivo
PLOS ONE, 2014Co-Authors: Nidhi Shrivastav, James C Delaney, Bogdan I Fedeles, Lauren E Frick, James J Foti, Graham C Walker, John M. EssigmannAbstract:DinB, the E. coli translesion synthesis polymerase, has been shown to bypass several N2-alkylguanine adducts in vitro, including N2-furfurylguanine, the structural analog of the DNA adduct formed by the antibacterial agent nitrofurazone. Recently, it was demonstrated that the Fe(II)- and α-ketoglutarate-dependent dioxygenase AlkB, a DNA repair enzyme, can dealkylate in vitro a series of N2-alkyguanines, including N2-furfurylguanine. The present study explored, head to head, the in vivo relative contributions of these two DNA maintenance pathways (replicative bypass vs. repair) as they processed a series of structurally varied, biologically relevant N2-alkylguanine lesions: N2-furfurylguanine (FF), 2-tetrahydrofuran-2-yl-methylguanine (HF), 2-methylguanine, and 2-ethylguanine. Each lesion was chemically synthesized and incorporated site-specifically into an M13 bacteriophage genome, which was then replicated in E. coli cells deficient or proficient for DinB and AlkB (4 strains in total). Biochemical tools were employed to analyze the relative replication efficiencies of the phage (a measure of the bypass efficiency of each lesion) and the base composition at the lesion site after replication (a measure of the mutagenesis profile of each lesion). The main findings were: 1) Among the lesions studied, the bulky FF and HF lesions proved to be strong replication blocks when introduced site-specifically on a single-stranded vector in DinB deficient cells. This toxic effect disappeared in the strains expressing physiological levels of DinB. 2) AlkB is known to repair N2-alkylguanine lesions in vitro; however, the presence of AlkB showed no relief from the replication blocks induced by FF and HF in vivo. 3) The mutagenic properties of the entire series of N2-alkyguanines adducts were investigated in vivo for the first time. None of the adducts were mutagenic under the conditions evaluated, regardless of the DinB or AlkB cellular status. Taken together, the data indicated that the cellular pathway to combat bulky N2-alkylguanine DNA adducts was DinB-dependent lesion bypass.
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recognition and processing of a new repertoire of dna substrates by human 3 methyladenine dna glycosylase aag
Biochemistry, 2009Co-Authors: James C Delaney, Maria Kartalou, Gondichatnahalli M Lingaraju, Ayelet Maorshoshani, John M. Essigmann, Leona D SamsonAbstract:The human 3-methyladenine DNA glycosylase (AAG) recognizes and excises a broad range of purines damaged by alkylation and oxidative damage, including 3-methyladenine, 7-Methylguanine, hypoxanthine (Hx), and 1,N6-ethenoadenine (eA). The crystal structures of AAG bound to eA have provided insights into the structural basis for substrate recognition, base excision, and exclusion of normal purines and pyrimidines from its substrate recognition pocket. In this study, we explore the substrate specificity of full-length and truncated Δ80AAG on a library of oligonucleotides containing structurally diverse base modifications. Substrate binding and base excision kinetics of AAG with 13 damaged oligonucleotides were examined. We found that AAG bound to a wide variety of purine and pyrimidine lesions but excised only a few of them. Single-turnover excision kinetics showed that in addition to the well-known eA and Hx substrates, 1-methylguanine (m1G) was also excised efficiently by AAG. Thus, along with eA and ethanoa...
Timothy R Oconnor - One of the best experts on this subject based on the ideXlab platform.
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purification and characterization of human 3 methyladenine dna glycosylase
Nucleic Acids Research, 1993Co-Authors: Timothy R OconnorAbstract:Abstract A human cDNA coding sequence for a 3-methyladenine-DNA glycosylase was expressed in Escherichia coli. In addition to the full-length 3-methyladenine-DNA glycosylase coding sequence, two other sequences (resulting from differential RNA splicing and the truncated anpg cDNA) derived from that sequence were also expressed. All three proteins were purified to physical homogeneity and their N-terminal amino acid sequences are identical to those predicted by the nucleic acid sequences. The full-length protein has 293 amino acids coding for a protein with a molecular mass of 32 kDa. Polyclonal antibodies against one of the proteins react with the other two proteins, and a murine 3-methyladenine-DNA glycosylase, but not with several other E. coli DNA repair proteins. All three proteins excise 3-methyl-adenine, 7-Methylguanine, and 3-methylguanine as well as ethylated bases from DNA. The activities of the proteins with respect to ionic strength (optimum 100 mM KCl), pH (optimum 7.6), and kinetics for 3-methyladenine and 7-Methylguanine excision (average values: 3-methyladenine: Km 9 nM and kcat 10 min-1, 7-Methylguanine: Km 29 nM and kcat 0.38 min-1) are comparable. In contrast to these results, however, the thermal stability of the full-length and splicing variant proteins at 50 degrees C is less than that of the truncated protein.
Stephen A Smith - One of the best experts on this subject based on the ideXlab platform.
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in vivo repair of methylation damage in aag 3 methyladenine dna glycosylase null mouse cells
Nucleic Acids Research, 2000Co-Authors: Stephen A Smith, Bevin P EngelwardAbstract:3-Methyladenine (3MeA) DNA glycosylases initiate base excision repair by removing 3MeA. These glycosylases also remove a broad spectrum of spontaneous and environmentally induced base lesions in vitro. Mouse cells lacking the Aag 3MeA DNA glycosylase (also known as the Mpg, APNG or ANPG DNA glycosylase) are susceptible to 3MeA-induced S phase arrest, chromosome aberrations and apoptosis, but it is not known if Aag is solely responsible for repair of 3MeA in vivo. Here we show that in Aag–/– cells, 3MeA lesions disappear from the genome slightly faster than would be expected by spontaneous depurination alone, suggesting that there may be residual repair of 3MeA. However, repair of 3MeA is at least 10 times slower in Aag–/– cells than in Aag+/+ cells. Consequently, 24 h after exposure to [3H]MNU, 30% of the original 3MeA burden is intact in Aag–/– cells, while 3MeA is undetectable in Aag+/+ cells. Thus, Aag is the major DNA glycosylase for 3MeA repair. We also investigated the in vivo repair kinetics of another Aag substrate, 7-Methylguanine. Surprisingly, 7-Methylguanine is removed equally efficiently in Aag+/+ and Aag–/– cells, suggesting that another DNA glycosylase acts on lesions previously thought to be repaired by Aag.
