A common mechanism of action for the N-glycosylase activity of DNA N-glycosylase/AP lyases from E. coli and T4

Purmal, Andrei A.; Rabow, Lois E.; Lampman, Gary W.; Cunningham, Richard P.; Kow, Yoke W.

doi:10.1016/s0921-8777(96)00032-8

Cited by 28 publications

(20 citation statements)

References 45 publications

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“…It is well established that endonuclease III generates a 5′-cleavage product containing a 3′ 4-hydroxy-2-pentenal residue [β-elimination product; (27,28)], whereas endonuclease VIII generates a 3′ phosphoryl group [β,δ-elimination product; (27,28)], as evidenced by the different mobilities of their products (Figure 3, lanes 2 and 5). Endonucleases IV and V are AP endonucleases that generate hydroxyl groups at the 3′ termini of 5′-cleavage products (22,28). However, endonuclease V nicks DNA containing an AP site at the second phosphodiester bond 3′ to an AP site, thus generating a labeled 5′ cleavage product that is larger than that generated by endonuclease IV (Figure 3, lanes 3 and 4).…”

Section: Resultsmentioning

confidence: 99%

Escherichia coli HU protein has a role in the repair of abasic sites in DNA

Kow

Imhoff

Weiss

et al. 2007

Nucleic Acids Research

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HU is one of the most abundant DNA binding proteins in Escherichia coli. We find that it binds strongly to DNA containing an abasic (AP) site or tetrahydrofuran (THF) (apparent Kd ≈50 nM). It also possesses an AP lyase activity that cleaves at a deoxyribose but not at a THF residue. The binding and cleavage of an AP site was observed only with the HUαβ heterodimer. Site-specific mutations at K3 and R61 residues led to a change in substrate binding and cleavage. Both K3A(α)K3A(β) and R61A(α)R61A(β) mutant HU showed significant reduction in binding to DNA containing AP site; however, only R61A(α)R61A(β) mutant protein exhibited significant loss in AP lyase activity. Both K3A(α)K3A(β) and R61K(α)R61K(β) showed slight reduction in AP lyase activities. The function of HU protein as an AP lyase was confirmed by the ability of hupA or hupB mutations to further reduce the viability of an E. coli dut(Ts) xth mutant, which generates lethal AP sites at 37°C; the hupA and hupB derivatives, respectively, had a 6-fold and a 150-fold lower survival at 37°C than did the parental strain. These data suggest, therefore, that HU protein plays a significant role in the repair of AP sites in E. coli.

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Section: Resultsmentioning

confidence: 99%

Escherichia coli HU protein has a role in the repair of abasic sites in DNA

Kow

Imhoff

Weiss

et al. 2007

Nucleic Acids Research

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“…100,208,[210][211][212] Other evidence offered in favor of endocyclic mechanisms was the fact that a number of bifunctional enzymes could degrade urea nucleoside (9) and N-hydroxylamine compounds (10). 207 This evidence was not unambiguous, however, because (i) the reaction products were only characterized by band positions on gel electrophoresis and some reactions gave unexpected products, (ii) all the bifunctional enzymes degraded 9 and 10 at identical rates despite widely differing N-glycoside specificities, implying that, however they were degraded, it was not using the glycosylase catalytic machinery, and (iii) the mechanism of 10 breakdown was not characterized. It was only demonstrated that the enzymes were capable of acting on 10, not that breakdown was relevant to the glycosylase mechanism.…”

Section: Endocyclic Versus Exocyclic Hydrolysismentioning

confidence: 99%

Toward a Detailed Understanding of Base Excision Repair Enzymes: Transition State and Mechanistic Analyses of N-Glycoside Hydrolysis and N-Glycoside Transfer

2006

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“…Evidence supporting this mechanism emerged recently. For example, Fpg, EndoIII, and bacteriophage T4 endonuclease V are active on O-alkoxyamine-modified, ringopened abasic sites that share a double bond between C1Ј and the hydroxylamine nitrogen (69). KCN was found to inhibit DNA strand nicking and base release by mOgg1 (18).…”

Section: Substrate Specificity Of Mogg1: Recognition and Processing Omentioning

confidence: 99%

“…However, the geometry of the protonated pyrrolidine analog is different from that of a planar oxycarbenium ion and more closely resembles an oxonium ion; thus, the evidence does not support this mechanism well. More likely, following ring opening, an amino group of the enzyme carries out imine exchange, displacing the base and forming an enzyme-DNA covalent complex (53,69). Base protonation may still be important, facilitating imine exchange.…”

Section: Substrate Specificity Of Mogg1: Recognition and Processing Omentioning

confidence: 99%

Substrate Specificity and Reaction Mechanism of Murine 8-Oxoguanine-DNA Glycosylase

Zharkov

Rosenquist

Gerchman

et al. 2000

Journal of Biological Chemistry

185

178

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Genomic DNA is prone to oxidation by reactive oxygen species. A major product of DNA oxidation is the miscoding base 8-oxoguanine (8-oxoG). The mutagenic effects of 8-oxoG in mammalian cells are prevented by a DNA repair system consisting of 8-oxoguanine-DNA glycosylase (Ogg1), adenine-DNA glycosylase, and 8-oxodGTPase. We have cloned, overexpressed, and characterized mOgg1, the product of the murine ogg1 gene. mOgg1 is a DNA glycosylase/AP lyase belonging to the endonuclease III family of DNA repair enzymes. The AP lyase activity of mOgg1 is significantly lower than its glycosylase activity. mOgg1 releases 8-oxoG from DNA when paired with C, T, or G, but efficient DNA strand nicking is observed only with 8-oxoG:C. Binding of mOgg1 to oligonucleotides containing 8-oxoG:C is strong (K D ‫؍‬ 51.5 nM), unlike other mispairs. The average residence time for mOgg1 bound to substrate containing 8-oxoG:C is 18.3 min; the time course for accumulation of the NaBH 4 -sensitive intermediate suggests a two-step reaction mechanism. Various analogs of 8-oxoG were tested as substrates for mOgg1. An electron-withdrawing or hydrogen bond acceptor moiety at C8 is required for efficient binding of mOgg1. A substituent at C6 and a keto group at C8 are required for cleavage. The proposed mechanism of 8-oxoG excision involves protonation of O 8 or the deoxyribose oxygen moiety. -7,8-dihydro-2Ј-deoxyguanosine (8-oxodG) 1 is found in DNA following oxidative damage mediated by reactive oxygen species (1, 2). In the absence of conformational restraints, including Watson-Crick pairing, 8-oxodG tends to adopt the syn conformation (3, 4). The lactim-lactam equilibrium at the N 7 -O 8 (C 8 ) tautomeric center favors the C8-keto configuration (3, 5), which in syn forms a stable Hoogstein pair with dA (4, 6). As a result of this structural preference, dATP frequently is incorporated opposite template 8-oxodG, and 8-oxodGTP is incorporated opposite template dA during DNA synthesis (7,8). Unrepaired, these mispairs lead, respectively, to G 3 T and A 3 C transversions. 8-OxoIn prokaryotes, several DNA repair enzymes known as the "GO system" prevent mutagenesis via 8-oxoG (9). This system consists of MutT, an 8-oxodGTPase that prevents incorporation of 8-oxodG into DNA from the triphosphate pool (10); Fpg (MutM), an 8-oxoguanine-DNA glycosylase that preferentially excises 8-oxoG paired with C (11); and MutY, an adenine-DNA glycosylase that preferentially excises A paired with 8-oxoG (9), initiating a round of base excision repair that restores the 8-oxoG:C pair, a substrate for Fpg. Eukaryotic homologs have been discovered for the components of the GO system; these include the 8-oxoguanine-DNA glycosylases, Ogg1 and Ogg2, isolated from yeast (12-14). Ogg1 also has been cloned from humans (15-23), mice (18), and rats (24), and a functional analog of yeast Ogg2 was detected in human cells (25). Ogg1 appears to be the primary repair enzyme for 8-oxoG in mammals (26). Interestingly, despite a functional equivalence, eukaryotic 8-oxoguanine-DNA glycos...

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A common mechanism of action for the N-glycosylase activity of DNA N-glycosylase/AP lyases from E. coli and T4

Cited by 28 publications

References 45 publications

Escherichia coli HU protein has a role in the repair of abasic sites in DNA

Escherichia coli HU protein has a role in the repair of abasic sites in DNA

Toward a Detailed Understanding of Base Excision Repair Enzymes: Transition State and Mechanistic Analyses of N-Glycoside Hydrolysis and N-Glycoside Transfer

Substrate Specificity and Reaction Mechanism of Murine 8-Oxoguanine-DNA Glycosylase

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