Aberrant activity of the DNA repair enzyme AlkB

Henshaw, Timothy F.; Feig, Michael; Hausinger, Robert P.

doi:10.1016/j.jinorgbio.2003.10.021

Cited by 52 publications

(61 citation statements)

References 34 publications

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“…This peak was attributed to the LMCT transition of OH-Trp coordinated to Fe III . The hydroxylation of the Trp side chain of AlkB, identified to be Trp 178 , suggests formation of an oxidative intermediate in AlkB [45]. This behavior is consistent with other members of the non-heme Fe II /αKG-dependent family [48][49][50].…”

Section: Repair Mechanismsupporting

confidence: 80%

“…The mononuclear Fe II center of AlkB with bound αKG has been confirmed both with the overexpression and isolation of the native protein directly from E. coli [41] and by the addition of excess Fe II and αKG to apo-AlkB [30][31][32]38,[42][43][44][45][46]. Studies of the metal center has lead to an observation of an UV-vis band at 560 nm for the native protein [41].…”

Section: The Mononuclear Iron Centermentioning

confidence: 89%

“…Addition of O 2 saturated H 2 O to iron-loaded AlkB led to the development of a visible chromophore centered at 595 nm (ε 590 960 M −1 cm −1 ) [45]. This peak was attributed to the LMCT transition of OH-Trp coordinated to Fe III .…”

Section: Repair Mechanismmentioning

confidence: 99%

“…TauD has been a preferred enzyme for study in this family due to its simplicity of substrate and its ease with which it can be prepared in large quantity, as well as the characterization of its crystal structure. The uncoupled reaction, leading to irreversible modifications of AlkB [45], was demonstrated as αKG decomposition was found to be partially uncoupled from DNA repair, accounting for the observed stimulation of AlkB activity in the presence of ascorbate [31,44]. The uncoupled αKG turnover was also stimulated by binding of the small molecules 1-meA, 1-methyldeoxyadenosine (1-me(dA)), 3-meC and 3-methyldeoxycytidine (3-me(dC)), but they were not repaired by AlkB [44].…”

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confidence: 99%

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Oxidative dealkylation DNA repair mediated by the mononuclear non-heme iron AlkB proteins

Mishina

2006

Journal of Inorganic Biochemistry

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DNA can be damaged by various intracellular and environmental alkylating agents to produce alkylation base lesions. These base damages, if not repaired promptly, may cause genetic changes that lead to diseases such as cancer. Recently, it was discovered that some of the alkylation DNA base damage can be directly removed by a family of proteins called the AlkB proteins that utilize a mononuclear non-heme iron(II) and α-ketoglutarate as cofactor and cosubstrate. These proteins activate dioxygen and perform an unprecedented oxidative deal-kylation of the alkyl adducts on DNA heteroatoms. This review summarizes the discovery of this activity and the recent research advances in studying this unique DNA repair pathway. The focus is placed on the chemical mechanism and function of these proteins. KeywordsDNA repair; AlkB; ABH; Direct repair; Mononuclear non-heme iron; Dioxygenase; Oxidative dealkylation; Demethylation Direct repair of alkylation DNA damageCellular DNA is subject to nonenzymatic alkylation (methylation) by environmental or chemical mutagens, resulting in adducts that are toxic and mutagenic [1][2][3][4][5][6]. Organisms have evolved a variety of mechanisms to repair these mutagenic damages. Escherichia coli (E. coli) responds to this threat by activating an adaptive responsive pathway, mediated by the E. coli Ada protein [7]. Upon receiving a methyl group from the damaged DNA, Ada turns into a transcriptional activator and activates its own expression and that of the alkB gene and two other genes, alkA and aidB ( Fig. 1(a)) [4,7,8]. The upregulated Ada is a bifunctional protein, which uses a N-terminal Cys38 residue to remove a methyl group fromS p -methylphosphotriester and a C-terminal Cys321 residue to remove a methyl adduct from O 6 -methylguanaine ( Fig. 1(b)) [9][10][11]. Both repair functions are irreversible and represent rare examples of direct repair of DNA damage. AlkA is a glycosylase that cleaves methylated bases from DNA and performs the first step of the well-known base excision repair of base lesions [12][13][14]. The precise function of AidB is still unknown. The function of the last member of this adaptive response, AlkB, also remained unknown until recently despite extensive research efforts. E. coli AlkBSince the first genetic study in 1983 isolating the E. coli mutant with specific sensitivity to the alkylating agent methylmethane sulfonate, MMS [15], the activity of AlkB was undisclosed The Fe II /αKG-dependent dioxygenase superfamily is widespread from bacteria to humans [19] and is the largest known non-heme iron protein family [20][21][22][23]. It represents a wide diversity of enzymes that catalyze the hydroxylation of unactivated C-H groups of a variety of substrates by coupling reductive activation of dioxygen with a decarbox-ylation of αKG, the co-substrate, to succinate. In the event of oxidation one of the oxygen atoms from O 2 is incorporated into the succinate and the other as a hydroxyl in the product [24]. This group of enzymes is known for their importance in ...

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Section: Repair Mechanismsupporting

confidence: 80%

Section: The Mononuclear Iron Centermentioning

confidence: 89%

Section: Repair Mechanismmentioning

confidence: 99%

mentioning

confidence: 99%

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Oxidative dealkylation DNA repair mediated by the mononuclear non-heme iron AlkB proteins

Mishina

2006

Journal of Inorganic Biochemistry

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“…During the uncoupled reaction of TauD, ϳ60% of the inactivated complexes can be recovered by reduction, whereas approximately one-third of the molecules are transformed into a Fe(III)-succinate-catecholate species (15) that are produced by enzyme self-hydroxylation at the side chain of Tyr 73 . For TfdA and AlkB, analogous hydroxylation of a tryptophan side chain has been observed (16,17). The precise location of the relevant hydroxylated aromatic residue within the protein framework varies among enzymes.…”

mentioning

confidence: 98%

Succinate Complex Crystal Structures of the α-Ketoglutarate-dependent Dioxygenase AtsK

Müller

Stückl

Wakeley

et al. 2005

Journal of Biological Chemistry

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The alkylsulfatase AtsK from Pseudomonas putida S-313 is a member of the non-heme iron(II)-␣-ketoglutarate-dependent dioxygenase superfamily. In the initial step of their catalytic cycle, enzymes belonging to this widespread and versatile family coordinate molecular oxygen to the iron center in the active site. The subsequent decarboxylation of the cosubstrate ␣-ketoglutarate yields carbon dioxide, succinate, and a highly reactive ferryl (IV) species, which is required for substrate oxidation via a complex mechanism involving the transfer of radical species. Non-productive activation of oxygen may lead to harmful side reactions; therefore, such enzymes need an effective built-in protection mechanism. One of the ways of controlling undesired side reactions is the self-hydroxylation of an aromatic side chain, which leads to an irreversibly inactivated species. Here we describe the crystal structure of the alkylsulfatase AtsK in complexes with succinate and with Fe(II)/succinate. In the crystal structure of the AtsKFe(II)-succinate complex, the side chain of Tyr 168 is coordinated to the iron, suggesting that Tyr 168 is the target of enzyme self-hydroxylation. This is the first structural study of an Fe(II)-␣-ketoglutarate-dependent dioxygenase that presents an aromatic side chain coordinated to the metal center, thus allowing structural insight into this protective mechanism of enzyme self-inactivation.The alkylsulfatase AtsK from Pseudomonas putida S-313 is a member of the family of non-heme iron(II), ␣-ketoglutarate-dependent dioxygenases (1, 2). This remarkable superfamily is widespread among prokaryotic and eukaryotic organisms, and its members catalyze a broad diversity of energetically demanding biosynthetic and degradative reactions including hydroxylation processes, stereoselective desaturation of inactivated carbon-carbon single bonds, and oxidative ring closure (3). The catalytic mechanism of this enzyme superfamily has been the topic of numerous spectroscopic and structural investigations (Fig. 1). This family of enzymes all bind iron using a 2-His-1-carboxylate facial triad, a feature that is one of nature's recurring multifunctional bioinorganic motifs such as the heme group or iron-sulfur clusters. They catalyze the oxidative conversion of the cofactor ␣KG 1 into CO 2 and succinate, simultaneously generating an activated oxygen species bound to the oxidized Fe center.The generally accepted mechanism proceeds via a Fe(IV)-oxo intermediate (4). The interaction of this activated intermediate with the enzyme substrate generates a transient radical species that subsequently decomposes to the reaction product. A reaction mechanism that involves radical formation would at first sight seem to be a hazardous method for an organism to accomplish the relatively straightforward cleavage reaction that is catalyzed by AtsK (i.e. the release of sulfate from an alkyl sulfate ester), because harmful side reactions could take place in the absence of substrate. Such enzymes must therefore have an effective built-in pro...

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Oxidative Demethylation of DNA and RNA Mediated by Non‐Heme Iron‐Dependent Dioxygenases

Zhu

Xia

et al. 2014

Chemistry An Asian Journal

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DNA/RNA methylation can be generated by methyltransferases and thus plays a critical role in regulating cellular processes; alternatively, nucleic acid methylation can be produced by methylation agents and is cytotoxic/mutagenic if left unrepaired. Oxidative demethylation mediated by non-heme iron-dependent dioxygenases is an efficient way to reverse either the cellular roles of regulatory methylation or the cytotoxic/mutagenic effects of methylation damage. In this Focus Review we summarize recent advances in the study of nucleic acid dioxygenases exemplified by the TET and AlkB family proteins, with an emphasis on chemical insights from the recent literature. Comparison of the chemical mechanisms of these dioxygenases revealed that differences in the mechanism also contribute significantly to their distinct biological functions.

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Aberrant activity of the DNA repair enzyme AlkB

Cited by 52 publications

References 34 publications

Oxidative dealkylation DNA repair mediated by the mononuclear non-heme iron AlkB proteins

Oxidative dealkylation DNA repair mediated by the mononuclear non-heme iron AlkB proteins

Succinate Complex Crystal Structures of the α-Ketoglutarate-dependent Dioxygenase AtsK

Oxidative Demethylation of DNA and RNA Mediated by Non‐Heme Iron‐Dependent Dioxygenases

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