α‐Ketoglutarate in biological hydroxylations

Holme, Elisabeth; Lindstedt, Göran; Lindstedt, Sven; Tofft, Marianne

doi:10.1016/0014-5793(68)80092-4

Cited by 39 publications

(22 citation statements)

References 19 publications

(1 reference statement)

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“…In the case of some prokaryotic enzymes however, e.g., deacetoxy/deacetyl cephalosporin C synthase and clavaminic acid synthase (at least with some substrates) less than stoichiometric incorporation of oxygen into the alcohol product occurs [9,10]. However, with deacetoxy/deacetyl cephalosporin C synthase, prior work has established that a single oxygen from dioxygen is incorporated into succinate during catalysis [9], as was the case for the more typical reaction catalysed by thymine-7-hydroxylase [11] catalysis. When using (±)-DHQ as a substrate, ANS catalysis also leads to sub-stoichiometric incorporation of oxygen from dioxygen into its flavonoid product, but this reaction is a special case in which the nascent hydroxyl is lost by dehydration [13].…”

Section: Discussionmentioning

confidence: 99%

“…This is thought to be due to solvent exchange of one of the reactive ironoxygen intermediates, although the identity of the particular intermediate(/s) that undergo exchange has not been defined. Results with some eukaryotic 2OG dependent hydroxylases, such as thymine 7-hydroxylase (T7H) [11], mammalian type I prolyl 4-hydroxylase [12], enzymes of flavonoid biosynthesis [13] and human prolyl and asparaginyl hydroxylases involved in hypoxic sensing, PHD1 [14] and FIH [15], show that >90% incorporation of oxygen from dioxygen occurs on hydroxylation of their substrates (in the case of some reactions catalysed by the flavonoid oxygenases exchange may occur after initial hydroxylation by a non-oxidative process [13]). This contrasts with results for eukaryotic lysyl hydroxylase where only approximately 10% of 18 O was reported to be incorporated into the peptide product [16] (data reviewed in [8]).…”

Section: Introductionmentioning

confidence: 99%

“…The enzymes thymine-7-hydroxylase and c-butyrobetaine hydroxylase have been reported to incorporate >95% and 68% 18 O from 18 O 2 into succinate, respectively [11,19]. These data were obtained by mass spectrometric analysis of the succinate co-product derivatised with bis(trimethylsilyl)trifluoroacetamide to give the bis-trimethylsilyl ester.…”

Section: Introductionmentioning

confidence: 99%

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Incorporation of oxygen into the succinate co‐product of iron(II) and 2‐oxoglutarate dependent oxygenases from bacteria, plants and humans

et al. 2005

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The ferrous iron and 2-oxoglutarate (2OG) dependent oxygenases catalyse two electron oxidation reactions by coupling the oxidation of substrate to the oxidative decarboxylation of 2OG, giving succinate and carbon dioxide coproducts. The evidence available on the level of incorporation of one atom from dioxygen into succinate is inconclusive. Here, we demonstrate that five members of the 2OG oxygenase family, AlkB from Escherichia coli, anthocyanidin synthase and flavonol synthase from Arabidopsis thaliana, and prolyl hydroxylase domain enzyme 2 and factor inhibiting hypoxia-inducible factor-1 from Homo sapiens all incorporate a single oxygen atom, almost exclusively derived from dioxygen, into the succinate co-product.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Incorporation of oxygen into the succinate co‐product of iron(II) and 2‐oxoglutarate dependent oxygenases from bacteria, plants and humans

et al. 2005

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“…They are involved in a broad spectrum of primary and secondary biosynthetic pathways including the post-translational hydroxylation of proline and lysine residues in procollagens (Kivirikko et al, 1989), the biosynthesis of carnitine ) and blood-coagulation-factor VII (Stenflo et al, 1989) in mammals, the conversion of thymidine into uracil (Holme et al, 1970(Holme et al, , 1971Bankel et al, 1977) and of penicillins into cephalosporins (Baldwin and Abraham, 1988) in bacteria and fungi, the biosynthesis of clavulanic acid (Elson et al, 1987) and the macrolide antibioticum tylosin (Omura et al, 1984) in fungi, the formation of hydroxyproline-rich glycoproteins in plants (Chrispeels, 1969;Tanaka et al, 1980) and the biosynthesis of plant secondary products such a5 flavonoids Britsch et al, 1981), the alkaloids scopolamine (Hashimot0 and Yamada, 1986) and vindoline (De Carolis et al, 1990) and gibberellins (Hedden and Graebe, 1982).…”

Section: Discussionmentioning

confidence: 99%

Molecular characterization of flayanone 3β‐hydroxylases

Britsch

Dedio²,

Saedler³

et al. 1993

European Journal of Biochemistry

101

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A heterologous cDNA probe from Petunia hybrida was used to isolate flavanone-3P-hydroxylase-encoding cDNA clones from carnation (Dianthus caryophyllus), china aster (Callisteplzus chinensis) and stock (Matthiola incana). The deduced protein sequences together with the known sequences of the enzyme from l ? hybrida, barley (Hordeum vulgare) and snapdragon (Antirrhinum majus) enabled the determination of a consensus sequence which revealed an overall 84% similarity (53 % identity) of flavanone 3P-hydroxylases from the different sources. Alignment with the sequences of other known enzymes of the same class and to related non-heme iron-(11) enzymes demonstrated the strict genetic conservation of 14 anuno acids, in particular, of three histidines and an aspartic acid. The conservation of the histidine motifs provides strong support for the possible conservation of structurally similar iron-binding sites in these enzymes. The putative role of histidines as chelators of ferrous ions in the active site of tlavanone 3P-hydroxylases was corroborated by diethyl-pyrocarbonate modification of the partially purified recombinant Petunia enzyme.Flavanone 3P-hydroxylase (FHT) catalyzes 3P-hydroxylation of 2S-flavanones to 2R,3R-dihydroflavonols [( +)-dihydroflavonols] which are intermediates in the biosynthesis of flavonols, anthocyanidins, catechins and proanthocyanidins in plants (Heller and Forkmann, 1988;Forkmann, 1991). The enzyme from Petunia hybrida has been purified to homogeneity and characterized as a typical 2-oxoglutarate-dependent dioxygenase (Britsch and Grisebach, 1986;Britsch, 1990). Recently, we have isolated a near-full-length FHTencoding cDNA from a Petunia library (Britsch et al., 1992). Heterologous expression of the cDNA in Escherichia coli led to the formation of highly active recombinant FHT, thereby verifying the identity of the cDNA clone.Antibodies against FHT from Petunia readily crossreact with FHT from a number of other sources and were used to characterize specific FHT mutants (Britsch, 1990; Koch, 1992). Therefore, structural similarities of the FHT protein from different plants can be assumed. A comparison of the deduced amino acid sequence of the Petunia FHT with the sequence of FHT from barley (Meldgaard, 1992) indeed showed a high degree of identity (75%; Britsch et al., 1992).Consequently, the FHT cDNA from Petunia should serve as a useful heterologous probe to clone this enzyme from other plant sources. Utilizing the 945-bp EcoRI fragment from Petunia FHT as a hybridization probe, we were able to clone the corresponding cDNAs from carnation, china aster and stock. In this study, we report the cloning and sequence analysis of these particular FHT enzymes, the analysis of the consensus sequence derived from six individual FHT sequences and the modification of purified recombinant enzyme with diethyl pyrocarbonate, performed to elucidate the possible function of strictly conserved histidines. Furthermore, the sequences were also compared with other known 2-oxoglutarate-dependent dioxygenases f...

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“…One atom of molecular oxygen is incorporated into succinate and the other into the primary products. [15][16][17] The Rhodotorula glutinis T7H sequence includes His 211, Asp 213, and His 268 as putative metal-binding ligands and Arg 284 as a likely αKG stabilizing residue, but no structural studies have been carried out to confirm these assignments.Efforts to characterize T7H have focused on the enzymes from fungi such as Neurospora crassa, R. glutinis, and Aspergillus nidulans that can grow on thymine as their sole nitrogen source. These microorganisms possess a salvage pathway enabling the host to utilize thymine as a pyrimidine source in the absence of the common "de novo" pathway of uracil biosynthesis (Fig.…”

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confidence: 99%

FeII/α-ketoglutarate hydroxylases involved in nucleobase, nucleoside, nucleotide, and chromatin metabolism

2008

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Fe II /α-ketoglutarate-dependent hydroxylases uniformly possess a double-stranded β-helix fold with two conserved histidines and one carboxylate coordinating their mononuclear ferrous ions. Oxidative decomposition of the α-keto acid is proposed to generate a ferryl-oxo intermediate capable of hydroxylating unactivated carbon atoms in a myriad of substrates. This perspective focuses on a subgroup of these enzymes that are involved in pyrimidine salvage, purine decomposition, nucleoside and nucleotide hydroxylation, DNA/RNA repair, and chromatin modification. The varied reaction schemes are presented, and selected structural and kinetic information is summarized. IntroductionFerrous ion and α-ketoglutarate (αKG)-dependent dioxygenases comprise an enzyme superfamily with members present in animals, plants, protists, fungi, bacteria and even viruses. Most representatives of this large group of enzymes couple the oxidative decarboxylation of αKG to various hydroxylation reactions ( Fig. 1), but others are capable of catalyzing desaturation, epoxidation, halogenation, ring formation, or ring expansion reactions, which allows them to fill a wide variety of biological roles including antibiotic biosynthesis, hypoxic sensing, DNA repair, and various types of metabolite transformations. [1][2][3] Not surprisingly, the primary substrates also are diverse and include proteins, lipids, nucleic acids, and a myriad of small molecules that can be used for synthesis or undergo degradation.The sequences of Fe II /αKG dioxygenases show little overall identity, yet the diverse reactions all are catalyzed by proteins with the same core fold and use similar chemistries. 4 The hallmark of these enzymes is their use of Fe II to activate molecular oxygen according to a mechanism (Scheme 1) first proposed by Hanauske-Abel and Günzler in 1982 for prolyl 4-hydroxylase. 5 (A) In the absence of substrates, a "facial triad" (usually two His plus one Asp or Glu occurring in a His-X-Asp/Glu-X n -His motif) weakly bind one face of the hexacoordinate metal. 6,7 (B) The αKG co-substrate displaces two waters and coordinates Fe II in a bidentate fashion with its C1 carboxyl and C2 keto oxygens. The binding of αKG is Correspondence to: Robert P. Hausinger, hausinge@msu.edu. NIH Public Access Author ManuscriptDalton Trans. Author manuscript; available in PMC 2010 July 20. This perspective focuses on the modification of nucleobases, nucleosides, nucleotides, and chromatin by the Fe II and αKG dependent hydroxylases (Table 1). We discuss the degradation and salvage of free bases by the fungal enzymes thymine 7-hydroxylase and xanthine hydroxylase. Next, nucleoside hydroxylases are briefly described. We then examine the developmental modification of DNA in kinetoplastid protozoans and summarize evidence that the JBP1 and JBP2 proteins possess thymidine hydroxylase activity. The oxidative repair of DNA by Escherichia coli AlkB is discussed, and the properties of mammalian homologues are summarized. Finally, we present emerging evidence pointing to...

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α‐Ketoglutarate in biological hydroxylations

Cited by 39 publications

References 19 publications

Incorporation of oxygen into the succinate co‐product of iron(II) and 2‐oxoglutarate dependent oxygenases from bacteria, plants and humans

Incorporation of oxygen into the succinate co‐product of iron(II) and 2‐oxoglutarate dependent oxygenases from bacteria, plants and humans

Molecular characterization of flayanone 3β‐hydroxylases

FeII/α-ketoglutarate hydroxylases involved in nucleobase, nucleoside, nucleotide, and chromatin metabolism

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