The Mononuclear Molybdenum Enzymes

Hille, Russ

doi:10.1021/cr950061t

Cited by 1,482 publications

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References 424 publications

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“…Furthermore, EPR experiments using cyanide with a different isotope composition suggest that cyanide is not coordinated to Mo(V) ions. Mo(V) EPR signals having nearly axial symmetry have been observed in several Mo enzymes with well-documented crystal structures, such as the members of the XO family [4] and formatereduced Ec Fdh-H [34]. The crystal structures of these proteins show Mo sites in square pyramidal coordination, indicating that the Mo(V) site of the cyanide species in Dd NapA is more compatible with this coordination than distorted hexa coordination (Scheme 1).…”

Section: Discussionmentioning

confidence: 97%

“…Most NRs are mononuclear molybdenum-containing enzymes present in several living organisms which have, in addition to a molybdenum active site, additional redox cofactors such as iron-sulfur and heme centers that mediate electron transfer reactions between the electron donor and the nitrate [4]. NRs have been classified into four groups according to different Electronic Supplementary Material Supplementary material is available for this article at http://dx.doi.org/10.1007/s00775-006-0110-0 and is accessible for authorized users.…”

Section: Introductionmentioning

confidence: 99%

“…Nas, Nar, and Nap, which are only found in prokaryotic organisms, belong to the dimethyl sulfoxide reductase family, whereas Euk-NRs belong to the sulfite oxidase family of molybdo proteins [4]. Euk-NR and Nas are cytoplasmic enzymes involved in nitrate assimilation, whereas Nar are membranebound enzymes involved exclusively in denitrification.…”

Section: Introductionmentioning

confidence: 99%

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EPR and redox properties of periplasmic nitrate reductase from Desulfovibrio desulfuricans ATCC 27774

et al. 2006

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Nitrate reductases are enzymes that catalyze the conversion of nitrate to nitrite. We report here electron paramagnetic resonance (EPR) studies in the periplasmic nitrate reductase isolated from the sulfatereducing bacteria Desulfovibrio desulfuricans ATCC 27774. This protein, belonging to the dimethyl sulfoxide reductase family of mononuclear Mo-containing enzymes, comprises a single 80-kDa subunit and contains a Mo bis(molybdopterin guanosine dinucleotide) cofactor and a [4Fe-4S] cluster. EPR-monitored redox titrations, carried out with and without nitrate in the potential range from 200 to À500 mV, and EPR studies of the enzyme, in both catalytic and inhibited conditions, reveal distinct types of Mo(V) EPR-active species, which indicates that the Mo site presents high coordination flexibility. These studies show that nitrate modulates the redox properties of the Mo active site, but not those of the [4Fe-4S] center. The possible structures and the role in catalysis of the distinct Mo(V) species detected by EPR are discussed.

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Section: Discussionmentioning

confidence: 97%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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EPR and redox properties of periplasmic nitrate reductase from Desulfovibrio desulfuricans ATCC 27774

et al. 2006

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“…Replacement of the product (sulfate) by water or hydroxide (B→C) and subsequent oneelectron oxidations (Mo(IV/V/VI)) and concomitant deprotonations (C→D→A) return the Mo center to the fully oxidized resting state. [6][7][8] Previous steady-state kinetic studies of the R160Q hSO mutant showed a nearly 1000-fold decrease in k cat /K m sulfite , which suggested that the positive charge on Arg 160 in hSO makes an important contribution to the binding of sulfite. 2 Flash photolysis studies of intramolecular electron transfer (IET) between the molybdenum and heme domains showed that IET was also reduced about 1000-fold in R160Q compared to wild-type hSO.…”

Section: Introductionmentioning

confidence: 98%

Structural Studies of the Molybdenum Center of the Pathogenic R160Q Mutant of Human Sulfite Oxidase by Pulsed EPR Spectroscopy and ¹⁷O and ³³S Labeling

et al. 2008

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Electron paramagnetic resonance (EPR) investigation of the Mo(V) center of the pathogenic R160Q mutant of human sulfite oxidase (hSO) confirms the presence of three distinct species whose relative abundances depend upon pH. Species 1 is exclusively present at pH ≤ 6, and remains in significant amounts even at pH 8. Variable-frequency electron spin echo envelope modulation (ESEEM) studies of this species prepared with 33 S-labeled sulfite clearly show the presence of coordinated sulfate, as has previously been found for the "blocked" form of Arabidopsis thaliana at low pH (Astashkin, A. V.; Johnson-Winters, K.; Klein, E. L.; Byrne, R. S.; Hille, R.; Raitsimring, A. M.; Enemark, J. H. J. Am. Chem. Soc. 2007, 129, 14800). The ESEEM spectra of Species 1 prepared in 17 O-enriched water show both strongly and weakly magnetically coupled 17 O atoms that can be assigned to an equatorial sulfate ligand and the axial oxo ligand, respectively. The nuclear quadrupole interaction (nqi) of the axial oxo ligand is substantially stronger than those found for other oxo-Mo(V) centers studied previously. Additionally, pulsed electron-nuclear double resonance (ENDOR) measurements reveal a nearby weakly coupled exchangeable proton. The structure for Species 1 proposed from the pulsed EPR results using isotopic labeling is a six-coordinate Mo(V) center with an equatorial sulfate ligand that is hydrogen bonded to an exchangeable proton. Six-coordination is supported by the 17 O nqi parameters for the axial oxo group of the model compound, (dttd)Mo 17 O( 17 Otms), where H 2 dttd = 2,3:8,9-dibenzo-1,4,7,10-tetrathiadecane; tms = trimethylsilyl. Reduction of R160Q to Mo(V) with Ti(III) gives primarily Species 2, another low pH form, whereas reduction with sulfite at higher pH values gives a mixture of Species 1 and 2, as well as the "primary" high pH form of wild-type SO. The occurrence of significant amounts of the "sulfate-blocked" form of R160Q (Species 1) at physiological pH suggests that this species may be a contributing factor to the lethality of this mutation.

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“…AO is structurally very similar to XOR, containing one FAD, two nonidentical [2Fe-2S] centres and one molybdopterin, in a homodimeric structure. Both enzymes have broad and overlapping specificity for reducing substrates but, unlike XOR, AO cannot be converted to a dehydrogenase form, being unable to utilise NAD + as the oxidising substrate [28].…”

Section: Introductionmentioning

confidence: 99%

NADH oxidase activity of rat and human liver xanthine oxidoreductase: potential role in superoxide production

et al. 2007

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To characterise the NADH oxidase activity of both xanthine dehydrogenase (XD) and xanthine oxidase (XO) forms of rat liver xanthine oxidoreductase (XOR) and to evaluate the potential role of this mammalian enzyme as an O 2•-source, kinetics and electron paramagnetic resonance (EPR) spectroscopic studies were performed. A steady-state kinetics study of XD showed that it catalyses NADH oxidation, leading to the formation of one O 2•-molecule and half a H 2 O 2 molecule per NADH molecule, at rates 3 times those observed for XO (29.2 ± 1.6 and 9.38 ± 0.31 min -1 , respectively). EPR spectra of NADHreduced XD and XO were qualitatively similar, but they were quantitatively quite different. While NADH efficiently reduced XD, only a great excess of NADH reduced XO. In agreement with reductive titration data, the XD specificity constant for NADH (8.73 ± 1.36 lM -1 min -1 ) was found to be higher than that of the XO specificity constant (1.07 ± 0.09 lM -1 min -1 ). It was confirmed that, for the reducing substrate xanthine, rat liver XD is also a better O 2•-source than XO. These data show that the dehydrogenase form of liver XOR is, thus, intrinsically more efficient at generating O 2•-than the oxidase form, independently of the reducing substrate. Most importantly, for comparative purposes, human liver XO activity towards NADH oxidation was also studied, and the kinetics parameters obtained were found to be very similar to those of the XO form of rat liver XOR, foreseeing potential applications of rat liver XOR as a model of the human liver enzyme.Keywords Xanthine oxidoreductase Á Xanthine oxidase Á Xanthine dehydrogenase Á Rat liver Á Reactive oxygen species Á NADH Abbreviations AFR Activity-to-flavin ratio AO Aldehyde oxidase EPR Electron paramagnetic resonance FAD Flavin adenine dinucleotide ROS Reactive oxygen species Sim Simulated XD Xanthine dehydrogenase XO Xanthine oxidase XOR Xanthine oxidoreductase

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The Mononuclear Molybdenum Enzymes

Cited by 1,482 publications

References 424 publications

EPR and redox properties of periplasmic nitrate reductase from Desulfovibrio desulfuricans ATCC 27774

EPR and redox properties of periplasmic nitrate reductase from Desulfovibrio desulfuricans ATCC 27774

Structural Studies of the Molybdenum Center of the Pathogenic R160Q Mutant of Human Sulfite Oxidase by Pulsed EPR Spectroscopy and ¹⁷O and ³³S Labeling

NADH oxidase activity of rat and human liver xanthine oxidoreductase: potential role in superoxide production

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The Mononuclear Molybdenum Enzymes

Cited by 1,482 publications

References 424 publications

EPR and redox properties of periplasmic nitrate reductase from Desulfovibrio desulfuricans ATCC 27774

EPR and redox properties of periplasmic nitrate reductase from Desulfovibrio desulfuricans ATCC 27774

Structural Studies of the Molybdenum Center of the Pathogenic R160Q Mutant of Human Sulfite Oxidase by Pulsed EPR Spectroscopy and 17O and 33S Labeling

NADH oxidase activity of rat and human liver xanthine oxidoreductase: potential role in superoxide production

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Structural Studies of the Molybdenum Center of the Pathogenic R160Q Mutant of Human Sulfite Oxidase by Pulsed EPR Spectroscopy and ¹⁷O and ³³S Labeling