Trapping and spectroscopic characterization of an Fe<sup>III</sup>-superoxo intermediate from a nonheme mononuclear iron-containing enzyme

Mbughuni, Michael M.; Chakrabarti, Mrinmoy; Hayden, Joshua A.; Bominaar, Emile L.; Hendrich, Michael P.; Münck, Eckard; Lipscomb, John D.

doi:10.1073/pnas.1010015107

Cited by 144 publications

(325 citation statements)

References 45 publications

(57 reference statements)

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“…Ringcleaving dioxygenases active toward these noncatecholic compounds belong to the cupin superfamily and utilize a mononuclear Fe II center for catalysis. Their catalytic strategy has been proposed to involve a one-electron transfer to dioxygen, possibly via transient formation of an Fe III -O 2 · Ϫ intermediate, as observed in type I extradiol dioxygenases (83,84), which are members of the VOC superfamily (5). However, it should be emphasized that compared to the thoroughly studied extradiol dioxygenase reaction mechanism, much less experimental evidence is available on the catalytic mechanism of the cupin-type ring-cleaving dioxygenases.…”

Section: Discussionmentioning

confidence: 99%

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Ring-Cleaving Dioxygenases with a Cupin Fold

Fetzner

2012

Appl Environ Microbiol

166

141

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ABSTRACTRing-cleaving dioxygenases catalyze key reactions in the aerobic microbial degradation of aromatic compounds. Many pathways converge to catecholic intermediates, which are subject toorthoormetacleavage by intradiol or extradiol dioxygenases, respectively. However, a number of degradation pathways proceed via noncatecholic hydroxy-substituted aromatic carboxylic acids like gentisate, salicylate, 1-hydroxy-2-naphthoate, or aminohydroxybenzoates. The ring-cleaving dioxygenases active toward these compounds belong to the cupin superfamily, which is characterized by a six-stranded β-barrel fold and conserved amino acid motifs that provide the 3His or 2- or 3His-1Glu ligand environment of a divalent metal ion. Most cupin-type ring cleavage dioxygenases use an FeIIcenter for catalysis, and the proposed mechanism is very similar to that of the canonical (type I) extradiol dioxygenases. The metal ion is presumed to act as an electron conduit for single electron transfer from the metal-bound substrate anion to O2, resulting in activation of both substrates to radical species. The family of cupin-type dioxygenases also involves quercetinase (flavonol 2,4-dioxygenase), which opens up two C-C bonds of the heterocyclic ring of quercetin, a wide-spread plant flavonol. Remarkably, bacterial quercetinases are capable of using different divalent metal ions for catalysis, suggesting that the redox properties of the metal are relatively unimportant for the catalytic reaction. The major role of the active-site metal ion could be to correctly position the substrate and to stabilize transition states and intermediates rather than to mediate electron transfer. The tentative hypothesis that quercetinase catalysis involves direct electron transfer from metal-bound flavonolate to O2is supported by model chemistry.

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Section: Discussionmentioning

confidence: 99%

“…Subsequent rearrangement and O-O bond cleavage gives a seven-membered lactone intermediate and an Fe II -bound hydroxide ion, which hydrolyzes the lactone to yield the 2-hydroxymuconate semialdehyde product (Fig. 1b) (44,71,72,78,83,84).…”

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confidence: 99%

Ring-Cleaving Dioxygenases with a Cupin Fold

Fetzner

2012

Appl Environ Microbiol

166

141

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“…The Fe 3+ in the active site of IDOs will not bind O 2 and no direct spectroscopic evidence for redox cycling has been obtained (1). This means that IDOs must activate O 2 using a unique mechanism, which differs from that of the much more common Fe 2+ containing oxygenases that readily form an Fe-O 2 complex after the substrate binds in the active site (7)(8)(9).…”

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confidence: 99%

Crystal structures of alkylperoxo and anhydride intermediates in an intradiol ring-cleaving dioxygenase

Knoot

Purpero

Lipscomb

2014

Proc. Natl. Acad. Sci. U.S.A.

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Intradiol aromatic ring-cleaving dioxygenases use an active site, nonheme Fe 3+ to activate O 2 and catecholic substrates for reaction. The inability of Fe 3+ to directly bind O 2 presents a mechanistic conundrum. The reaction mechanism of protocatechuate 3,4-dioxygenase is investigated here using the alternative substrate 4-fluorocatechol. This substrate is found to slow the reaction at several steps throughout the mechanistic cycle, allowing the intermediates to be detected in solution studies. When the reaction was initiated in an enzyme crystal, it was found to halt at one of two intermediates depending on the pH of the surrounding solution. The X-ray crystal structure of the intermediate at pH 6.5 revealed the key alkylperoxo-Fe 3+ species, and the anhydride-Fe 3+ intermediate was found for a crystal reacted at pH 8.5. Intermediates of these types have not been structurally characterized for intradiol dioxygenases, and they validate four decades of spectroscopic, kinetic, and computational studies. In contrast to our similar in crystallo crystallographic studies of an Fe 2+ -containing extradiol dioxygenase, no evidence for a superoxo or peroxo intermediate preceding the alkylperoxo was found. This observation and the lack of spectroscopic evidence for an Fe 2+ intermediate that could bind O 2 are consistent with concerted formation of the alkylperoxo followed by Criegee rearrangement to yield the anhydride and ultimately ringopened product. Structural comparison of the alkylperoxo intermediates from the intra-and extradiol dioxygenases provides a rationale for site specificity of ring cleavage.dioxygenase | oxygen activation | X-ray crystallography | reaction intermediate | Fe (III)

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“…EPR and Mossbauer spectroscopy characterization of both the Fe-and Mn-substituted 2,3-HPCD provide evidence of M III -radical intermediates, which suggests sequential oxidation and reduction of the metal cofactor may take place during such dioxygenase activity (48,49). Fig.…”

Section: Discussionmentioning

confidence: 99%

“…7 display possible reaction pathways for both the dioxygenase and nitroxygenase activities of Mn-QDO. By analogy to the 2,3-HPCD mechanistic studies (47)(48)(49), the Mn-QDO dioxygenase reaction is proposed to go through an initial tertiary complex, A, in which an initial SET has yielded an Mn III -superoxide species. A second SET from substrate to metal allows subsequent radical coupling between the metal-bound superoxide and substrate.…”

Section: Discussionmentioning

confidence: 99%

Nitrosyl hydride (HNO) replaces dioxygen in nitroxygenase activity of manganese quercetin dioxygenase

Kumar

Zapata

Ramirez

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

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Quercetin dioxygenase (QDO) catalyzes the oxidation of the flavonol quercetin with dioxygen, cleaving the central heterocyclic ring and releasing CO. The QDO from Bacillus subtilis is unusual in that it has been shown to be active with several divalent metal cofactors such as Fe, Mn, and Co. Previous comparison of the catalytic activities suggest that Mn(II) is the preferred cofactor for this enzyme. We herein report the unprecedented substitution of nitrosyl hydride (HNO) for dioxygen in the activity of Mn-QDO, resulting in the incorporation of both N and O atoms into the product. Turnover is demonstrated by consumption of quercetin and other related substrates under anaerobic conditions in the presence of HNO-releasing compounds and the enzyme. As with dioxygenase activity, a nonenzymatic base-catalyzed reaction of quercetin with HNO is observed above pH 7, but no enhancement of this basal reactivity is found upon addition of divalent metal salts. Unique and regioselective N-containing products ( 14 N∕ 15 N) have been characterized by MS analysis for both the enzymatic and nonenzymatic reactions. Of the several metallo-QDO enzymes examined for nitroxygenase activity under anaerobic condition, only the Mn(II) is active; the Fe(II) and Co(II) substituted enzymes show little or no activity. This result represents an enzymatic catalysis which we denote nitroxygenase activity; the unique reactivity of the Mn-QDO suggests a metalmediated electron transfer mechanism rather than metal activation of the substrate's inherent base-catalyzed reactivity.nitroxyl | quercetinase D ioxygenases are enzymes that incorporate both atoms of dioxygen into a targeted substrate, e.g., the enzyme catechol 1,2-dioxygenase cleaves the C-C bond between catechol's two hydroxyl groups forming the dicarboxylate cis,cis-muconic acid (1). Although highly divergent, there are two broadly defined classes of nonheme dioxygenases: those that directly bind dioxygen at the metal center or those in which dioxygen binds to the substrate that has been activated by interaction with the metal center (2). Such questions also apply to quercetin dioxygenase (QDO), which catalyzes the oxidation of the flavonol quercetin (1) with dioxygen ( Fig. 1). During turnover, QDO cleaves the central heterocyclic ring of quercetin, releasing CO, and yielding the corresponding depside. The QDO from Bacillus subtilis is unusual in that it has been shown to be active with several divalent metal cofactors such as Cu, Fe, Mn, and Co. Comparison of the catalytic activities suggest that Mn(II) is the preferred cofactor for this enzyme (3).The simple molecule nitrosyl hydride (HNO) is the singly reduced and protonated form of nitric oxide, NO. HNO has garnered much interest because it demonstrates a physiological reactivity distinctly different from its congener nitric oxide (4, 5). In a series of papers, we have reported the preparation and characterization of the HNO adduct of myoglobin (HNO-Mb) by reduction of 7) and by trapping of free HNO by deoxymyoglobin (Mb-Fe II ) (8)...

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Trapping and spectroscopic characterization of an Fe^III-superoxo intermediate from a nonheme mononuclear iron-containing enzyme

Cited by 144 publications

References 45 publications

Ring-Cleaving Dioxygenases with a Cupin Fold

Ring-Cleaving Dioxygenases with a Cupin Fold

Crystal structures of alkylperoxo and anhydride intermediates in an intradiol ring-cleaving dioxygenase

Nitrosyl hydride (HNO) replaces dioxygen in nitroxygenase activity of manganese quercetin dioxygenase

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Trapping and spectroscopic characterization of an FeIII-superoxo intermediate from a nonheme mononuclear iron-containing enzyme

Cited by 144 publications

References 45 publications

Ring-Cleaving Dioxygenases with a Cupin Fold

Ring-Cleaving Dioxygenases with a Cupin Fold

Crystal structures of alkylperoxo and anhydride intermediates in an intradiol ring-cleaving dioxygenase

Nitrosyl hydride (HNO) replaces dioxygen in nitroxygenase activity of manganese quercetin dioxygenase

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Trapping and spectroscopic characterization of an Fe^III-superoxo intermediate from a nonheme mononuclear iron-containing enzyme