α-Amine Desaturation of d -Arginine by the Iron(II)- and 2-(Oxo)glutarate-Dependent l -Arginine 3-Hydroxylase, VioC

Noah P. Dunham, Andrew J. Mitchell, José M. Del Río Pantoja, Carsten Krebs, Joseph M. Bollinger, Jr., Amie Kathleen Boal

Research output: Contribution to journalArticle

5 Citations (Scopus)

Abstract

When challenged with substrate analogues, iron(II)- and 2-(oxo)glutarate-dependent (Fe/2OG) oxygenases can promote transformations different from those they enact upon their native substrates. We show here that the Fe/2OG enzyme, VioC, which is natively an l-arginine 3-hydroxylase, catalyzes an efficient oxidative deamination of its substrate enantiomer, d-Arg. The reactant complex with d-Arg retains all interactions between enzyme and substrate functional groups, but the required structural adjustments and opposite configuration of C2 position this carbon more optimally than C3 to donate hydrogen (H) to the ferryl intermediate. The simplest possible mechanism, C2 hydroxylation followed by elimination of ammonia, is inconsistent with the demonstrated solvent origin of the ketone oxygen in the product. Rather, the reaction proceeds via a hydrolytically labile C2-iminium intermediate, demonstrated by its reductive trapping in solution with NaB2H4 to produce racemic [2H]Arg. Of two alternative pathways to the iminium species, C2 hydroxylation followed by dehydration versus direct desaturation, the latter possibility appears to be more likely, because the former mechanism would be expected to result in detectable incorporation of 18O from 18O2. The direct desaturation of a C-N bond implied by this analysis is analogous to that recently posited for the reaction of the l-Arg 4,5-desaturase, NapI, thus lending credence to the prior mechanistic proposal. Such a pathway could also potentially be operant in a subset of reactions catalyzed by Fe/2OG N-demethylases, which have instead been purported to enact C-N bond cleavage by methyl hydroxylation and elimination of formaldehyde.

Original languageEnglish (US)
Pages (from-to)6479-6488
Number of pages10
JournalBiochemistry
Volume57
Issue number46
DOIs
StatePublished - Nov 20 2018

Fingerprint

Glutarates
Hydroxylation
Mixed Function Oxygenases
Amines
Arginine
Iron
Substrates
N Demethylating Oxidoreductases
Oxygenases
Deamination
Enzymes
Ketones
Dehydration
Ammonia
Formaldehyde
Enantiomers
Hydrogen
Carbon
Functional groups
Oxygen

All Science Journal Classification (ASJC) codes

  • Biochemistry

Cite this

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title = "α-Amine Desaturation of d -Arginine by the Iron(II)- and 2-(Oxo)glutarate-Dependent l -Arginine 3-Hydroxylase, VioC",
abstract = "When challenged with substrate analogues, iron(II)- and 2-(oxo)glutarate-dependent (Fe/2OG) oxygenases can promote transformations different from those they enact upon their native substrates. We show here that the Fe/2OG enzyme, VioC, which is natively an l-arginine 3-hydroxylase, catalyzes an efficient oxidative deamination of its substrate enantiomer, d-Arg. The reactant complex with d-Arg retains all interactions between enzyme and substrate functional groups, but the required structural adjustments and opposite configuration of C2 position this carbon more optimally than C3 to donate hydrogen (H•) to the ferryl intermediate. The simplest possible mechanism, C2 hydroxylation followed by elimination of ammonia, is inconsistent with the demonstrated solvent origin of the ketone oxygen in the product. Rather, the reaction proceeds via a hydrolytically labile C2-iminium intermediate, demonstrated by its reductive trapping in solution with NaB2H4 to produce racemic [2H]Arg. Of two alternative pathways to the iminium species, C2 hydroxylation followed by dehydration versus direct desaturation, the latter possibility appears to be more likely, because the former mechanism would be expected to result in detectable incorporation of 18O from 18O2. The direct desaturation of a C-N bond implied by this analysis is analogous to that recently posited for the reaction of the l-Arg 4,5-desaturase, NapI, thus lending credence to the prior mechanistic proposal. Such a pathway could also potentially be operant in a subset of reactions catalyzed by Fe/2OG N-demethylases, which have instead been purported to enact C-N bond cleavage by methyl hydroxylation and elimination of formaldehyde.",
author = "Dunham, {Noah P.} and Mitchell, {Andrew J.} and {Del R{\'i}o Pantoja}, {Jos{\'e} M.} and Carsten Krebs and {Bollinger, Jr.}, {Joseph M.} and Boal, {Amie Kathleen}",
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α-Amine Desaturation of d -Arginine by the Iron(II)- and 2-(Oxo)glutarate-Dependent l -Arginine 3-Hydroxylase, VioC. / Dunham, Noah P.; Mitchell, Andrew J.; Del Río Pantoja, José M.; Krebs, Carsten; Bollinger, Jr., Joseph M.; Boal, Amie Kathleen.

In: Biochemistry, Vol. 57, No. 46, 20.11.2018, p. 6479-6488.

Research output: Contribution to journalArticle

TY - JOUR

T1 - α-Amine Desaturation of d -Arginine by the Iron(II)- and 2-(Oxo)glutarate-Dependent l -Arginine 3-Hydroxylase, VioC

AU - Dunham, Noah P.

AU - Mitchell, Andrew J.

AU - Del Río Pantoja, José M.

AU - Krebs, Carsten

AU - Bollinger, Jr., Joseph M.

AU - Boal, Amie Kathleen

PY - 2018/11/20

Y1 - 2018/11/20

N2 - When challenged with substrate analogues, iron(II)- and 2-(oxo)glutarate-dependent (Fe/2OG) oxygenases can promote transformations different from those they enact upon their native substrates. We show here that the Fe/2OG enzyme, VioC, which is natively an l-arginine 3-hydroxylase, catalyzes an efficient oxidative deamination of its substrate enantiomer, d-Arg. The reactant complex with d-Arg retains all interactions between enzyme and substrate functional groups, but the required structural adjustments and opposite configuration of C2 position this carbon more optimally than C3 to donate hydrogen (H•) to the ferryl intermediate. The simplest possible mechanism, C2 hydroxylation followed by elimination of ammonia, is inconsistent with the demonstrated solvent origin of the ketone oxygen in the product. Rather, the reaction proceeds via a hydrolytically labile C2-iminium intermediate, demonstrated by its reductive trapping in solution with NaB2H4 to produce racemic [2H]Arg. Of two alternative pathways to the iminium species, C2 hydroxylation followed by dehydration versus direct desaturation, the latter possibility appears to be more likely, because the former mechanism would be expected to result in detectable incorporation of 18O from 18O2. The direct desaturation of a C-N bond implied by this analysis is analogous to that recently posited for the reaction of the l-Arg 4,5-desaturase, NapI, thus lending credence to the prior mechanistic proposal. Such a pathway could also potentially be operant in a subset of reactions catalyzed by Fe/2OG N-demethylases, which have instead been purported to enact C-N bond cleavage by methyl hydroxylation and elimination of formaldehyde.

AB - When challenged with substrate analogues, iron(II)- and 2-(oxo)glutarate-dependent (Fe/2OG) oxygenases can promote transformations different from those they enact upon their native substrates. We show here that the Fe/2OG enzyme, VioC, which is natively an l-arginine 3-hydroxylase, catalyzes an efficient oxidative deamination of its substrate enantiomer, d-Arg. The reactant complex with d-Arg retains all interactions between enzyme and substrate functional groups, but the required structural adjustments and opposite configuration of C2 position this carbon more optimally than C3 to donate hydrogen (H•) to the ferryl intermediate. The simplest possible mechanism, C2 hydroxylation followed by elimination of ammonia, is inconsistent with the demonstrated solvent origin of the ketone oxygen in the product. Rather, the reaction proceeds via a hydrolytically labile C2-iminium intermediate, demonstrated by its reductive trapping in solution with NaB2H4 to produce racemic [2H]Arg. Of two alternative pathways to the iminium species, C2 hydroxylation followed by dehydration versus direct desaturation, the latter possibility appears to be more likely, because the former mechanism would be expected to result in detectable incorporation of 18O from 18O2. The direct desaturation of a C-N bond implied by this analysis is analogous to that recently posited for the reaction of the l-Arg 4,5-desaturase, NapI, thus lending credence to the prior mechanistic proposal. Such a pathway could also potentially be operant in a subset of reactions catalyzed by Fe/2OG N-demethylases, which have instead been purported to enact C-N bond cleavage by methyl hydroxylation and elimination of formaldehyde.

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