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The Mechanism of 2‐Nitropropane Dioxygenase: a Model for Flavin Semiquinone Intermediates in Enzymatic Catalysis
Author(s) -
Gadda Giovanni,
Francis Kevin,
Belaineh Merid
Publication year - 2007
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.21.5.a272
Subject(s) - chemistry , flavin group , photochemistry , substrate (aquarium) , catalysis , dioxygenase , semiquinone , kinetic isotope effect , catalytic cycle , stereochemistry , enzyme , organic chemistry , quinone , oceanography , physics , deuterium , quantum mechanics , geology
2‐Nitropropane dioxygenase (E.C.1.13.11.32) is an FMN‐dependent enzyme that catalyzes the oxidation of nitroalkanes and nitronates to their corresponding carbonyl compounds and nitrite. Mechanistic studies using pH, solvent viscosity, substrate kinetic isotope effects and rapid‐mixing techniques have been used to investigate the chemical mechanism of substrate oxidation in the reaction catalyzed by 2‐nitropropane dioxygenase. The resulting data are consistent with a catalytic base, His‐196 in the enzyme from Neurospora crassa , initiating the oxidative pathway by abstracting a proton from the α‐carbon of the neutral substrate to generate an enzyme‐bound alkylnitronate intermediate. This kinetic step is followed by a single electron transfer from the alkylnitronate to the enzyme‐bound flavin that results in the transient formation of an anionic flavosemiquinone and a substrate radical. An electrostatic catalyst in the active site of the enzyme facilitates the formation of the anionic flavosemiquinone. After the flavin‐linked one‐electron reduction of molecular oxygen, the resulting superoxide reacts with the substrate radical generating a peroxynitroalkane intermediate before decaying to give the carbonyl product of the reaction. Catalysis with alkylnitronates as substrates proceeds in a similar fashion except that the initial proton abstraction step is excluded.