Component Interactions and Electron Transfer in Toluene/o-Xylene Monooxygenase
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Lippard_Component interactions.pdf
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Author(s)
Liang, Alexandria D
Lippard, Stephen J.
Date Issued
November 2014
Journal
Biochemistry
Publisher
American Chemical Society (ACS)
Citation
Liang, Alexandria Deliz and Lippard, Stephen J. "Component Interactions and Electron Transfer in Toluene/o-Xylene Monooxygenase." Biochemistry 53, no. 47 (2014 November): 7368–7375 © 2014 American Chemical Society
Version
Final published version
Abstract
The multicomponent protein toluene/o-xylene monooxygenase (ToMO) activates molecular oxygen to oxidize aromatic hydrocarbons. Prior to dioxygen activation, two electrons are injected into each of two diiron(III) units of the hydroxylase, a process that involves three redox active proteins: the ToMO hydroxylase (ToMOH), Rieske protein (ToMOC), and an NADH oxidoreductase (ToMOF). In addition to these three proteins, a small regulatory protein is essential for catalysis (ToMOD). Through steady state and pre-steady state kinetics studies, we show that ToMOD attenuates electron transfer from ToMOC to ToMOH in a concentration-dependent manner. At substoichiometric concentrations, ToMOD increases the rate of turnover, which we interpret to be a consequence of opening a pathway for oxygen transport to the catalytic diiron center in ToMOH. Excess ToMOD inhibits steady state catalysis in a manner that depends on ToMOC concentration. Through rapid kinetic assays, we demonstrate that ToMOD attenuates formation of the ToMOC–ToMOH complex. These data, coupled with protein docking studies, support a competitive model in which ToMOD and ToMOC compete for the same binding site on the hydroxylase. These results are discussed in the context of other studies of additional proteins in the superfamily of bacterial multicomponent monooxygenases.
MIT Department
Massachusetts Institute of Technology. Department of Chemistry
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DOI of Published Version
https://doi.org/10.1021/bi500892n
