Experimental and computational evidence of metal-O2 activation and rate-limiting proton-coupled electron transfer in a copper amine oxidase

Yi Liu, Arnab Mukherjee, Nadav Nahumi, Mehmet Ozbil, Doreen Brown, Alfredo M. Angeles-Boza, David M. Dooley, Rajeev Prabhakar, Justine P. Roth

Research output: Contribution to journalArticle

17 Citations (Scopus)

Abstract

The mechanism of O2 reduction by copper amine oxidase from Arthrobacter globiformus (AGAO) is analyzed in relation to the cobalt-substituted protein. The enzyme utilizes a tyrosine-derived topaquinone cofactor to oxidize primary amines and reduce O2 to H 2O2. Steady-state kinetics indicate that amine-reduced CuAGAO is reoxidized by O2 >103 times faster than the CoAGAO analogue. Complementary spectroscopic studies reveal that the difference in the second order rate constant, kcat/KM(O2), arises from the more negative redox potential of CoIII/II in relation to CuII/I. Indistinguishable competitive oxygen-18 kinetic isotope effects are observed for the two enzymes and modeled computationally using a calibrated density functional theory method. The results are consistent with a mechanism where an end-on (η1)-metal bound superoxide is reduced to an η1-hydroperoxide in the rate-limiting step.

Original languageEnglish
Pages (from-to)218-229
Number of pages12
JournalJournal of Physical Chemistry B
Volume117
Issue number1
DOIs
StatePublished - Jan 10 2013

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Amine Oxidase (Copper-Containing)
oxidase
Amines
Protons
amines
electron transfer
Metals
Chemical activation
activation
Copper
copper
Kinetics
protons
Electrons
enzymes
Enzymes
Cobalt
oxygen 18
Superoxides
Isotopes

ASJC Scopus subject areas

  • Physical and Theoretical Chemistry
  • Materials Chemistry
  • Surfaces, Coatings and Films

Cite this

Experimental and computational evidence of metal-O2 activation and rate-limiting proton-coupled electron transfer in a copper amine oxidase. / Liu, Yi; Mukherjee, Arnab; Nahumi, Nadav; Ozbil, Mehmet; Brown, Doreen; Angeles-Boza, Alfredo M.; Dooley, David M.; Prabhakar, Rajeev; Roth, Justine P.

In: Journal of Physical Chemistry B, Vol. 117, No. 1, 10.01.2013, p. 218-229.

Research output: Contribution to journalArticle

Liu, Yi ; Mukherjee, Arnab ; Nahumi, Nadav ; Ozbil, Mehmet ; Brown, Doreen ; Angeles-Boza, Alfredo M. ; Dooley, David M. ; Prabhakar, Rajeev ; Roth, Justine P. / Experimental and computational evidence of metal-O2 activation and rate-limiting proton-coupled electron transfer in a copper amine oxidase. In: Journal of Physical Chemistry B. 2013 ; Vol. 117, No. 1. pp. 218-229.
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AU - Nahumi, Nadav

AU - Ozbil, Mehmet

AU - Brown, Doreen

AU - Angeles-Boza, Alfredo M.

AU - Dooley, David M.

AU - Prabhakar, Rajeev

AU - Roth, Justine P.

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N2 - The mechanism of O2 reduction by copper amine oxidase from Arthrobacter globiformus (AGAO) is analyzed in relation to the cobalt-substituted protein. The enzyme utilizes a tyrosine-derived topaquinone cofactor to oxidize primary amines and reduce O2 to H 2O2. Steady-state kinetics indicate that amine-reduced CuAGAO is reoxidized by O2 >103 times faster than the CoAGAO analogue. Complementary spectroscopic studies reveal that the difference in the second order rate constant, kcat/KM(O2), arises from the more negative redox potential of CoIII/II in relation to CuII/I. Indistinguishable competitive oxygen-18 kinetic isotope effects are observed for the two enzymes and modeled computationally using a calibrated density functional theory method. The results are consistent with a mechanism where an end-on (η1)-metal bound superoxide is reduced to an η1-hydroperoxide in the rate-limiting step.

AB - The mechanism of O2 reduction by copper amine oxidase from Arthrobacter globiformus (AGAO) is analyzed in relation to the cobalt-substituted protein. The enzyme utilizes a tyrosine-derived topaquinone cofactor to oxidize primary amines and reduce O2 to H 2O2. Steady-state kinetics indicate that amine-reduced CuAGAO is reoxidized by O2 >103 times faster than the CoAGAO analogue. Complementary spectroscopic studies reveal that the difference in the second order rate constant, kcat/KM(O2), arises from the more negative redox potential of CoIII/II in relation to CuII/I. Indistinguishable competitive oxygen-18 kinetic isotope effects are observed for the two enzymes and modeled computationally using a calibrated density functional theory method. The results are consistent with a mechanism where an end-on (η1)-metal bound superoxide is reduced to an η1-hydroperoxide in the rate-limiting step.

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