title: Experimental and computational evidence of metal-O2 activation and rate-limiting proton-coupled electron transfer in a copper amine oxidase.
abstract: The mechanism of O(2) 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 O(2) to H(2)O(2). Steady-state kinetics indicate that amine-reduced CuAGAO is reoxidized by O(2) >10(3) times faster than the CoAGAO analogue. Complementary spectroscopic studies reveal that the difference in the second order rate constant, k(cat)/K(M)(O(2)), arises from the more negative redox potential of Co(III/II) in relation to Cu(II/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 (η(1))-metal bound superoxide is reduced to an η(1)-hydroperoxide in the rate-limiting step.
authors.affiliation: Department of Chemistry, Johns Hopkins University, 3400 North Charles Street, Baltimore, Maryland 21218, United States.
authors.forename: Yi
authors.initials: Y
authors.lastname: Liu
authors.forename: Arnab
authors.initials: A
authors.lastname: Mukherjee
authors.forename: Nadav
authors.initials: N
authors.lastname: Nahumi
authors.forename: Mehmet
authors.initials: M
authors.lastname: Ozbil
authors.forename: Doreen
authors.initials: D
authors.lastname: Brown
authors.forename: Alfredo M
authors.initials: AM
authors.lastname: Angeles-Boza
authors.forename: David M
authors.initials: DM
authors.lastname: Dooley
authors.forename: Rajeev
authors.initials: R
authors.lastname: Prabhakar
authors.forename: Justine P
authors.initials: JP
authors.lastname: Roth
mesh_terms: D006631:Amine Oxidase (Copper-Containing) / Q000737:chemistry*; D004578:Electron Spin Resonance Spectroscopy; D007700:Kinetics; D008670:Metals / Q000737:chemistry*; D008958:Models, Molecular; D010100:Oxygen / Q000737:chemistry*; D013056:Spectrophotometry, Ultraviolet
chemical_list: D008670:Metals; D006631:Amine Oxidase (Copper-Containing); D010100:Oxygen