Copper(I)-phenolate complexes as models of the reduced active site of galactose oxidase: Synthesis, characterization, and O reactivity

The Cu(I)-phenolate complexes LCu and LCu and the Cu(I)-phenol complex [HLCu(CNCHMe)] BArF were prepared and structurally characterized by X-ray crystallography, where L and L are ligands comprised of a 2,4-di-tert-butylphenolate linked to 1-isopropyl-1,5-diazacyclooctane or 1,4-diisopropyl-1,4,7-triazacyclononane, respectively. The reduced galactose oxidase (GAO) structural models LCu and LCu were found to be highly reactive with O, and through combined stopped-flow kinetic and EPR, UV-vis, and resonance Raman spectroscopic studies of the oxygenation of LCu at low temperature, new intermediates relevant to those postulated for the active site oxidation step of the GAO catalytic cycle were identified. The oxygenation was shown by kinetics experiments to proceed via initial binding of O to yield a green, unusually thermodynamically stable 1:1 adduct, LCu(O). Symmetric (η) binding of a superoxo ligand was indicated by oxygen-isotope-sensitive features in resonance Raman spectra obtained in batch experiments; peaks at v(O) = 1120 cm, v(OO) = 1093 cm, and v(O) = 1058 cm were assigned as O-O stretching vibrations. These data represent the first experimental evidence for such superoxide coordination in complexes of tetradentate tripodal ligands and provide new precedent for how O may bind at the reduced GAO active site. The 1: 1 Cu/O adduct subsequently evolves into a metastable purple species that is only observable under conditions of substoichiometric O. The kinetics of formation of this transient species are second order overall (rate = k[LCu(O)] [L1Cu]). It exhibits an absorption band with λ = 565 nm (ε = 17,900 M cm) and multiple oxygen-isotope-sensitive v(Cu-O) and v(O-O) features in the respective regions 500-550 cm and 700-850 cm in Raman spectra, with excitation-wavelength-dependent intensities that correlate with the 565 nm absorption feature. On the basis of the combined data available, the presence of multiple isomeric peroxodicopper species in the transient purple solution is postulated.