Kinetic analysis of oxygen utilization during cofactor biogenesis in a copper-containing amine oxidase from yeast.

A detailed kinetic analysis of oxygen consumption during TPQ biogenesis has been carried out on a yeast copper amine oxidase. O(2) is consumed in a single, exponential phase, the rate of which responds linearly to dissolved oxygen concentration. This behavior is observed up to conditions of maximally obtainable oxygen concentrations. In contrast, no viscosity effect is observed on rate, implicating a high K(m) for O(2). Binding of oxygen appears to occur faster than its consumption and to result in displacement of the precursor tyrosine onto copper to form a charge-transfer species, described in the the preceding paper of this issue [Dove, J. E., Schwartz, B., Williams, N. K., and Klinman, J. P. (2000) Biochemistry 39, 3690-3698). Reaction between this intermediate and O(2) is proposed to occur in a rate-limiting step, and to proceed more rapidly when the tyrosine is deprotonated. This rate-limiting step in cofactor biogenesis does not display a solvent isotope effect and is, thus, uncoupled from proton transfer. Comparisons are drawn between the proposed biogenesis mechanism and that for the oxidation of reduced cofactor during catalytic turnover in the mature enzyme.