STRUCTURAL CHARACTERIZATION OF THE COPPER SITE IN GALACTOSE-OXIDASE USING X-RAY-ABSORPTION SPECTROSCOPY

X-ray absorption spectroscopy has been used to characterize the local structural environment of the Cu ion in the reductively inactivated, oxidatively activated, and active+substrate oxidation state derivatives of galactose oxidase. In all three cases, the local environment of the Cu is best modeled by a single shell of low-Z (N or O) scatterers. This is generally consistent with the structure determined crystallographically, although the EXAFS bond lengths are slightly, but significantly, shorter than those found crystallographically. The best-fit average bond lengths are 1.97, 1.95, and 1.98 Angstrom for inactive, active, and active+substrate, respectively. The Cu-II ion in the active and inactive derivatives has an apparent coordination number of 4, consistent with the equatorial ligation seen crystallographically. The Cu-I ion in the reduced+substrate derivative appears to have either a lower coordination number or a significantly more distorted local environment. The observed Cu-I-(N/O) bond length favors a model where the Cu becomes 3-coordinate in the substrate-reduced complex.