An investigation on reduction process of cucumber ascorbate oxidase.

Reduction process of cucumber ascorbate oxidase with L-ascorbate was investigated in detail through absorption and electron paramagnetic resonance (EPR) spectra under anaerobic condition. One of the three type I coppers (the type I copper which is oxidized rapidly (Sakurai, T. et al. (1985) Biochem. Biophys. Res. Commun. 131, 647-652)) and a pair of type III coppers only which contribute to the absorption at 330 nm were reduced faster than other two type I and the other pair of type III coppers, respectively. The principal active site of ascorbate oxidase was confirmed to be comprised of one type I, one type II and a pair of type III coppers. Type II copper seemed to be reduced after all type I and type III coppers have been reduced.