EPR study of the dinuclear active copper site of tyrosinase from Streptomyces antibioticus

The [Cu(I)-Cu(II)] half-met form of the dinuclear copper site of tyrosinase has been probed bg continuous wave electron paramagnetic resonance (EPR) a nd hyperfine sublevel correlation (HYSCORE) spectroscopy in the presence an d absence of inhibitors, In all cases the EPR spectrum is indicative of a d (x)(-y)(2)(2) ground state for the unpaired electron. From the cross-peaks observed in the HYSCORE spectra, proton hyperfine coupling constants were o btained that are compatible with a hydroxide ion in an equatorial coordinat ion position of the paramagnetic copper, After changing the water solvent t o D2O or after addition of the inhibitors p-nitrophenol or L-mimosine, the proton signals disappear, The relevance of these findings for understanding the catalytic cycle is discussed. (C) 2000 Federation of European Biochemi cal Societies.