Visible and magnetic circular dichroism studies on cobalt(II)-substituted rhus vernicifera laccase

The visible and magnetic circular dichroism (MCD) spectra of Co(II) derivatives of Rhus vernicifera laccase are reported. Anaerobic incorporation of 1 g-atom of Co(II) into apolaccase gave bands at 528(ϵ = 248), 558 (254) and 589 nm (shoulder) attributable to dd transitions. The MCD spectrum in the corresponding region is similar to that of Co(II)-substituted hemocyanin, indicating that the Co(II) ion incorporated into apolaccase is tetrahedral. On increasing the amount of Co(II) ion acting on the apolaccase, both the intensities of the absorption and the MCD spectra increased, and 2 g-atoms of tetrahedral Co(II) ion were introduced into the apolaccase. Very similar absorption and MCD spectra were obtained when laccase whose type I copper site was occupied by Hg(II) and both type II and type III copper sites were vacant (TlHg apolaccase) was treated with Co(II); this clearly supports the hypothesis that Co(II) cannot be incorporated into a type I copper site but may possibly be incorporated into a type III copper site. A tetrahedral Co(II) ion was also introduced into a type II copper site of type II copper-depleted (T2D) laccase, although its MCD bands were shifted ca. 20 nm to the longer wavelength region from the MCD bands due to tetrahedral Co(II) ion incorporated into type III copper site(s). The present study demonstrate that a tetrahedral Co(II) ion is introduced into type II or type III copper site(s) of laccase.