Studies on cobalt myoglobins and hemoglobins. Electron paramagnetic resonance investigation of iron-cobalt hybrid hemoglobins and its implication for the heme-heme interaction and for the alkaline Bohr effect.

An artificial hybrid hemoglobin cu(Co)P(Fe), the (Y and p subunits of which contain cobaltous protoporphyrin IX and ferrous protoporphyrin IX, respectively, and its complementary hybrid cu(Fe)P(Co), were prepared and examined by electron paramagnetic resonance spectroscopy. The fully oxy EPR spectra of cu(Co)p(Fe) and cu(Fe)P(Co) differ in their hyperfine splitting from each other, showing different environment of the paramagnetic center (principally the oxygen bound to the cobaltous ion). Fully deoxy a(Fe)P(Co) shows an EPR spectrum similar to those of deoxy CoHb pdsH chain or deoxy CoMb. The EPR spectrum of fully deoxy cu(Co)p(Fe), being different from those deoxy CoHb (yes” chain or deoxy CoMb, exhibits a broad gL component at g = 2.39 and g,, component at g = 2.03 with poorly resolved superhyperfine splitting, which is probably caused by the in-plane anisotropy of the cobaltous ions in the (Y subunits. Comparison of these EPR spectra with that of deoxy CoHb suggests that the electronic state similar to the cobaltous ions in the (Y subunits of fully deoxy cu(Co)P(Fe) also exists in the (Y subunits of deoxy CoHb. Ligation of carbon monoxide to the ferrous (Y subunits somewhat sharpens the g, signal of cu(Fe)fi(Co). Upon ligation of carbon monoxide to the ferrous fi subunits, the anomalous EPR spectrum of fully deoxy a(Co)/3(Fe) changes to that very similar to the deoxy CoHb @” chain, showing that the ligation in the /3 subunits drastically changes the electronic state of the (Y subunits. The apparent amplitude of the EPR signal of fully deoxy cu(Co)P(Fe) at g = 2.39 decreases, whereas that at g = 2.33 increases upon raising pH. The deoxy EPR spectrum of CoHb also shows a pH dependence similar to that of