Cyanide compounds of ferroperoxidase and myoglobin and their reversible photodissociation

Abstract

Ferriperoxidase (per3) combines reversibly with cyanide over a wide range of pH (4.6 - 10.3) forming a stable ferriperoxidase-cyanide complex (per3CN) with an absorption spectrum very similar to that of metmyoglobin cyanide (MetmbCN) or catalase cyanide. Ferroperoxidase (per2) also combines reversibly with cyanide but within a much more limited range of pH (8.5 - 9.5), forming the less stable per2CN which can be obtained either by addition of HCN to per2 or of Na2S2O4 to per3 CN. The affinity of per2 for cyanide is about 1000 times lower than that of per3, the dissociation constants at pH 9 being 1.3 x 10-3 [image] for per2CN and 2 x 10-6 [image] for per3CN. The absorption spectrum of per2CN is identical with that of HbCN and MbCN. It is of the hemochromogen type, the positions and intensities of bands being: [alpha]-565.5 m[mu] ([image]mM= 135). On acidifying a solution of per2CN the above bands gradually disappear and are replaced, not by those of per2, but by the absorption bands of per3CN, the change being complete at pH 6. At pH 5.5-6.0 per3CN does not undergo reduction even in the presence of Na2S2O4; in this respect the compound resembles catalase. Per2 combines re-versibly with CO, forming a light-sensitive per2CO compound. The affinity of per2 for CO is much greater than for cyanide, the partition constant K = ((per2co) (HCN))/((per2CN) (CO)) being about 2500. Per2 combines also with methylcarbylamine, forming a compound very similar to the carbylamine compounds of Hb and Mb. Per2CN under-goes a reversible photodissociation which can easily be followed by direct microspectroscopic observation. The light sensitivity of this compound is of special interest since hitherto, only CO compounds of hemes were known to be light-sensitive. A method is described for quantitative spectrophotometric study of the kinetics of reversible dissociation of per2CN. The photodissociation was effected by means of Hg-blue light the wavelength of which (435.8 m[mu].) lies close to the in-tense [gamma]-band of per2CN. Examination of MbCN by the same method revealed a less marked but still distinct light sensitivity. An attempt to calculate, from the spectrophotometric experiments, the quantum efficiency in photodissociation of per2CN gave an approximate value of [phi]= 0.8 (moles per2CN dissociated per mole light quantum absorbed). In view of the uncertainties in the calculation this result indicates only that the efficiency is of the order of 1.