Reactivity of ferric Aplysia and sperm whale myoglobins towards imidazole : X-ray and binding study

Abstract

Spectral and equilibrium binding properties for the reactions of sperm whale and Aplysia metmyoglobins with imidazole have been studied in solution and in single crystals, as a function of pH. Both in solution and in the crystal the high-spin forms of these two proteins show clear spectral differences, which have been correlated to the absence of the iron-bound water molecule in Aplysia metmyoglobin. On the other hand, both the hydroxyl and imidazole adducts give identical spectra for the two myoglobins. Under all the conditions explored, the binding constants for the reaction of ferric sperm whale and Aplysia myoglobins with imidazole are very similar when measured in solution and in single crystals, thus indicating that the crystalline lattice does not exert large strains on the structure of the active site. Three-dimensional difference Fourier studies of the imidazole adducts of the two proteins show that this ligand binds to the iron distal sites in a similar manner, despite the absence of the distal histidine in Aplysia myoglobin. In the case of sperm whale myoglobin, the distal histidine side-chain is displaced by the exogenous imidazole, and this fact is associated with structural changes in the sidechains mainly in the neighborhood of the heme. In both proteins the exogenous imidazole ring is approximately perpendicular to the heme plane and points towards the external part of the heme crevice along the Fe-(metinic)CHA direction.