Optical Rotatory Dispersion and Circular Dichroism of Horse Myoglobin and Its Derivatives

Abstract

Optical rotatory dispersion and circular dichroism of solutions of horse ferri-myo-globin and its derivatives (ferro-, carbonyl-, oxy-, cyano-, azide-, apo- and reconstituted myoglobin) were measured over a wavelength range of 650 to 185mμ. All myoglobins except apo-myoglobin showed distinct positive Cotton effects and circular dichroic bands in the region of Soret absorptions. However, the crossover points of the Cotton effects were essentially identical with those of the positive maxima of the circular dichroic bands and the absorption maxima of the Soret bands. The magnitude of the Soret Cotton effect and circular dichroic band of carbonylmyoglobin was most prominent. In the near-ultraviolet region between 260-280mμ all proteins showed positive circular dichroic bands, which may be due to asymmetry of the aromatic amino acid groups. The intensities of these bands were enhanced giving a blue shift when the myoglobin was complexed with the ligand molecules. In the ultraviolet region intense negative circular dichroic bands near 222mμ and 207mμ were present in accord with the cor-responding Cotton effects having a trough at 233mμ and a peak near 200mμ. Helix contents, based on the 233mμ trough and the band intensity at 222mμ, were estimated for all the proteins. Within experimental error, horse ferrimyoglobin and its deriva-tives have almost the same helix content of about 70%, whereas apomyoglobin has less helix content, approximately 50%. This indicates that the addition of ligands to the heme group causes no drastic conformational change in the protein moiety, while the removal of heme has an effect on the diminution of the helix content. Reconstituted ferrimyoglobin regained essentially the same conformation as the native protein.