The conformational changes of catalase molecule caused by ligand molecules

Abstract

Circular dichroism and optical rotatory dispersion of bovine liver catalase have been measured over wide wavelength range of 200–650 mμ. Native catalase showed a small negative CD band in the Soret absorption region, which corresponded to a small negative Cotton effect. The denaturation with acid (pH 3.0), alkali (pH 12.0) and 8 M urea caused almost complete loss of this Soret CD band and the Cotton effect. Addition of the ligand molecules such as KCN and NaN 3 to native catalase caused an appearance of new small CD band near 450 mμ. In the near-ultraviolet region native molecule exhibited a CD band at 285 mμ, while the denaturations caused blue shift of the band with a large enhancement in intensity at pH 12.0 as well as diminutions at pH 3.0 and by 8 M urea. The liganded catalase molecules showed a new band near 340 mμ in addition to the CD band at 280–290 mμ. These CD bands may be related to aromatic amino acids and also their interaction with the heme groups in the molecules. In the ultraviolet region all proteins except for the denatured one with 8 M urea exhibited two intense negative CD bands and a deep trough of Cotton effect at 233 mμ, which indicated the presence of α-helical structure in their molecules. It was concluded from these data that the helix content of the native molecule was about 50%, whereas the acid- and alkali-denatured molecules showed the less helical content of 20–25% and 15–20%, respectively. Treatment with 8 M urea seemed to cause complete loss of helical conformation in the molecule. However, addition of ligands (KCN and NaN 3) to the heme groups of catalase caused reduction of helical content in the molecule whose enzymic activity was completely lost. The estimated helical content was approx. 30%.