Magnetic and natural circular dichroism spectra of cytochromes P-450 11 β and P-450 scc purified from bovine adrenal cortex

Abstract

Magnetic (MCD) and natural circular dichroism (CD) spectra of various complexes of cytochrome P-450 11 β ( P-450 11 β ) and cytochrome P-450 scc ( P-450 scc) from bovine adrenal cortex were measured from 250 nm to 700 nm. MCD and CD spectral contours of cytochromes P-450 11 β and P-450 scc in the Soret and visible regions were, as a whole, analogous to those of cytochromes P-450 from rabbit liver microsomes and also from Pseudomonas putida in their high-spin ferric, high-spin ferrous and ferrous-CO complexes. MCD spectrum of the low-spin ferric P-450 scc free from the substrate, cholesterol, was very similar to that caused by addition of 20α-hydroxycholesterol, a reaction intermediate. However, it was distinct from those of the low-spin ferric P-450 11 β and P-450 scc complexes caused by addition of external nitrogenous ligands. The electronic states of the heme in the low-spin ferric P-450 free from substrates seemed to be subtly different from those of low-spin complexes coordinated with external nitrogenous ligands. Soret CD spectra of ferric low-spin complexes were not so different from each other. Upon reduction of high-spin ferric P-450 11 β or P-450 scc, the Soret CD magnitudes increased significantly in contrast with those of other P-450s, the Soret CD magnitudes of which decrease upon reduction. This may reflect an increased proximity of the neighbouring aromatic groups upon reduction of high-spin P-450 11 β or P-450 scc. High substrate specificity of adrenal P-450 s compared with liver P-450s can be explained in view of the above findings. The CD spectra in the near ultraviolet region (250–350 nm) were found to be quite sensitive to the spin change for ferric P-450 scc, while the MCD spectra in this region did not reflect substantially the spin state of the enzyme. MCD parameters of cytochrome P-450s were compared to those of other hemoproteins in diagrams describing selected MCD spectral values of hemoproteins so far available and were discussed in connection with the structures of the heme environment of P-450.