Interaction of Polycyclic Hydrocarbons with Cytochrome P-450. III. Effects of Hydrocarbon Binding on the Interaction of Some Ligands with P-4481 Heme1

Abstract

The binding of polycyclic hydrocarbons to P-448 ₁ affected the interaction of exogenous ligands, such as ethyl isocyanide and CO in the ferrous state and 1-methyl imidazole in the ferric state, with the heme to various extents depending on the structure of the hydrocarbons. The effect of the hydrocarbons on dissociation constants ( K_d ) on the three ligands was essentially similar. K_d of ethyl isocyanide and CO for hydrocarbon-free P-448 ₁ were 18μ m and 1.1 μ m , respectively. The affinities of both the ligands increased about 3-fold when hydrocarbons of a small molecular size, such as phenanthrene, were bound to P-448 ₁ . Addition of a benzene ring to the outside of the phenanthrene molecule resulted in a remarkable decrease in the affinities of both the ligands. The binding of larger hydrocarbons in size produced a stronger inhibitory effect on the interaction of ethyl isocyanide with P-448 ₁ . Thus, K_d of the isocyanide for benz[a]anthracene-bound and dibenzf[a,c]-anthracene-bound P-448 ₁ were estimated to be 260 μ m and 3 m m , respectively. Spectral changes hardly occurred on addition of 4 m m ethyl isocyanide to 3-methyl-cholanthrene-bound or 7,8-benzoflavone-bound P-448 ₁ . The effect of the hydrocarbon binding on K_d of the ligands was depressed when a benzene ring which links two aromatic rings in the hydrocarbon molecule was replaced by a single C-C bond. A probable structure of the active area of P-448 ₁ composed of the substrate site and the heme is proposed based on the effect of the hydrocarbon binding on the interaction between the exogenous ligands and the P-448 ₁ heme iron reported in this series of papers. The binding of hydrocarbons to P-448 ₁ increased the intensity ratio of the 430 nm to the 453 nm peak of the absorption spectrum of the ethyl isocyanide compound. When a large and long hydrocarbon in shape such as dibenz[a,h]anthracene was bound to P-448 ₁ , a red shift of the Soret absorption peak and modification of the Soret CD pattern of the CO compound were observed.