Association of hydrophobic substances with hemin. Characterization of the reverse type I binding spectrum and its relationship to cytochrome P-450.

When hydrophobic compounds were added to a solution of protoferriheme, a a reverse type I spectral change was produced when observed by difference spectroscopy. The spectrum had a peak at 422 nm and a trough at 387 nm, and the characteristics were dependent on the pH of the sample. An association constant for the complex could be determined and was also found to be pH sensitive, with the association constant dropping to zero at values below pH 7.0 and above pH 8.5. The determination of the delta Absmax for the ethylbenzene-hemin complex at various hemin concentrations indicates monomeric heme to be the species responsible for binding the hydrocarbon with the concomitant generation of the reverse type I spectral change.