Chiral orientation of prosthetic heme in the cytochrome P-450 active site.

Abstract

The absolute orientation of the prosthetic heme group in the active site of a hemoprotein may influence its substrate selectivity and catalytic properties. The only method available until now to determine the chiral orientation of the heme in a hemoprotein has been high resolution x-ray crystallography. The orientation of the heme in cytochrome P-450, therefore, is unknown because a crystallographic structure is not available for any form of this enzyme. We report here that the absolute configurations of the N-ethylprotoporphyrin IX adducts formed from the prosthetic hemes of cytochrome P-450 and hemoglobin during catalytic turnover of appropriate substrates are identical. The prosthetic heme in the inactivated cytochrome P-450 enzyme, therefore, has exactly the same orientation, relative to the fifth iron ligand, as the heme in hemoglobin. The approach described here can be used to determine the prosthetic heme orientation in other hemoproteins, including other cytochrome P-450 isozymes, for which x-ray structures are not available.