Probing the cytochrome P-450 2B1 active site with diamantoid compounds

• Published in: General physiology and biophysics Vol. 14; no. 3; pp. 225 - 239
• Main Authors: Hodek, P, Burkhard, J, Janků, J
• Format: Journal Article
• Language: English
• Published: Slovakia 01.06.1995
• Subjects: Adamantane - analogs & derivatives; Adamantane - chemistry; Adamantane - metabolism; Aminopyrine - metabolism; Animals; Aryl Hydrocarbon Hydroxylases; Binding Sites; Binding, Competitive; Biophysical Phenomena; Biophysics; Cytochrome P-450 Enzyme System - chemistry; Cytochrome P-450 Enzyme System - metabolism; In Vitro Techniques; Kinetics; Microsomes, Liver - enzymology; Molecular Structure; Rats; Rats, Wistar; Steroid Hydroxylases - chemistry; Steroid Hydroxylases - metabolism
• ISSN: 0231-5882

Hydrocarbone diamantane has been shown to be a specific substrate with a high affinity for the binding site of PB-inducible cytochrome P-450 2B1 (Hodek et al. 1988). Using a difference spectroscopy approach, a battery of diamantane analogues and diamantane oxygen containing derivatives were examined for their interaction with P-450 2B1 active site. Of the compounds (diamantane and its analogues, adamantane and triamantane) tested, diamantane had the lowest value of a spectral dissociation constant Ks = 0.5 mumol/l, indicating that diamantane was accommodated well to the cytochrome P-450 2B1, hence values of 0.46 nm and 0.66 nm for the width and length of the diamantane molecule, respectively, were used to describe of the dimensions the cytochrome P-450 binding site. Adamantane (Ks = 1.3 mumol/l) is relatively small and thus it binds loosely whereas triamantane (Ks = 4.3 mumol/l) is bulky enough to fit the binding site. This conclusion has been confirmed by spectral competition experiments as well as metabolic studies. Of all oxygen containing derivatives diamantane 1,6-dicarboxylic acid dimethylester only exhibited a pronounced ligand interaction with cytochrome P-450. Using molecular dimensions of this derivative the distance of 0.56 nm from the heme iron to the center of the substrate binding site was estimated.