Binding of nitrosamines to cytochrome P-450 of liver microsomes.
The interactions of 5 carcinogenic and 1 non-carcinogenic nitrosamines with hepatic microsomal cytochrome (cyt.) P-450 were investigated, using both optical difference and electron paramagnetic resonance (EPR) spectroscopic methods. Liver microsomes from phenobarbital (PB)-pretreated mice and 3-methylcholanthrene (3-MC)-pretreated rats were used, in order to have an increased specific content of cyt. P-450 and cyt. P-448 respectively. The optical and EPR spectral data obtained in the oxidised state suggest that nitrosamines are able to bind both as substrates and as ligands to the hemoprotein cyt. P-450, depending on the concentration of nitrosamine, its chemical identity and the cytochrome species present. After reduction with dithionite or NADPH in the optical difference spectrum a Soret band developed between 444 and 453 nm to an extent, which is dependent on the particular nitrosamine present. This initial nitrosamine-induced spectrum might represent a ferrous nitric oxide (NO)-cyt. P-450 complex. It appears unstable and is converted kinetically into a spectrum lacking a Soret band, but with a predominant absorbance minimum at about 425 nm. A visible band is located at 585 nm. In the EPR spectrum a sharp 3-line signal around g = 2.01 appears concomitantly. Both spectral parameters are typical of a NO-cyt. P-420 complex. These results, in conjunction with metabolic studies, indicate that nitrosamines are denitrosated by a reductive process in which cyt. P-450 appears to be involved. The resulting NO-cyt. P-450 complex denatures to a NO-cyt. P-420 complex when the dioxygen level is not sufficiently high to complete successfully.
D000818:Animals; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D004578:Electron Spin Resonance Spectroscopy; D004790:Enzyme Induction / Q000187:drug effects; D007700:Kinetics; D008024:Ligands; D008297:Male; D008748:Methylcholanthrene / Q000494:pharmacology; D051379:Mice; D008862:Microsomes, Liver / Q000378:metabolism*; D009602:Nitrosamines / Q000378:metabolism*; D010084:Oxidation-Reduction; D010634:Phenobarbital / Q000494:pharmacology; D011485:Protein Binding; D051381:Rats; D013057:Spectrum Analysis
D008024:Ligands; D009602:Nitrosamines; D008748:Methylcholanthrene; D003577:Cytochrome P-450 Enzyme System; D010634:Phenobarbital