Purification and characterization of two unique forms of cytochrome P-450 from rabbit nasal microsomes

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Abstract
Two forms of cytochrome P-450, designated P-450NMa and P-450NMb, were purified to electrophoretic homogeneity from rabbit nasal microsomes. The purified cytochromes, which contained 14-16 nmol of P-450/mg of protein, exhibited apparent monomeric molecular weights of 49500 and 51000, respectively. As indicated by several criteria, including the amino acid composition, absorption spectra, and peptide maps, the two nasal forms of P-450 are distinct from each other. Furthermore, as judged by the NH2-terminal amino acid sequences, they are distinct from all other P-450 cytochromes described to date. In the ferric form, P-450NMa is in the low-spin state, whereas P-450NMb is predominantly in the high-spin state. When reconstituted with NADPH-cytochrome P-450 reductase and phospholipid, P-450NMa is very active in the oxidation of ethanol as well as several nasal procarcinogens, including the N-deethylation of N-nitrosodiethylamine, the O-deethylation of phenacetin, and the N-demethylation of hexamethylphosphoramide. P-450NMb also metabolizes these substrates, but at lower rates. Both nasal forms are also active with testosterone, with P-450NMa oxidizing the substrate in the 17 position to give androstenedione and P-450NMb catalyzing hydroxylation in the 15.alpha.-, 16.alpha.-, and 19-positions. The two cytochromes represent the major portion of the total P-450 in nasal microsomes, but the corresponding forms could not be detected in hepatic microsomes.