The reaction of phenylhydrazine with microsomal cytochrome P-450. Catalysis of heme modification.

作者：H. G. Jonen, Jurgen Werringloer, Russell A. Prough, R.W. Estabrook · 发表于：Journal of Biological Chemistry · 年份：1982 · DOI：10.1016/s0021-9258(18)34737-9 · 被引用次数：77 · 研究领域：Pharmacogenetics and Drug Metabolism、Drug Transport and Resistance Mechanisms、Porphyrin Metabolism and Disorders

Phenylhydrazine interacted with oxidized and reduced cytochrome P-450 of rat liver microsomes to produce binding difference spectra typical of many nitrogenous compounds. The phenylhydrazine-induced difference spectrum observed with oxidized microsomal cytochrome P-450 was converted, in a time-dependent process, to yield a new spectral intermediate with an absorbance maximum around 480 nm. The time required to form this new phenylhydrazine-induced spectral intermediate was decreased from hours to minutes when either NADPH or NADH was added to the reaction mixture. Phenyldiazene generated by addition of the decarboxylation product of methyl phenyldiazenecarboxylate or by addition of potassium ferricyanide and phenylhydrazine (2:1 molar equivalents) instantly formed the new spectral intermediate. This suggests that phenyldiazene is formed during the NADPH-dependent reaction. The appearance of the new spectral intermediate occurred concomitant with the loss of CO-reactive cytochrome P-450 (less than 90%) and loss of absorbance at 418 nm. The interpretation of the optical spectral changes was supported by a loss of the low spin signals characteristic of oxidized cytochrome P-450 as determined by electron paramagnetic resonance spectroscopy. The loss of CO-reactive cytochrome P-450 apparently resulted from the formation of a binary complex of phenyldiazene and the heme of oxidized cytochrome P-450 giving rise to the 480 nm spectral intermediate. In addition, the diazene-bound heme...