Received for publication January 27, 2005.
Revised March 15, 2005.
Accepted for publication March 16, 2005.

ENZYME-SELECTIVE EFFECTS OF NITRIC OXIDE ON AFFINITY AND MAXIMUM VELOCITY OF VARIOUS RAT CYTOCHROMES P450

Abstract

Nitric oxide (NO) has recently been shown to decrease cytochrome P450 (P450) enzyme activity rapidly (<= 30 min), concentration-dependently, and enzyme-selectively in the rat liver. Interestingly, among all the studied P450 enzymes, only CYP2D1 was not affected by NO donors. However, these studies were conducted using only a single concentration of the substrates, thus lacking information about the possible simultaneous changes in both maximum velocity (V_max) and affinity (K_m) of the enzymes. In the present study, we systematically evaluated the effects of NO on the enzyme kinetic parameters of marker substrates for a range of P450 enzymes, including 2D1. Livers were perfused (1 h) in the absence (Control) or presence of two NO donors with different mechanisms of NO release. At the end of the perfusion, microsomes were prepared and used for kinetic analysis. Except for 2D1, NO reduced the V_max of all the model reactions studied, although to a varying degree. However, the effects of NO donors on K_m were more diverse. Whereas the K_m values for testosterone 6- (3A2) and 16- (2C11) hydroxylation significantly decreased, the values for chlorzoxazone 6-hydroxylation (2E1), dextromethorphan N-demethylation (3A2), and high affinity ethoxyresorufin O-dealkylation (1A1/2) significantly increased in the presence of NO donors. Further, the K_m values for the high affinity component of dextromethorphan O-demethylation and benzyloxyresorufin O-dealkylation remained unchanged. These results indicate that NO can potentially change both the V_max and K_m of various substrates selectively and confirm our previous findings that the activity of CYP2D1 is not affected by NO donors.

Key words: CYP inhibition, cytochrome P450 isoforms, enzyme inhibitors, enzyme kinetics, isolated perfused liver, kinetics, liver microsomes

This article has been cited by other articles:

				R. Vuppugalla and R. Mehvar SELECTIVE EFFECTS OF NITRIC OXIDE ON THE DISPOSITION OF CHLORZOXAZONE AND DEXTROMETHORPHAN IN ISOLATED PERFUSED RAT LIVERS Drug Metab. Dispos., July 1, 2006; 34(7): 1160 - 1166. [Abstract] [Full Text] [PDF]

				H. C. Yohe, K. A. O'Hara, J. A. Hunt, T. J. Kitzmiller, S. G. Wood, J. L. Bement, W. J. Bement, J. G. Szakacs, S. A. Wrighton, J. M. Jacobs, et al. Involvement of Toll-like receptor 4 in acetaminophen hepatotoxicity Am J Physiol Gastrointest Liver Physiol, June 1, 2006; 290(6): G1269 - G1279. [Abstract] [Full Text] [PDF]