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Biochimie. 1996;78(8-9):706-13.

Cytochrome P450 conformation and substrate interactions as probed by CO binding kinetics.

Author information

1
Laboratory of Molecular Carcinogenesis, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.

Erratum in

  • Biochimie 1997;79(1):20.

Abstract

The kinetics of CO binding to cytochrome P450, as measured by the flash photolysis technique, is a powerful probe of P450 structure-function relationships. The kinetics are sensitive to P450 conformation and dynamics and are modulated by P450 interactions with substrates and other components of the microsomal membrane. Application of a difference method to kinetic data analysis distinguishes the kinetic behavior of individual P450 forms in the microsomal membrane. This approach shows that substrates differentially modulate the kinetics via: 1) changes in P450 conformation/dynamics that either accelerate or reduce the binding rate; and/or 2) steric effects that reduce the rate. Both mechanisms are observed, the relative contributions of each varying in a substrate- and P450-dependent manner. In addition to microsomes, substrate interactions with individual P450s can be similarly probed using expressed P450s. Experiments with baculovirus-expressed human P450 3A4 show that this P450 consists of multiple conformers with distinct substrate specificities, an observation which provides a basis for its recognition of a wide array of structurally diverse substrates. These studies thus demonstrate the utility of CO binding kinetics in elucidating fundamental P450-substrate interactions in a biological membrane environment.

PMID:
9010599
DOI:
10.1016/s0300-9084(97)82528-x
[Indexed for MEDLINE]

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