Drug-induction decreases the mobility of cytochrome P-450 in rat liver microsomes: protein rotation study.
Effect of drug-induction on the rotation of cytochrome P-450 and on lipid fluidity in rat liver microsomes was examined. Rotational diffusion of cytochrome P-450 was examined by observing the decay of absorption anisotropy, r(t), after photolysis of the heme.CO complex by a vertically polarized laser flash. Analysis of r(t) was based on a "rotation-about-membrane normal" model. Microsomal lipid fluidity was measured by observing fluorescence anisotropy of DPH incorporated in the lipid bilayer. The absorption anisotropy decayed within 2 ms to a time-independent value. Rotational diffusion of cytochrome P-450 was dependent on the drug-induction with PB, MC, and PCB when compared with non-induced CON-microsomes. The observed values for the normalized time-independent anisotropy r(infinity)/r(0) are r(infinity)/r(0) = 0.41 (CON-microsomes), 0.54 (PB-microsomes), 0.52 (MC-microsomes), and 0.57 (PCB-microsomes). The average rotational relaxation time phi = 580-690 microseconds was almost unchanged over all microsomes presently examined. A significantly high value of r(infinity)/r(0) = 0.41-0.57 implies the co-existence of mobile and immobile populations of cytochrome P-450. Based on the assumption that the heme tilts about 55 degrees from the membrane plane for all species of P-450s besides P-450PB, 59% (CON-microsomes), 46% (PB-microsomes), 48% (MC-microsomes), and 43% (PCB-microsomes), respectively, of the cytochrome P-450 in microsomes is calculated to be mobile. Upon drug-induction the microsomal membrane was fluidized to some extent as judged by the steady-state fluorescence anisotropy of 0.156 for CON-microsomes and 0.139-0.148 for drug-induced microsomes.(ABSTRACT TRUNCATED AT 250 WORDS)
Kawato
S
S
Institute of Physics, College of Arts and Sciences, University of Tokyo.
Ashikawa
I
I
Iwase
T
T
Hara
E
E
D000818:Animals; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism*; D004790:Enzyme Induction; D005454:Fluorescence Polarization; D007425:Intracellular Membranes / Q000201:enzymology; D007700:Kinetics; D008297:Male; D008560:Membrane Fluidity; D008748:Methylcholanthrene / Q000494:pharmacology; D008862:Microsomes, Liver / Q000187:drug effects / Q000201:enzymology*; D010634:Phenobarbital / Q000494:pharmacology; D010782:Photolysis; D011078:Polychlorinated Biphenyls / Q000494:pharmacology; D051381:Rats; D011919:Rats, Inbred Strains; D012016:Reference Values
D008748:Methylcholanthrene; D003577:Cytochrome P-450 Enzyme System; D011078:Polychlorinated Biphenyls; D010634:Phenobarbital