Cytochrome P-450 transfer from adrenocortical submitochondrial particles to liposome membranes.
The transfer of cytochrome P-450 from bovine adrenocortical submitochondrial particles (smp) to unilamellar liposome membranes was investigated using a table top ultracentrifuge. Submitochondrial particles were incubated with liposome membranes at 25 degrees C and precipitated by ultracentrifugation at 200000g for a few minutes at 25 degrees C. All liposome vesicles were recovered in the supernatant. Almost no proteins were detected in the supernatant when only smp were incubated and centrifuged. SDS-PAGE revealed one main protein band for the supernatant when smp were incubated with liposome vesicles at 25 degrees C. This band was reactive to anti-P-450scc IgG. Inaccuracy in time for kinetic studies of the transfer was less than 0.5 min. Transfer of P-450scc from smp to liposome membranes was further demonstrated by the decrease in side-chain cleavage activity of smp for endogenous cholesterol after incubation. Cytochromes P-450 accounted for about 70% of the transferred proteins in the liposome membranes, the amount of which increased exponentially with the incubation time. The inverse value of the relaxation time of the transfer increased linearly with the smp concentration and decreased hyperbolically with the liposome concentration. These results coincide with a mechanism by which cytochrome P-450 dissociates from smp membranes, diffuses, and binds to the liposome membranes. In the transfer of cytochrome P-450, the dissociation from smp membranes was deduced to be the rate-limiting step.
Kominami
S
S
Faculty of Integrated Arts and Sciences, Hiroshima University, Higashihiroshima, Japan.
Onizuka
M
M
Tasaka-Marumoto
C
C
Takemori
S
S
D000302:Adrenal Cortex / Q000378:metabolism*; D000818:Animals; D001692:Biological Transport; D015153:Blotting, Western; D002417:Cattle; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D007700:Kinetics; D008081:Liposomes / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism; D013367:Submitochondrial Particles / Q000378:metabolism*
D008081:Liposomes; D008565:Membrane Proteins; D003577:Cytochrome P-450 Enzyme System