[Purification of cytochrome P-450 and NADPH cytochrome p-450 reductase from human liver].
Two methods for the purification of cytochromes-P450 from microsomes of human liver are described. Method A: Cyt-P450 were solubilized from microsomes using a non ionic detergent, the Lubrol. The Cyt-P450 were purified by affinity, hydrophobicity followed by ion-exchange chromatography on DEAE-5PW column (HPLC) with an overall yield of 18% and a specific activity of 10 nmole/mg of protein. The recovery of NADPH Cyt-P450 reductase by method A (affinity) is about 60% with a specific activity of 16.2 U.I./mg of protein. Method B: Cyt-P450 were solubilized from microsomes using a zwitterionic detergent, the CHAPS. Cyt-P450 were filtered and separated by chromatofocusing on Mono-P column (HPLC). By this method it was possible to increase strongly the specific activity keeping a yield of 50% of Cyt-P450. Also it was possible to apply this method to small samples of human liver like biopsies (0.5 to 2.5 g).
Isa
M
M
Laboratoire de Biochimie Générale, U.L.B. Bruxelles, Belgique.
Cumps
J
J
Fossoul
C
C
Atassi
G
G
D003577:Cytochrome P-450 Enzyme System / Q000302:isolation & purification*; D006801:Humans; D008862:Microsomes, Liver / Q000201:enzymology*; D009251:NADPH-Ferrihemoprotein Reductase / Q000302:isolation & purification*
D003577:Cytochrome P-450 Enzyme System; D009251:NADPH-Ferrihemoprotein Reductase
Purification des cytochromes-P450 et de la NADPH cytochrome-P450 réductase de foie humain.