Different influences of two fractions of lung cytochrome b5 on reconstituted lung benzphetamine N-demethylase system.
Chromatography of lung microsomal cytochrome b5 obtained from DEAE-cellulose columns, yielded two distinct cytochrome b5 fractions. These cytochrome b5 fractions were further purified by Sephadex G-100 gel filtration chromatography. The specific cytochrome b5 content of fraction 1 and fraction 2 was found to be 16.5 and 16.4 nmol/mg protein respectively. Both fractions were free of cytochrome P-450, NADPH-cytochrome P-450 reductase and NADH-cytochrome b5 reductase activities. The effects of lung cytochrome b5 (fraction 1 and fraction 2) and liver cytochrome b5 on benzphetamine N-demethylase activity were examined. Four different reconstitution systems were used. Lung cytochrome b5 fraction 2 and liver cytochrome b5 stimulated N-demethylase activity in all four systems when b5:P-450 molar ratio was low, i.e. 0.25 or 0.5. Both cytochrome b5 samples inhibited N-demethylase activity when b5:P-450 ratio exceeded 1:1 molar ratio. In contrast lung cytochrome b5 fraction 1 stimulated N-demethylase activity in all four systems. Maximal enhancement of the activity was obtained when b5:P-450 ratio was 0.5. The greatest increase in N-demethylation activity was observed in the reconstitution system with the lowest concentration of cytochrome P-450 reductase, conditions which most closely resemble intact microsomes.
D000818:Animals; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism; D015786:Cytochromes b5 / Q000302:isolation & purification / Q000378:metabolism*; D004579:Electron Transport; D008099:Liver / Q000378:metabolism; D008168:Lung / Q000201:enzymology*; D008861:Microsomes / Q000378:metabolism; D009243:NAD / Q000378:metabolism / Q000494:pharmacology; D009249:NADP / Q000378:metabolism; D009251:NADPH-Ferrihemoprotein Reductase; D010088:Oxidoreductases / Q000378:metabolism; D010089:Oxidoreductases, N-Demethylating / Q000302:isolation & purification / Q000378:metabolism*; D012756:Sheep
D009243:NAD; D009249:NADP; D015786:Cytochromes b5; D003577:Cytochrome P-450 Enzyme System; D010088:Oxidoreductases; C019781:benzphetamine N-demethylase; D010089:Oxidoreductases, N-Demethylating; D009251:NADPH-Ferrihemoprotein Reductase