Stimulatory effects of lung cytochrome b5 on benzphetamine N-demethylation in a reconstituted system containing lung cytochrome P450 LgM2

Abstract

• 1.

  1. Cytochrome b5 was partially purified from sheep lung microsomes in the presence of detergents Emuigen 913 and cholate by three consecutive DEAE-cellulose and Sephadex G-100 gel filtration chromatographies.

• 2.

  2. The specific content ofcytochrome b5 was 16.5 nmol/mg protein and purified cytochrome b5 fractions were free of cytochrome P450, NADPH-cytochrome P450 reductase and NADH-cytochrome b5 reductase activities.

• 3.

  3. The influences of increasing concentrations of lung cytochrome b5 on benzphetamine N-demethylation reactions were examined in four different reconstitution systems containing lung cytochrome P 450 LgM2, lung cytochrome P450 reductase and lipid. In each system concentration of reductase was doubled with respect to former system.

• 4.

  4. In all systems cytochrome b 5 stimulated benzphetamine Ndemethylase activity especially when cytochrome b5 was present at 0.5:1 molar ratio with respect to cytochrome /P450 LgM2.

• 5.

  5. Besides, the greatest fold of increase in benzphetamine N-demethylation activity due to addition of cytochrome b5 was observed in System 1 with the lowest concentration of reductase.