Isolation, partial purification, and characterization of the cytochrome P-450-dependent monooxygenase system from the midgut of the earthworm Lumbricus terrestris.
1. Cytochrome P-450 was purified from microsomes of the midgut of the earthworm Lumbricus terrestris up to a maximal specific content of 5.5 nmol P-450/mg protein. 2. At least 3 different cytochromes P-450 with apparent molecular weights of 48,000, 51,000 and 53,000 were identified by SDS-PAGE. 3. Western blot analysis with various polyclonal antibodies did not show structural epitopes common to the cytochromes P-450 of rodents or yeast and L. terrestris. 4. The microsomes contained about 43 pmol P-450/mg protein corresponding to 0.51 nmol P-450/g midgut and 64 pmol P-450/g body weight, respectively, and converted benzyloxyresorufin into resorufin with a Vmax of 2.12 pmol resorufin/min.mg protein and a Km of 770 nM benzyloxyresorufin at 25 degrees C, pH 8.0. 5. The microsomes exhibited a NADPH-cytochrome P-450 reductase activity of 9.4 nmol cytochrome c/min.mg protein. 6. The apparent molecular weight of the threefold-purified reductase was 63,000.
Institut für Pharmakologie und Toxikologie, Philipps-Universität Marburg, Federal Republic of Germany.
A G
AG
Berghout
E
E
Wenzel
J
J
Büld
K J
KJ
Netter
D000818:Animals; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry / Q000302:isolation & purification*; D004064:Digestive System / Q000201:enzymology; D008861:Microsomes / Q000201:enzymology; D009251:NADPH-Ferrihemoprotein Reductase / Q000302:isolation & purification; D009835:Oligochaeta / Q000201:enzymology*; D010105:Oxygenases / Q000737:chemistry / Q000302:isolation & purification*
D003577:Cytochrome P-450 Enzyme System; D010105:Oxygenases; D009251:NADPH-Ferrihemoprotein Reductase