Isolation and characterization of a cytochrome P450 of the IIA subfamily from human liver microsomes

Eur J Biochem. 1991 Sep 1;200(2):511-7. doi: 10.1111/j.1432-1033.1991.tb16212.x.


Antibodies raised against cytochrome P450, which is overexpressed in mouse hepatic tumors, (P450tu) crossreact with two human liver microsomal proteins (49 kDa and 52 kDa). We have quantified these proteins in 60 human liver samples and found great interindividual variability in both of them. The concentration of the 49-kDa protein varies up to 144 fold in the various samples and represents typically 10% of the total mincrosomal P450 content. Its immunologically determined concentration correlates well (R = 0.78) with the microsomal coumarin-7-hydroylase (COH) activity. This activity is strongly and completely inhibited by anti-P450tu antibody (IC50 = 0.13 mg IgG/mg microsomal protein). The crossreacting 49-kDa protein shows an unusually high substrate specificity towards coumarin; it presents all human COH and part of 7-ethoxycoumarin O-deethylase (ECOD). Besides these two activities, we did not find any activity with other typical P450 substrates. In primary cultures of human hepatocytes, it is inducible by phenobarbital and dexamethasone, but not by pyrazole and beta-naphthoflavone. We isolated this protein from human liver microsomes and purified it to homogeneity by a combination of aminooctyl-amino-Sepharose chromatography and immunoaffinity chromatography. The protein was identified as a cytochrome P450 of the IIA subfamily. Its N-terminal amino-acid sequence was identical with the first 20 residues deduced from the nucleotide sequence of P450IIA6.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • 7-Alkoxycoumarin O-Dealkylase / metabolism
  • Amino Acid Sequence
  • Blotting, Western
  • Cells, Cultured
  • Cross Reactions
  • Cytochrome P-450 Enzyme System / immunology
  • Cytochrome P-450 Enzyme System / isolation & purification*
  • Cytochrome P-450 Enzyme System / metabolism
  • Electrophoresis, Polyacrylamide Gel
  • Humans
  • Isoenzymes / immunology
  • Isoenzymes / isolation & purification*
  • Isoenzymes / metabolism
  • Microsomes, Liver / enzymology*
  • Molecular Sequence Data
  • Substrate Specificity


  • Isoenzymes
  • Cytochrome P-450 Enzyme System
  • 7-Alkoxycoumarin O-Dealkylase