Open AccessIsolation and sequence determination of a cDNA clone related to human cytochrome P-450 nifedipine oxidase.
Author(s) -
Philippe Beaune,
Diane R. Umbenhauer,
Richard W. Bork,
R. Stephen Lloyd,
F. Peter Guengerich
Publication year - 1986
Publication title -
proceedings of the national academy of sciences of the united states of america
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.011
H-Index - 771
eISSN - 1091-6490
pISSN - 0027-8424
DOI - 10.1073/pnas.83.21.8064
Subject(s) - microbiology and biotechnology , complementary dna , biology , cytochrome , pregnenolone , cdna library , peptide sequence , nucleic acid sequence , genomic library , hemeprotein , gene , biochemistry , heme , enzyme , steroid , hormone
Human liver cytochrome P-450NF is the form of cytochrome P-450 responsible for the oxidation of the calcium-channel blocker nifedipine, which has been reported to show polymorphism in clinical studies. By screening a bacteriophage lambda gt11 expression cDNA library, we isolated two clones: NF95 with an insert length of 0.8 kilobases which gave a stable fusion protein and NF25 with an insert length of 2.2 kilobases. The two clones were both sequenced and shown to be identical in their overlapping section. The sequence of NF25 is 77% similar to that reported for a rat cytochrome "P-450PCN" cDNA (PCN = pregnenolone-16 alpha-carbonitrile). The similarity decreases to 45-53% when the sequence is compared to human cytochromes P-450 belonging to other families [i.e., "pH P-450(1)," "P1-450," "P3-450," and "P-450MP." The deduced amino acid sequence is 73% similar to that of rat cytochrome P-450PCN, and the first 21 amino acids are identical to those reported for human liver cytochrome "P-450p." Sections of these clones were nick-translated and used as probes for analyses of human mRNA and genomic DNA. The number and size of bands indicate that P-450NF belongs to a multigene family, the so-called pregnenolone-16 alpha-carbonitrile-inducible family.