Genomic cloning and protein expression of a novel rat brain cytochrome P-450 CYP2D18* catalyzing imipramine N-demethylation

Hidenori Kawashima, David J. Sequeira, David Nelson, Henry W. Strobel

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Abstract

We have previously reported the isolation of two cDNA clones, designated 2d-29 and 2d-35, which have identical open reading frames and code for a novel brain cytochrome p-450 (P-450) belonging to the CYP2D subfamily, and noted that the mRNA of clone 2d-35 seems to be expressed in the brain but not in the liver (1). Although the deduced amino acid sequence of these clones differs from that of the liver CYP2D4 by only 5 amino acids distributed in the C-terminal region, this new P-450 cDNA clone contained a unique 5'- extension, and we posit in this report by analysis of a genomic clone that this 5'-untranslated sequence is derived from a gene distinct from that of CYP2D4. Thus, this novel P-450 was named P-450 2D18 according to the recommended nomenclature (2). The expressibility of this cDNA was confirmed by in vitro translation using a reticulocyte system, and protein expression was performed using COS-M6 cells. Immunoblot analysis showed a cross-reacting band of the predicted size range with anti-P-450 2D6 antiserum, which was not seen in control cells. Furthermore, the CYP2D18-ex-pressed COS cell lysate showed N-demethylation activity toward imipramine, whereas another brain P- 450 CYP4F6-expressed COS cell lysate showed 10-hydroxylation activity. This is the first report that associates an individual P-450 isozyme in brain with a particular metabolic alteration of the antidepressant imipramine.

Original languageEnglish (US)
Pages (from-to)28176-28180
Number of pages5
JournalJournal of Biological Chemistry
Volume271
Issue number45
DOIs
StatePublished - Nov 19 1996

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Imipramine
Cloning
Cytochrome P-450 Enzyme System
Rats
Organism Cloning
Brain
Clone Cells
COS Cells
Complementary DNA
Liver
Proteins
Amino Acids
Hydroxylation
Terminology
Antidepressive Agents
Isoenzymes
Reticulocytes
Immune Sera
Genes
Open Reading Frames

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Genomic cloning and protein expression of a novel rat brain cytochrome P-450 CYP2D18* catalyzing imipramine N-demethylation. / Kawashima, Hidenori; Sequeira, David J.; Nelson, David; Strobel, Henry W.

In: Journal of Biological Chemistry, Vol. 271, No. 45, 19.11.1996, p. 28176-28180.

Research output: Contribution to journalArticle

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