Publications

• Molecular cloning and expression of CYP2J2, a human cytochrome P450 arachidonic acid epoxygenase highly expressed in heart.

  Wu S., Moomaw C.R., Tomer K.B., Falck J.R., Zeldin D.C.

  A cDNA encoding a human cytochrome P450 arachidonic acid epoxygenase was isolated from a human liver cDNA library. Sequence analysis revealed that this 1,876-base pair cDNA contained an open reading frame and encoded a new 502-amino acid protein designated CYP2J2. Blot hybridization analysis of RN ... >> More

  A cDNA encoding a human cytochrome P450 arachidonic acid epoxygenase was isolated from a human liver cDNA library. Sequence analysis revealed that this 1,876-base pair cDNA contained an open reading frame and encoded a new 502-amino acid protein designated CYP2J2. Blot hybridization analysis of RNA prepared from human tissues revealed that CYP2J2 was highly expressed in the heart. Recombinant CYP2J2 protein was prepared using the baculovirus expression system and purified to near electrophoretic homogeneity. The enzyme metabolized arachidonic acid predominantly via olefin epoxidation to all four regioisomeric cis-epoxyeicosatrienoic acids (catalytic turnover 65 pmol of product formed/nmol of cytochrome P450/min at 30 degrees C). Epoxidation of arachidonic acid by CYP2J2 at the 14,15-olefin was highly enantioselective for (14R, 15S)-epoxyeicosatrienoic acid (76% optical purity). Immunoblotting of microsomal fractions prepared from human tissues using a polyclonal antibody raised against the recombinant hemoprotein confirmed primary expression of CYP2J2 protein in human heart. The in vivo significance of CYP2J2 was suggested by documenting the presence of epoxyeicosatrienoic acids in the human heart using gas chromatography/mass spectroscopy. Importantly, the chirality of CYP2J2 products matched that of the epoxyeicosatrienoic acid enantiomers present, in vivo, in human heart. We propose that CYP2J2 is one of the enzymes responsible for epoxidation of endogenous arachidonic acid pools in human heart and that epoxyeicosatrienoic acids may, therefore, play important functional roles in cardiac physiology. << Less

  J. Biol. Chem. 271:3460-3468(1996) [PubMed] [EuropePMC]

  This publication is cited by 6 other entries.

• The kidney cytochrome P-450 2C23 arachidonic acid epoxygenase is upregulated during dietary salt loading.

  Holla V.R., Makita K., Zaphiropoulos P.G., Capdevila J.H.

  Excess dietary salt intake induces the activity of the kidney arachidonate epoxygenase and markedly increases the urinary excretion of its metabolites. The epoxyeicosatrienoic acids, products of the kidney P-450 arachidonate epoxygenase, inhibit distal nephron Na(+) reabsorption. Nucleic acid hybr ... >> More

  Excess dietary salt intake induces the activity of the kidney arachidonate epoxygenase and markedly increases the urinary excretion of its metabolites. The epoxyeicosatrienoic acids, products of the kidney P-450 arachidonate epoxygenase, inhibit distal nephron Na(+) reabsorption. Nucleic acid hybridization studies demonstrated the expression of P-450s 2C23, 2C24, and 2C11 as the predominant kidney 2C isoforms and the lack of significant dietary salt-dependent transcriptional regulation of these proteins. Recombinant P-450s 2C11, 2C23, and 2C24 catalyze arachidonate metabolism to mixtures of epoxy- and monohydroxylated acids. Whereas the arachidonate 11,12-olefin was the preferred target for epoxidation by P-450 2C23 (57% of total products), P-450s 2C11 and 2C24 epoxidized the 11,12-olefins and 14,15-olefins with nearly equal efficiency. Stereochemical comparisons demonstrated that the regiochemical and enantiofacial selectivity of P-450 2C23 matched that of the kidney microsomal epoxygenase and that excess dietary salt does not alter the regiochemical or stereochemical selectivity of the kidney arachidonate epoxygenase. Inhibition and immunoelectrophoresis experiments using antibodies raised against recombinant P-450s 2C11 and 2C23 demonstrated that P-450 2C23 is the major 2C arachidonic acid epoxygenase in the rat kidney and the renal P-450 isoform regulated by excess dietary salt intake. << Less

  J. Clin. Invest. 104:751-760(1999) [PubMed] [EuropePMC]

  This publication is cited by 5 other entries.

• Molecular cloning, expression and characterization of an endogenous human cytochrome P450 arachidonic acid epoxygenase isoform.

  Zeldin D.C., DuBois R.N., Falck J.R., Capdevila J.H.

  A cDNA containing an open reading frame coding for a human cytochrome P450 arachidonic acid epoxygenase was isolated from a male human kidney cDNA library. Sequence analysis showed that, with few exceptions, this cDNA was nearly identical to the published sequence for human liver Cyp 2C8 (S. T. Ok ... >> More

  A cDNA containing an open reading frame coding for a human cytochrome P450 arachidonic acid epoxygenase was isolated from a male human kidney cDNA library. Sequence analysis showed that, with few exceptions, this cDNA was nearly identical to the published sequence for human liver Cyp 2C8 (S. T. Okino et al., 1987, J. Biol. Chem. 262, 16072-16079) and encoded a polypeptide of 490 amino acids. Nucleic acid hybridization indicated that: (a) Cyp 2C8 and 2C10 were expressed at comparable levels in the human liver and (b) compared to Cyp 2C10, the steady state concentrations of Cyp 2C8 transcripts in the human kidney were substantially lower. The kidney 2C8 cDNA was cloned into a pBlue BacIII vector, expressed using a baculovirus/Sf9 insect cell system, and the recombinant Cyp 2C8 protein was purified by a combination of hydrophobic and hydroxylapatite chromatography. Purified recombinant Cyp 2C8 and 2C10 were reconstituted in the presence of NADPH and NADPH-cytochrome P450 reductase and shown to metabolize arachidonic via olefin epoxidation with both proteins generating, almost exclusively, epoxygenase-derived products (94 and 90% of total products, respectively). Catalytic turnover (1.05 and 0.75 nmol of product/nmol of hemoprotein/min at 30 degrees C for Cyp 2C8 and 2C10, respectively) was inhibited by the addition of purified cytochrome b5. Metabolism by recombinant 2C8 was both regio- and enantioselective for 11(R), 12(S)- and 14(R), 15(S)-epoxyeicosatrienoic acids (82% optical purity, each). Compared to Cyp 2C8, arachidonic acid epoxidation by Cyp 2C10 was less regio- and stereo-selective and generated mixtures of 8(S), 9(R)-, 11(S), 12(R)-, and 14(R), 15(S)-epoxyeicosatrienoic acids (with optical purities of 66, 69, 63%, respectively). Importantly, recombinant Cyp 2C8 and 2C10 epoxidized the arachidonic acid 11, 12-olefin with opposite enantiofacial selectivities. Only for Cyp 2C8 did the chirality of the products match that of the enantiomers present, in vivo, in human kidney cortex (A. Karara et al., 1990, FEBS Lett. 268, 227-230). Hence, we propose that Cyp 2C8 is one of the human cytochrome P450 isoforms responsible for the metabolism of endogenous arachidonic acid pools. << Less

  Arch. Biochem. Biophys. 322:76-86(1995) [PubMed] [EuropePMC]

  This publication is cited by 5 other entries.