Identification of CYP2C23 expressed in rat kidney as an arachidonic acid epoxygenase.
A cDNA was isolated from a rat kidney cDNA library using mixed probes of CYP (cytochrome P450) 2C6, 2C7, 2C8, 2C9 and 2C18 cDNAs. The 3'-terminal and 5'-terminal regions of the cDNA were sequenced and were identical with those of cytochrome P450 2C23 (CYP2C23) except for a one-base deletion and a one-base addition in coding region. These changes caused a frame shift and changed the deduced amino acid sequence relative to the previously published sequence. This cDNA was expressed using a baculovirus expression system, and the resultant P450 had a lambda max of 450 nm when reduced and complexed with carbon monoxide. Specific content of the expressed P450 ranged from 0.27 to 0.43 nmol/mg of cell lysate protein. Arachidonic acid metabolism catalyzed by expressed CYP2C23 indicated that CYP2C23 efficiently produced epoxyeicosatrienoic acids (EETs). These EETs were characterized further by gas-liquid chromatography/negative ion chemical ionization mass spectrometry (GC/NCIMS) and were found to include 8,9-EET, 11,12-EET and 14,15-EET in a ratio of 1:2:1. No 5,6-EET was detected. A low rate of lauric acid hydroxylation at the (omega-1)-position was found, but the enzyme was unable to metabolize prostaglandin E1. These studies suggest that CYP2C23 is responsible, in part, for the production of EETs in rat kidney.
Imaoka
S
S
Laboratory of Molecular Carcinogenesis, National Cancer Institute, National Institutes of Health, Bethesda, Maryland.
Wedlund
P J
PJ
Ogawa
H
H
Kimura
S
S
Gonzalez
F J
FJ
Kim
H Y
HY
D000527:Alprostadil / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism; D001483:Base Sequence; D003001:Cloning, Molecular; D000090762:Cytochrome P-450 CYP2J2; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics* / Q000378:metabolism*; D015342:DNA Probes; D018076:DNA, Complementary / Q000235:genetics / Q000302:isolation & purification; D015870:Gene Expression / Q000235:genetics; D007668:Kidney / Q000201:enzymology* / Q000502:physiology*; D007850:Lauric Acids / Q000378:metabolism; D008969:Molecular Sequence Data; D010105:Oxygenases / Q000235:genetics* / Q000378:metabolism*; D051381:Rats; D013053:Spectrophotometry
C000717351:Cyp2c23 protein, rat; D015342:DNA Probes; D018076:DNA, Complementary; D007850:Lauric Acids; C030358:lauric acid; D016718:Arachidonic Acid; D003577:Cytochrome P-450 Enzyme System; D010105:Oxygenases; D000090762:Cytochrome P-450 CYP2J2; D000527:Alprostadil