title: Cloning and expression of three rabbit kidney cDNAs encoding lauric acid omega-hydroxylases.
abstract: cDNAs encoding three cytochrome P-450 enzymes were cloned from a rabbit kidney cDNA library. These three cDNAs exhibit greater than 90% nucleotide sequence identity across the coding region. This degree of sequence identity is also seen with P450IVA4, an enzyme that catalyzes the omega-hydroxylation of prostaglandins and that is elevated during pregnancy and induced by progesterone in rabbit lung. The 3' untranslated regions of the three cDNAs display very little sequence identity, suggesting that they are the products of distinct genes. The predicted amino acid sequences derived from each cDNA and for P450IVA4 exhibit about 85% identity. Each cDNA was inserted into an expression vector for transient transfection of COS-1 cells. The transfected cells each expressed a protein recognized by antibodies to P450IVA4. Microsomes isolated from the cells transfected with each cDNA efficiently catalyzed the omega-hydroxylation of lauric acid with rates that greatly exceed that catalyzed by microsomes isolated from the host cell line. One of the cDNAs encodes an enzyme that omega-hydroxylates prostaglandin A1; however, the specific activity was 2 orders of magnitude lower than that for lauric acid. Our results indicate that the substrate selectivity of the kidney P-450s encoded by these cDNAs is distinct from that of the lung P450IVA4 and that multiple enzymes comprise P-450 class IVA in the rabbit.
authors.affiliation: Department of Molecular and Experimental Medicine, Research Institute of Scripps Clinic, La Jolla, California 92037.
authors.forename: E F
authors.initials: EF
authors.lastname: Johnson
authors.forename: D L
authors.initials: DL
authors.lastname: Walker
authors.forename: K J
authors.initials: KJ
authors.lastname: Griffin
authors.forename: J E
authors.initials: JE
authors.lastname: Clark
authors.forename: R T
authors.initials: RT
authors.lastname: Okita
authors.forename: A S
authors.initials: AS
authors.lastname: Muerhoff
authors.forename: B S
authors.initials: BS
authors.lastname: Masters
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D042926:Cytochrome P-450 CYP4A; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics*; D004247:DNA / Q000235:genetics*; D015870:Gene Expression; D015698:Genomic Library; D007527:Isoenzymes / Q000235:genetics; D007668:Kidney / Q000201:enzymology / Q000502:physiology; D008168:Lung / Q000201:enzymology / Q000502:physiology; D051379:Mice; D008807:Mice, Inbred BALB C; D006899:Mixed Function Oxygenases / Q000235:genetics*; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D010957:Plasmids; D011817:Rabbits; D014162:Transfection
chemical_list: D007527:Isoenzymes; D004247:DNA; D003577:Cytochrome P-450 Enzyme System; D006899:Mixed Function Oxygenases; D042926:Cytochrome P-450 CYP4A