Expression and catalytic activity of mouse leukotriene B4 omega-hydroxylase, CYP4F14.
We have isolated a cDNA for a mouse leukotriene B4 omega-hydroxylase, CYP4F14. The cDNA encoded a protein with 524 amino acids, whose sequence similarity is 95% that of rat CYP4F1. The microsomes from yeast cells transfected with CYP4F14 expression vector showed 0.1 nmol P450/mg protein and catalyzed omega-hydroxylations of leukotriene B4, 6-trans-leukotriene B4, lipoxin A4, prostaglandin A1, and several hydroxyeicosatetraeonic acids (HETEs), with 8-HETE being the most active substrate. In contrast, no activity was detected toward lipoxin B4, laurate, and arachidonate. The mRNA for CYP4F14 had three different 5' untranslated sequences. Analysis of the CYP4F14 gene showed that two exon I sequences with different transcription start sites are located in the gene, and two splicing signals on the 3' end of intron I are alternatively used. The mRNA for this P450 was detected only in the liver by Northern blot analysis, whereas a small amount of the mRNA was detected in the brain using RT-PCR. Administration of clofibrate had no effect on microsomal 6-trans-leukotriene B4 omega-hydroxylase activity, but resulted in a marked reduction in the content of mRNA for this P450 in the liver. These findings indicate that CYP4F14 is very similar to CYP4F1 except for its expression in the brain and 5' untranslated sequences.
Kikuta
Y
Y
Department of Applied Biological Science, Faculty of Engineering, Fukuyama University, Hiroshima, Japan. kikuta@fubac.fukuyama-u.ac.jp
Kasyu
H
H
Kusunose
E
E
Kusunose
M
M
D020121:5' Untranslated Regions; D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism; D001483:Base Sequence; D015152:Blotting, Northern; D001921:Brain / Q000378:metabolism; D002384:Catalysis; D002994:Clofibrate / Q000494:pharmacology; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D000072470:Cytochrome P450 Family 4; D018076:DNA, Complementary / Q000378:metabolism; D005091:Exons; D006893:Hydroxyeicosatetraenoic Acids / Q000378:metabolism; D007438:Introns; D007848:Laurates / Q000378:metabolism; D007975:Leukotriene B4 / Q000378:metabolism; D044045:Lipoxins; D008099:Liver / Q000378:metabolism; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008862:Microsomes, Liver / Q000201:enzymology; D006899:Mixed Function Oxygenases / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D008957:Models, Genetic; D008969:Molecular Sequence Data; D011454:Prostaglandins A / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D014018:Tissue Distribution; D014158:Transcription, Genetic; D014162:Transfection
D020121:5' Untranslated Regions; D018076:DNA, Complementary; D006893:Hydroxyeicosatetraenoic Acids; D007848:Laurates; D044045:Lipoxins; D011454:Prostaglandins A; D012333:RNA, Messenger; D011994:Recombinant Proteins; C040527:lipoxin A4; D007975:Leukotriene B4; D016718:Arachidonic Acid; C047628:8-hydroxyeicosatetraenoic acid; D003577:Cytochrome P-450 Enzyme System; C042651:lipoxin B4; D006899:Mixed Function Oxygenases; D000072470:Cytochrome P450 Family 4; C040780:leukotriene B4 20-hydroxylase; D002994:Clofibrate