Evidence for two enzymatic pathways for omega-oxidation of docosanoic acid in rat liver microsomes.
We studied the omega-oxidation of docosanoic acid (C22:0) in rat liver microsomes. C22:0 and 22-hydroxy-docosanoic acid (omega-hydroxy-C22:0) were used as substrates, and the reaction products were analyzed by electrospray ionization mass spectrometry. In the presence of NADPH, omega-oxidation of C22:0 produced not only the hydroxylated product, omega-hydroxy-C22:0, but also the dicarboxylic acid of C22:0, docosanedioic acid (C22:0-DCA). When rat liver microsomes were incubated with omega-hydroxy-C22:0 in the presence of either NAD+ or NADPH, C22:0-DCA was formed readily. Formation of C22:0-DCA from either C22:0 or omega-hydroxy-C22:0 with NADPH as cofactor was inhibited strongly by miconazole and disulfiram, whereas no inhibition was found with NAD+ as cofactor. Furthermore, omega-oxidation of C22:0 was reduced significantly when molecular oxygen was depleted. The high sensitivity toward the more specific cytochrome P450 inhibitors ketoconazole and 17-octadecynoic acid suggests that hydroxylation of C22:0 and omega-hydroxy-C22:0 may be catalyzed by one or more cytochrome P450 hydroxylases belonging to the CYP4A and/or CYP4F subfamily. This study demonstrates that C22:0 is a substrate for the omega-oxidation system in rat liver microsomes and that the product of the first hydroxylation step, omega-hydroxy-C22:0, may undergo further oxidation via two distinct pathways driven by NAD+ or NADPH.
Sanders
Robert-Jan
RJ
Academic Medical Center, University of Amsterdam, Laboratory Genetic Metabolic Diseases, Department of Pediatrics/Emma Children's Hospital, 1105 AZ Amsterdam, The Netherlands.
Ofman
Rob
R
Valianpour
Fredoen
F
Kemp
Stephan
S
Wanders
Ronald J A
RJ
D000818:Animals; D065607:Cytochrome P-450 Enzyme Inhibitors; D005227:Fatty Acids / Q000378:metabolism*; D008862:Microsomes, Liver / Q000378:metabolism*; D010084:Oxidation-Reduction; D051381:Rats; D021241:Spectrometry, Mass, Electrospray Ionization
D065607:Cytochrome P-450 Enzyme Inhibitors; D005227:Fatty Acids; C007547:behenic acid