Nafenopin-, ciprofibroyl-, and palmitoyl-CoA conjugation in vitro: kinetic and molecular characterization of marmoset liver microsomes and expressed MLCL1.
Acyl-CoA conjugation of xenobiotic carboxylic acids is catalyzed by hepatic microsomal long-chain fatty acid CoA ligases (LCL, EC 6.2.1.3). Marmosets (Callithrix jacchus) are considered genetically closer to humans than rodents and are used in pharmacological and toxicological studies. We have demonstrated that marmoset liver microsomes catalyze nafenopin-, ciprofibroyl-, and palmitoyl-CoA conjugation and that only palmitoyl-CoA conjugation is significantly upregulated (1.7-fold, P < 0.02) by a high fat diet. Additionally, the apparent C(50) values for nafenopin-, ciprofibroyl-, and palmitoyl-CoA conjugation of 149.7, 413.4, and 3.4 microM were comparable to those reported for human liver microsomes viz, 213.7, 379.8, and 3.4 microM, respectively. Comparison with human data was enabled by the cloning of a full-length marmoset cDNA (MLCL1) that encoded a 698-amino-acid protein sharing 83% similarity with rat liver acyl-CoA synthetase (ACS1) and 93 and 90% similarity with human liver LCL1 and LCL2, respectively. MLCL1 transiently expressed in COS-7 cells activated nafenopin (C(50) 192.9 microM), ciprofibrate (C(50) 168.7 microM), and palmitic acid (C(50) 4.5 microM) to their respective CoA conjugates. This study also demonstrated that the sigmoidal kinetics observed for nafenopin- and ciprofibroyl-CoA conjugation were not unique to human liver microsomes but were also characteristic of marmoset liver microsomes and recombinant MLCL1. More extensive characterization of the substrate specificity of marmoset LCL isoforms will aid in determining further the suitability of marmosets as a model for human xenobiotic metabolism via acyl-CoA conjugation.
Drogemuller
C J
CJ
Department of Clinical Pharmacology, School of Medicine, Adelaide, 5042, Australia.
Nunthasomboon
S
S
Knights
K M
KM
D000595:Amino Acid Sequence; D000818:Animals; D002144:Callithrix / Q000235:genetics*; D002995:Clofibric Acid / Q000031:analogs & derivatives* / Q000378:metabolism*; D003066:Coenzyme A Ligases / Q000235:genetics* / Q000378:metabolism*; D058607:Fibric Acids; D006801:Humans; D008297:Male; D008862:Microsomes, Liver / Q000378:metabolism*; D008969:Molecular Sequence Data; D009255:Nafenopin / Q000378:metabolism*; D019308:Palmitic Acid / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity; D015262:Xenobiotics / Q000378:metabolism; D000098859:Long-Chain-Fatty-Acid-CoA Ligase
D058607:Fibric Acids; D011994:Recombinant Proteins; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D015262:Xenobiotics; D009255:Nafenopin; D019308:Palmitic Acid; D002995:Clofibric Acid; D003066:Coenzyme A Ligases; C090799:FAA2 protein, S cerevisiae; D000098859:Long-Chain-Fatty-Acid-CoA Ligase; C019304:ciprofibrate