title: Identification of a novel malonyl-CoA IC(50) for CPT-I: implications for predicting in vivo fatty acid oxidation rates.
abstract: Published values regarding the sensitivity (IC(50)) of CPT-I (carnitine palmitoyltransferase I) to M-CoA (malonyl-CoA) inhibition in isolated mitochondria are inconsistent with predicted in vivo rates of fatty acid oxidation. Therefore we have re-examined M-CoA inhibition kinetics under various P-CoA (palmitoyl-CoA) concentrations in both isolated mitochondria and PMFs (permeabilized muscle fibres). PMFs have an 18-fold higher IC(50) (0.61 compared with 0.034 μM) in the presence of 25 μM P-CoA and a 13-fold higher IC(50) (6.3 compared with 0.49 μM) in the presence of 150 μM P-CoA compared with isolated mitochondria. M-CoA inhibition kinetics determined in PMFs predicts that CPT-I activity is inhibited by 33% in resting muscle compared with >95% in isolated mitochondria. Additionally, the ability of M-CoA to inhibit CPT-I appears to be dependent on P-CoA concentration, as the relative inhibitory capacity of M-CoA is decreased with increasing P-CoA concentrations. Altogether, the use of PMFs appears to provide an M-CoA IC(50) that better reflects the predicted in vivo rates of fatty acid oxidation. These findings also demonstrate that the ratio of [P-CoA]/[M-CoA] is critical for regulating CPT-I activity and may partially rectify the in vivo disconnect between M-CoA content and CPT-I flux within the context of exercise and Type 2 diabetes.
authors.affiliation: Department of Human Health and Nutritional Sciences, University of Guelph, Guelph, ON, Canada. smithb@uoguelph.ca
authors.forename: Brennan K
authors.initials: BK
authors.lastname: Smith
authors.forename: Christopher G R
authors.initials: CG
authors.lastname: Perry
authors.forename: Timothy R
authors.initials: TR
authors.lastname: Koves
authors.forename: David C
authors.initials: DC
authors.lastname: Wright
authors.forename: Jeffrey C
authors.initials: JC
authors.lastname: Smith
authors.forename: P Darrell
authors.initials: PD
authors.lastname: Neufer
authors.forename: Deborah M
authors.initials: DM
authors.lastname: Muoio
authors.forename: Graham P
authors.initials: GP
authors.lastname: Holloway
mesh_terms: D000818:Animals; D002331:Carnitine / Q000378:metabolism; D002334:Carnitine O-Palmitoyltransferase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002463:Cell Membrane Permeability; D004305:Dose-Response Relationship, Drug; D005227:Fatty Acids / Q000378:metabolism*; D020128:Inhibitory Concentration 50; D007700:Kinetics; D008316:Malonyl Coenzyme A / Q000378:metabolism / Q000494:pharmacology*; D008931:Mitochondria, Muscle / Q000201:enzymology* / Q000378:metabolism; D018657:Muscle Fibers, Slow-Twitch / Q000201:enzymology / Q000378:metabolism; D018482:Muscle, Skeletal / Q000201:enzymology; D010084:Oxidation-Reduction; D010101:Oxygen Consumption; D010171:Palmitoyl Coenzyme A / Q000378:metabolism; D010805:Physical Conditioning, Animal; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D005227:Fatty Acids; D010171:Palmitoyl Coenzyme A; D008316:Malonyl Coenzyme A; D002334:Carnitine O-Palmitoyltransferase; D002331:Carnitine