Action in vivo and in vitro of 2-tetradecylglycidic acid, 2-tetradecylglycidyl-CoA and 2-tetradecylglycidylcarnitine on hepatic carnitine palmitoyltransferase.
The effects of 2-tetradecylglycidic acid (TDGA), TDGA-CoA and TDGA-carnitine were examined in purified hepatic CPT (carnitine palmitoyltransferase) and in hepatic mitochondria and inverted submitochondrial vesicles derived from Sprague-Dawley rats. Since TDGA has been reported as a specific inhibitor of carnitine palmitoyltransferase-A (CPT-A), the focus was on kinetics and inhibition of CPT-A, and the relationship of this key enzyme to beta-oxidation. After administration of TDGA in vivo to overnight-starved rats, the Vmax. of CPT in intact mitochondria and in inverted vesicles (CPT-B) was depressed by 66%. The S0.5 for palmitoyl-CoA and Km for carnitine were unchanged. The I50 (concn. giving 50% inhibition) for malonyl-CoA was significantly increased from 20 to 141 microM in intact mitochondria, but unchanged (199 versus 268 microM) in inverted vesicles. The addition in vitro of TDGA-CoA (0-1.0 microM) gave I50 values of 0.29 and 0.27 microM (S.E.M. = 0.19) in intact mitochondria from fed and 48 h-starved rats, and 0.81 and 1.57 microM (S.E.M. = 0.29) for inverted vesicles derived from fed and starved rats. Addition in vitro of TDGA-carnitine to mitochondria from starved rats yielded an I50 value of 27.7 mM (S.E.M. = 12.2) for L-[methyl-14C]carnitine release from palmitoyl-L-[methyl-14C]carnitine and 0.64 mM (S.E.M. = 0.07) for palmitoyl-L-[methyl-14C]carnitine formation from L-[methyl-14C]carnitine in intact mitochondria. Inverted vesicles were not measurably sensitive to TDGA-carnitine up to 500 microM for the assay of L-[methyl-14C]carnitine release, but were as sensitive as intact mitochondria when inhibition was determined in the direction of palmitoyl-L-[methyl-14C]carnitine formation (I50 = 0.54 +/- 0.07 microM). When TDGA-CoA was added to intact mitochondria, then incubated for 5 min at room temperature and subsequently washed out, Vmax. of CPT decreased from 5.8 to 3.5 (S.E.M. = 0.6) in intact mitochondria, and from 17.2 to 6.3 (S.E.M. = 4.8) in inverted vesicles. The Km for L-carnitine and the S0.5 for palmitoyl-CoA increased 2-fold with TDGA-CoA pretreatment in both intact mitochondria and inverted vesicles. Detergent solubilization (0.05% Triton X-100) resulted in a complete loss of TDGA-CoA sensitivity (up to 1.0 microM measured). Sonicated mitochondria exhibited an I50 of 0.72 +/- 0.03 microM.(ABSTRACT TRUNCATED AT 400 WORDS)
D000214:Acyl Coenzyme A / Q000031:analogs & derivatives*; D000217:Acyltransferases / Q000037:antagonists & inhibitors*; D000284:Administration, Oral; D000818:Animals; D001384:Azetidines; D002331:Carnitine / Q000031:analogs & derivatives* / Q000494:pharmacology; D002334:Carnitine O-Palmitoyltransferase / Q000037:antagonists & inhibitors*; D004852:Epoxy Compounds / Q000494:pharmacology*; D004988:Ethers, Cyclic / Q000494:pharmacology*; D005227:Fatty Acids / Q000494:pharmacology*; D005260:Female; D005454:Fluorescence Polarization; D007004:Hypoglycemic Agents / Q000494:pharmacology*; D066298:In Vitro Techniques; D007700:Kinetics; D008099:Liver / Q000187:drug effects; D008297:Male; D008316:Malonyl Coenzyme A / Q000494:pharmacology; D008930:Mitochondria, Liver / Q000187:drug effects / Q000201:enzymology*; D010171:Palmitoyl Coenzyme A / Q000031:analogs & derivatives* / Q000494:pharmacology; D051381:Rats; D011919:Rats, Inbred Strains
D000214:Acyl Coenzyme A; D001384:Azetidines; D004852:Epoxy Compounds; D004988:Ethers, Cyclic; D005227:Fatty Acids; D007004:Hypoglycemic Agents; C050896:2-tetradecylglycidylcarnitine; D010171:Palmitoyl Coenzyme A; D008316:Malonyl Coenzyme A; C042420:2-tetradecylglycidyl-coenzyme A; D000217:Acyltransferases; D002334:Carnitine O-Palmitoyltransferase; C021756:2-tetradecylglycidic acid; D002331:Carnitine