4-bromotiglic acid, a novel inhibitor of thiolases and a tool for assessing the cooperation between the membrane-bound and soluble beta-oxidation systems of rat liver mitochondria

An inhibitor of long-chain 3-ketoacyl-CoA thiolase has been developed as a tool for probing the cooperation between the two fatty acid beta-oxidation systems located in the inner mitochondrial membrane and in the mitochondrial matrix, respectively. 4-Bromotiglic acid was synthesized and found to inhibit palmitoylcarnitine-supported respiration of rat liver mitochondria in concentration-dependent and time-dependent fashions. Complete inhibition of respiration was achieved after incubating coupled mitochondria with 10 microM 4-bromotiglic acid for 2 min. Uncoupled mitochondria were resistant to the toxic effect of the inhibitor. Inhibition of octanoate-supported or octanoylcarnitine-supported respiration was partially reversed when the inhibitor was removed from the incubation medium. Such reversal was not observed with either palmitoylcarnitine or 2-methyldecanoic acid as the respiratory substrate. The severity of the irreversible inhibition declined with decreasing chain length of the acylcarnitine substrate. Of all beta-oxidation enzymes, only thiolases were inactivated by the inhibitor. Under conditions at which acetoacetyl-CoA thiolase and long-chain thiolase were completely inactivated, 3-ketoacyl-CoA thiolase retained some activity. It is concluded that the degradation of palmitic acid and longer-chain fatty acids is initiated by the beta-oxidation system of the inner membrane, whereas fatty acids shorter than palmitic acid can be oxidized to a certain degree by the matrix system alone. The effectiveness of the matrix system increases with decreasing chain length of the substrate.