Mechanism of carnitine acylcarnitine translocase-catalyzed import of acylcarnitines into mitochondria.

Mitochondrial imports of acylcarnitine and carnitine have been measured by new methods based on the monitoring of deacylation of acylcarnitines and the acetylation of carnitine in the matrix, subsequent to their entry. These methods have shown higher import rates than those calculated from the uptake of radioactive carnitines into mitochondria as employed until presently. This new approach has permitted the import of long chain acylcarnitine to be followed unambiguously; the results have confirmed that the carnitine acylcarnitine translocase is indeed involved in this import which also proceeds by a mole to mole exchange-diffusion against internal carnitine. Depletion of matrix carnitine greatly decreased the substrate import rates based on their uptake assay but much less so when the deacylation and acylation techniques were employed to monitor imports. These results have revealed that there is a small pool of carnitine in the matrix which readily equilibrates with the medium carnitine through the translocase but which equilibrates with the larger matrix carnitine pool slowly. This finding has necessitated reinterpretation of several previous observations on the translocase that were based on the assumption of a single matrix carnitine pool and in which the translocase was assumed to constitute the rate-limiting step in the activity measurements.