Function of Carnitine in the Fatty Acid Oxidase-deficient Insect Flight Muscle

Charles C Childress,Bertram Sacktor,David R Traynor

doi:10.1016/s0021-9258(18)96269-1

Charles C Childress, Bertram Sacktor + Show 1 more

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https://doi.org/10.1016/s0021-9258(18)96269-1

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Abstract

Abstract The metabolic function of carnitine in the fatty acid oxidase-deficient flight muscle of the blowfly, Phormia regina, was investigated. Mitochondria from flies did not oxidize palmitate or hexanoate in the presence of adenosine triphosphate, coenzyme A, carnitine, and serum albumin. Palmitoyl carnitine was oxidized at an extremely slow rate, QO2 of 20. The rate of synthesis of the carnitine ester was much slower and is rate limiting in fatty acid oxidation. Hexanoyl carnitine was apparently not metabolized. Flight muscle of flies, however, has a concentration of carnitine of over 4 µmoles per g, wet weight. These findings suggest that the primary function of carnitine in blowfly flight muscle is not related to utilization of fats. Carnitine increased the rate of oxidative decarboxylation of pyruvate-1-14C to acetyl-CoA. The malonate inhibition of the rate of production of 14CO2 from pyruvate-1-14C, but not from pyruvate-3-14C, was relieved by carnitine. The rate of oxidation of pyruvate-3-14C to 14CO2 was inhibited by carnitine. These effects of carnitine may be explained by the formation of acetyl carnitine from carnitine and acetyl-CoA, derived from pyruvate, catalyzed by an active carnitine acetyl transferase. Approximately 90% of the enzyme was found in the mitochondria of the muscle. Acetyl-CoA, in the presence of carnitine, was not oxidized by mitochondria, although acetyl carnitine was oxidized with a QO2 of over 300. This indicates that blowfly mitochondria are not permeable to acetyl-CoA but are permeable to acetyl carnitine. When both pyruvate and acetyl carnitine were available, pyruvate was used preferentially. The concentration of acetyl carnitine in flight muscle of blowflies increased 4-fold on initiation of flight, paralleling the increase in pyruvate. After about 1 min of flight, the level of acetyl carnitine decreased and attained a steady state at a concentration twice that in the muscle at rest. Functions for carnitine in carbohydrate utilization, resulting from the direct action of carnitine on pyruvate metabolism, are proposed. A comparative study of the metabolism of carnitine in blowflies and bees is described.

Highlights

Hexanoyl, and palmitoyl carnitines were synthesized as described by Bremer [15, 5], except that, following neutralization of the reaction mixture, the palmitoyl carnitine was extracted with butanol
In view of the permeability of blowfly mitochondria to carnitine, the possibility that intramitochondrial carnitine leaked into the washing during isolation of the mitochbndria has not been precluded
Carnitine was not detected in the thorax of the bee, indicating a level less than 0.02 pmole per g, wet weight

Summary

Methods

Sodium pyruvate and m-carnitine HCl were obtained from Calbiochem. Hexanoyl, and palmitoyl carnitines were synthesized as described by Bremer [15, 5], except that, following neutralization of the reaction mixture, the palmitoyl carnitine was extracted with butanol. The butanol extract was evaporated to dryness under reduced pressure. Palmitoyl carnitine was redissolved in a small volume of warm butanol and precipitated by addition of several volumes of ether. The product was subsequently recrystallized from butanol-ether and several times from water. M.p. 186”, hexanoyl carnitine, m.p. 145”, and palmitoyl carnitine, m.p. 141”, all reacted positively in the Hestrin test [1]. The Hestrin reaction for palmitoyl carnitine was carried out in ethanol because of the insolubility of palmitoyl hydroxamate. 1.24 f 0.06 (131%) 0.35 f 0.02 (37%) 0.94 f 0.12 (99%)

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