The Leakage and Destruction of CoA in Isolated Mitochondria

1 In mitochondria from rat liver, heart or kidney, the induction of large amplitude swelling was accompanied by a leakage of endogenous CoA into the incubation medium, almost as rapid as the leakage of pyridine nucleotides. 2,4-Dinitrophenol induced a small leakage of CoA despite the fact that only minor mitochondrial swelling occurred. In kidney mitochondria, swelling and loss of cofactors was pronounced even in an isoosmotic N-tris(hydroxymethyl)-methyl-2-aminoethanesulfonic acid/KCl-medium. 2 When no swelling occurred, endogenous CoA was stable. After the induction of swelling, CoA leaked out and was destroyed. 3 In “lysosome-free” mitochondria, the degradation of CoA after its leakage from mitochondria was much less than in ordinary mitochondrial preparations. Lysosomal contamination thus probably explains most of the breakdown of CoA in experiments with isolated mitochondria. 4 Tissue fractionation studies showed that at least three enzymes able to degrade CoA are present in rat liver. One enzyme is present in the lysosomes and the reaction product is dephospho-CoA. The lysosomal enzyme is extensively inhibited by Pi, and is probably identical with lysosomal acid phosphatase. The other enzymes are present in the microsomal and the nuclear fraction, and are not inhibited by Pi. Enzymes degrading dephospho-CoA are also present in the microsomal and the nuclear fraction, and are not inhibited by Pi. These Pi-insensitive enzymes are probably pyrophosphatases. 5 Pi affects CoA in mitochondrial preparations in at least three different ways. It induces swelling accompanied by a leakage of CoA into the incubation medium. It liberates CoA from CoA esters, mainly long chain acyl-CoA. It inhibits the degradation of CoA by lysosomal acid phosphatase. 6 The degradation of CoA to dephospho-CoA can be reversed by ATP, evidently by a rephosphorylation system present in mitochondrial preparations.