J Mol Cell Cardiol 17, 1161-1171 (1985) Regulation ofPyruvate Dehydrogenase During Infusion of Fatty Acids of Varying Chain Lengths in the Perfused Rat Heart Pirjo M. Latipfifi, Keijo J. Peuhkurinen, J. Kalervo Hiltunen and Ilmo E. Hassinen Department of Medical Biochemistry, University of Oulu, SF-90220 Oulu 22, Finland (Received October 1984, acceptedin revised form 5 February 1985) P. M. LA'rIPA,~, K . J . PEUHKURINEN, J. K. HILTUNENAND I. E. HASSINEN, Regulation of Pyruvate Dehydroge- nase During Infusion of Fatty Acids of Varying Chain Lengths in the Perfused Rat Heart. Journalof~lolecular and Cellular Cardiology (1985) 17, 1161-1171. The effects of a homologous series of fatty acids with a chain length of two to eight on the rate of pyruvate oxidation and covalent interconversions of the pyruvate dehydrogenase complex (PDH) were studied in isolated perfused rat hearts. In the Langendorff-pcrfused heart beating at 5 Hz against an aortic pressure of 59 mmHg (7,85 kPa), a positive linear correlation was found between the fraction of PDH existing in the active non-phosphorylated form of pyruvate dehydrogenase complex (PDHa) and the pyruvate oxidation rate until the PDH a fraction increased to 48%. This value resulted in a saturation of the citric acid cycle and further activation did not increase the metabolic flux. The PDH a content of the tissue was higher during infusion of odd carbon number fatty acids than during infusion of even carbon number fatty acids. Propionate caused an almost maximal (93 %) activation of PDH. A negative correlation Was found between the mitochondrial NADH/NAD + ratio and the PDH, content, A negative correlation was also found between the acetyl-CoA/CoA ratio and the tissue PDH acontent. The rate of labelled CO 2 production, the specific radioactivity of tissue alanine and the metabolic balance sheet demonstrated that the alanine aminotransferase reaction in the total tissue does not reach equilibrium with the mitochondrial pyruvate pool during propionate oxidation, but the equilibrium is reached during the oxidation of even-number carbon fatty acids. This suggests that pyruvate is formed from propionate-derived metabolites also in the cytosol, although the primary metabolism ofpropionate occurs in the mitochondria. The results indicate that the rate of pyruvate oxidation in the myocardium is mainly regulated by covalent interconversion of PDH. During propionate oxidation the PDH, content in the tissue can increase beyond the point of saturation of the citric acid cycle and this indicates that feedback inhibition of the enzyme is rate- determining under these conditions. KEY WORDS: Pyruvate oxidation; Covalent enzyme modification; Odd-numbered fatty acids; Metabolic com- partmentation. Introduction Control of the pyruvate dehydrogenase com- plex (PDH) is based on two different types of mechanism. The products, NADH and acetyl-CoA, act as competitive inhibitors of the reaction with respect to NAD + and CoA respectively [4, 8, 31], while the second type of regulation involves covalent modification of the enzyme complex [20, 23, 35]. Pyruvate dehydrogenase can exist in an inactive phos- phorylated form or an active non- phosphorylated form (PDHa) , interconversion between the two forms being catalyzed by a protein kinase requiring MgATP and by a phosphatase. Furthermore, it has been shown that NADH and acetyl-CoA stimulate the PDH kinase reaction, while free CoA and NAD § inhibit the kinase and NADH inhibits the PDH phosphatase [-1, 27]. Thus the NADH/NAD + and acetyl-CoA/CoA ratios are able to influence the PDH reaction at two levels of organization, direct product inhibi- tion and enzyme interconversion. Fatty acids or ketone bodies can convert pyruvate dehy- Address for correspondence: Ilmo Hassinen, Department of Medical Biochemistry, University of Oulu, Kajaanintie 52 A, SF-90220 Oulu 22, Finland. Abbreviation : PDH~, active (non-phosphorylated) form ofpyruvate dehydrogenase complex. 0022 2828/85/121161 + 11 $03.00/0 9 1985 Academic Press Inc. (London) Limited