Malonyl-Coenzyme A is a key intermediate in the fatty acid biosynthetic pathway. The cardiac tissue, unlike most other tissues, is not known to synthesize fatty acids. However, high levels of malonyl-CoA (4-5 uM) are found in the heart tissue. This project is an investigation of the pathways of formation and degradation of malonyl-CoA in the heart. It has been proposed that malonyl-CoA may serve as an inhibitor of fatty acid oxidation. Since fatty acid oxidation is the key energy yielding pathway for the cardiac tissue, such a control may be very significant. The synthesis of malonyl-CoA by protein fractions derived from heart tissue will be followed by the incorporation of 14C label from 14C bicarbonate into malonyl-CoA. The degradation of malonyl-CoA will be followed by the appearance of 14C carbon dioxide from [1-14C]malonyl-CoA or the appearance of acetyl-CoA, by a coupled spectrophotometric assay using malate dehydrogenase and citrate synthase. The enzymes involved in the synthesis and degradation will be purified by a combination of classical and affinity purification methods. The effect of hormones like epinephrine and insulin (which have pronounced effects on cardiac energetics) on the phosphorylation state and specific activities of heart acetyl-CoA carboxylase and malonyl-CoA decarboxylase will be studied in heart tissues perfused with these hormones. The concentration of malonyl-CoA and the activities of the above enzymes will also be studied in the ischemic heart. Elucidation of the pathways of malonyl-CoA metabolism and its regulation is imperative for a clearer understanding of cardiac energetics and its regulation. Investigation of the regulation of these enzymes will unravel the mechanism of insulin and epinephrine action on cardiac stimulation and lipolysis. The research will also address the relationship between ischemia induced inhibition of cardiac energy output and increased malonyl-CoA levels.
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