The overall objective of this research project is the elucidation of the mechanisms of regulation of the pyruvate dehydrogenase complex (PDC), primarily from liver and brain. Procedures will be developed for purification of liver and brain pyruvate dehydrogenase complexes. Studies will be conducted to determine the mechanisms by which different metabolites alter the activity of the regulatory enzymes of these complexes, the pyruvate dehydrogenase kinase and pyruvate dehydrogenase phosphatase. New kinetic approaches are proposed for characterizing the regulatory phosphorylation-dephosphorylation cycle. Liver PDC appears to function as a major rate limiting step in fatty acid synthesis from carbohydrate. Studies on liver PDC should give insights into mechanisms by which hormones, such as insulin and glucagon, and different metabolic conditions could elicit changes in the activities of the regulatory components of this complex. Glucose oxidation to CO2 is the major source of energy for brain function. Glucose oxidation in brain is stimulated by an effect of K ion on pyruvate oxidation. Recent data established that K ion markedly enhances inhibition of kidney pyruvate dehydrogenase kinase by ADP. Studies will determine whether this or other effects of K ion on regulation of brain PDC function to modulate this important rate limiting step. Several lines of evidence suggest that PDC may associate with inner mitochondrial membrane. Studies will be conducted to determine whether PDC binds to the inner mitochondrial membrane. If evidence for binding of PDC is obtained, studies will determine whether binding is at a specific site, what is the nature of the association, and what are the effects on the regulatory phosphorylation-dephosphorylation cycle.