The long term objectives of this application are to understand the regulatory mechanisms in intermediary metabolism, particularly in liver and heart which, in turn, account for the pathophysiological derangements occurring during fasting, uncontrolled diabetes mellitus and myocardial ischemia.
The specific aims outlined in this proposal concentrate on one of the more important proteins in cellular bioenergetics, namely the adenine nucleotide translocase (ANT) located in the inner mitochondrial membrane. Experiments will be designed to show that the ANT which is considered by most investigators to be the rate limiting step in overall oxidative phosphorylation is modulated by long chain fatty acyl CoA esters and membrane lipids. The hypothesis to be supported is that acyl CoA esters are natural ligands for the ANT thereby acting as the regulatory mechanism for this critical transport activity. Experiments will be carried out using liver and heart mitochondria, inverted submitochondrial particles and the purified ANT reconstituted into an artificial liposome system. For a number of studies the animals will be pretreated by dietary manipulation, streptozotocin to produce diabetes and thyroidectomy, in order to alter the lipid composition of the inner mitochondrial membrane which, in turn, influences the activity and kinetics of the ANT. Moreover, the ANT will be purified from these treated livers and the concentration of bound acyl CoA to the protein determined by a new method developed in this laboratory using high performance liquid chromatography. The results of these studies should establish conclusively the biochemical verification for modulation of adenine nucleotide transport by long chain acyl CoA esters and how this, in turn, accounts for the physiological and pathophysiological manifestations occurring in uncontrolled diabetes and fasting as well as other metabolic disorders.
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