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.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM014033-20
Application #
3268590
Study Section
Physical Biochemistry Study Section (PB)
Project Start
1979-01-01
Project End
1991-01-31
Budget Start
1987-02-01
Budget End
1988-01-31
Support Year
20
Fiscal Year
1987
Total Cost
Indirect Cost
Name
University of Wisconsin Madison
Department
Type
Schools of Medicine
DUNS #
161202122
City
Madison
State
WI
Country
United States
Zip Code
53715
Chung, C H; Woldegiorgis, G; Dai, G et al. (1992) Conferral of malonyl coenzyme A sensitivity to purified rat heart mitochondrial carnitine palmitoyltransferase. Biochemistry 31:9777-83
Chen, M T; Kaufman, L N; Spennetta, T et al. (1992) Effects of high fat-feeding to rats on the interrelationship of body weight, plasma insulin, and fatty acyl-coenzyme A esters in liver and skeletal muscle. Metabolism 41:564-9
Woldegiorgis, G; Fibich, B; Contreras, L et al. (1992) Restoration of malonyl-CoA sensitivity of soluble rat liver mitochondria carnitine palmitoyltransferase by reconstitution with a partially purified malonyl-CoA binding protein. Arch Biochem Biophys 295:348-51
Woldegiorgis, G; Contreras, L; Shrago, E (1990) Characterization of a low molecular weight protein of the ATP synthetase complex from beef heart and rat liver mitochondria with a high affinity monoclonal antibody. Biochem Biophys Res Commun 169:339-45
Kargas, G; Rudy, T; Spennetta, T et al. (1990) Separation and quantitation of long-chain free fatty acids in human serum by high-performance liquid chromatography. J Chromatogr 526:331-40
Ruoho, A E; Woldegiorgis, G; Kobayashi, C et al. (1989) Specific labeling of beef heart mitochondrial ADP/ATP carrier with N-(3-iodo-4-azidophenylpropionamido)cysteinyl- 5-(2'-thiopyridyl)cysteine-coenzyme A (ACT-CoA), a newly synthesized 125I-coenzyme A derivative photolabel. J Biol Chem 264:4168-72
Woldegiorgis, G; Duff, T; Contreras, L et al. (1989) Photoaffinity labeling of hamster brown adipose tissue mitochondria by an [125I] coenzyme A derivative: differential interaction with the uncoupling protein and ADP/ATP carrier. Biochem Biophys Res Commun 161:502-7
Molaparast-Saless, F; Shrago, E; Spennetta, T L et al. (1988) Determination of individual long-chain fatty acyl-CoA esters in heart and skeletal muscle. Lipids 23:490-2
Kim, J H; Woldgiorgis, G; Elson, C E et al. (1988) Age-related changes in respiration coupled to phosphorylation. I. Hepatic mitochondria. Mech Ageing Dev 46:263-77
Mares-Perlman, J A; Shrago, E (1988) Energy substrate utilization in freshly isolated Morris Hepatoma 7777 cells. Cancer Res 48:602-8

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