Demonstration of the occurrence of inactive fatty acid synthetase in rat liver by immunotitration and its in vitro partial activation.
Direct immunotitrations of rat liver fatty acid synthetase in crude tissue homogenates with monospecific rabbit anti-rat liver fatty acid synthetase antibody enabled us to make a comparison of fatty acid synthetase protein and activity (percentage of maximal activity) as a function of the nutritional state in normal, diabetic, and insulin- and glucagon-insulin treated animals. Previous results, in which large changes in fatty acid synthetase activity were related to protein synthesis and degradation rather than to enzyme activation, were confirmed. It was also shown that fatty acid synthetase activation does not occur immediately on synthesis but follows the synthesis of fatty acid synthetase protein. In order to characterize the enzymatically inactive protein found on immunotitration and to develop an in vitro system for fatty acid synthetase activation, conditions were sought to obtain large amounts of fatty acid synthetase protein free from, or low in, activity. It was found that treatment of hypophysectomized rats with triiodothyronine meets these requirements, yielding milligram quantities of inactive fatty acid synthetase protein with less than 2% of maximal activity. A part of the inactive fatty acid synthetase was found to be the apoenzyme as indicated by beta-ketoreductase and thioesterase activities, by its ability to incorporate label from [G3H]CoA, and by its partial in vitro activation, which led to an increase in overall synthetase activity in crude and partially purified cell-free systems. The components required for activation include magnesium ion and a transferase fraction prepared from livers of 48-h fasted, 12-h refed rats.
D000818:Animals; D000936:Antigen-Antibody Complex; D004789:Enzyme Activation; D064429:Fatty Acid Synthases / Q000276:immunology / Q000378:metabolism*; D007106:Immune Sera; D007074:Immunoglobulin G; D008099:Liver / Q000187:drug effects / Q000201:enzymology*; D008297:Male; D010204:Pantetheine / Q000378:metabolism; D051381:Rats; D014284:Triiodothyronine / Q000494:pharmacology
D000936:Antigen-Antibody Complex; D007106:Immune Sera; D007074:Immunoglobulin G; D014284:Triiodothyronine; D010204:Pantetheine; D064429:Fatty Acid Synthases