Enzymes of carbohydrate and fat metabolism in anti-insulin serum diabetes; inactivation by free fatty acids and the protective effect of cellular protein

The possibility that free fatty acids (FFA) and their CoA esters may directly inhibit the activity of enzymes of glycolysis and lipogenesis was studied in liver and adipose tissue of acutely diabetic rats. Despite a marked elevation in tissue FFA, the activity of glucose-6phosphate dehydrogenase, phosphofructokinase, pyruvate kinase, α-glycerophosphate dehydrogenase, aldolase, citrate synthetase, and several other enzymes was not affected 3 or 6 h after anti-insulin serum injection. The activities of hepatic and adipose tissue acetyl-CoA carboxylase, hepatic glucokinase and adipose tissue hexokinase were decreased. The tissue FFA levels were compared to linoleate concentrations required for halfmaximal inhibition (Ki) of several enzymes in tissue fractions afterin vitro contact of 30 min at 37 °C. Linoleate irreversibly inactivated the enzymes to a varying degree but its effect was dependent on the protein content of the system. The Ki of linoleate increased linearly with cellular protein concentration; when extrapolated to the protein level in the intact cell it became unphysiologically high, markedly exceeding the liver or adipose tissue FFA concentration of acutely diabetic rats. The Ki linoleate and the actual FFA concentrations were particularly discrepant in the cytoplasmic compartment, comprising roost of the investigated enzymes, but only a small proportion of tissue FFA, as determined by measurements of intracellular FFA distribution. Similar discrepancy was found for the interaction of palmityl-CoA with glucose-6-phosphate dehydrogenase and acetyl-CoA carboxylase. Thus, the selective decrease in the activity of glucokinase and acetyl-CoA carboxylase in acute diabetes was attributed to factors other than FFA or their CoA esters. The nature of FFA-enzyme interaction, causing enzyme inactivation rather than specific inhibition is discussed, concluding that FFA do not act as direct homeostatic modifiers of enzymes regulating carbohydrate and fat metabolism.