title: Physiological covalent regulation of rat liver branched-chain alpha-ketoacid dehydrogenase.
abstract: A radiochemical assay was developed for measuring branched-chain alpha-ketoacid dehydrogenase activity of Triton X-100 extracts of freeze-clamped rat liver. The proportion of active (dephosphorylated) enzyme was determined by measuring enzyme activities before and after activation of the complex with a broad-specificity phosphoprotein phosphatase. Hepatic branched-chain alpha-ketoacid dehydrogenase activity in normal male Wistar rats was 97% active but decreased to 33% active after 2 days on low-protein (8%) diet and to 13% active after 4 days on the same diet. Restricting protein intake of lean and obese female Zucker rats also caused inactivation of hepatic branched-chain alpha-ketoacid dehydrogenase complex. Essentially all of the enzyme was in the active state in rats maintained for 14 days on either 30 or 50% protein diets. This was also the case for rats maintained on a commercial chow diet (minimum 23% protein). However, maintaining rats on 20, 8, and 0% protein diets decreased the percentage of the active form of the enzyme to 58, 10, and 7% of the total, respectively. Fasting of chow-fed rats for 48 h had no effect on the activity state of hepatic branched-chain alpha-ketoacid dehydrogenase, i.e., 93% of the enzyme remained in the active state compared to 97% for chow-fed rats. However, hepatic enzyme of rats maintained on 8% protein diet was 10% active before starvation and 83% active after 2 days of starvation. Thus, dietary protein deficiency results in inactivation of hepatic branched-chain alpha-ketoacid dehydrogenase complex, presumably as a consequence of low hepatic levels of branched-chain alpha-ketoacids, established inhibitors of branched-chain alpha-ketoacid dehydrogenase kinase. With rats fed a low-protein diet and subsequently starved, inhibition of branched-chain alpha-ketoacid dehydrogenase kinase by branched-chain alpha-ketoacids generated as a consequence of endogenous proteolysis most likely promotes the greater branched-chain alpha-ketoacid dehydrogenase activity state.
mesh_terms: D042942:3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide); D000818:Animals; D004044:Dietary Proteins / Q000008:administration & dosage / Q000494:pharmacology*; D005260:Female; D007658:Ketone Oxidoreductases / Q000378:metabolism*; D007930:Leucine / Q000494:pharmacology; D008099:Liver / Q000201:enzymology*; D008297:Male; D009097:Multienzyme Complexes / Q000378:metabolism*; D009765:Obesity / Q000201:enzymology; D011854:Radiochemistry; D051381:Rats; D011919:Rats, Inbred Strains; D011924:Rats, Zucker; D013217:Starvation / Q000201:enzymology*; D013696:Temperature
chemical_list: D004044:Dietary Proteins; D009097:Multienzyme Complexes; D007658:Ketone Oxidoreductases; D042942:3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide); D007930:Leucine