Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.11.2 (PDK1)
 2,238 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The pyruvate dehydrogenase complex was purified to homogeneity from bakers' yeast (Saccharomyces cerevisiae). No pyruvate dehydrogenase kinase activity was detected at any stage of the purification. However, the purified pyruvate dehydrogenase complex was phosphorylated and inactivated with purified pyruvate dehydrogenase kinase from bovine kidney. The protein-bound radioactivity was localized in the pyruvate dehydrogenase alpha subunit. The phosphorylated, inactive pyruvate dehydrogenase complex was dephosphorylated and reactivated with purified pyruvate dehydrogenase phosphatase from bovine heart. Tryptic digestion of the 32P-labeled complex yielded a single phosphopeptide, which was purified to homogeneity. The sequence of the phosphopeptide was established to be Tyr-Gly-Gly-His-Ser(P)-Met-Ser-Asp-Pro-Gly-Thr-Thr-Tyr-Arg. This sequence is very similar to the sequence of a tryptic phosphotetradecapeptide derived from the alpha subunit of bovine kidney and heart pyruvate dehydrogenase: Tyr-His-Gly-His-Ser(P)-Met-Ser-Asp-Pro-Gly-Val-Ser-Tyr-Arg.
 ...
PMID:Phosphorylation-dephosphorylation of pyruvate dehydrogenase from bakers' yeast. 353 83

The pyruvate dehydrogenase kinase consists of a catalytic subunit (Kc) and a basic subunit (Kb) which appear to be anchored to the dihydrolipoyl transacetylase core component (E2) by another subunit, referred to as protein X (Rahmatullah, M., Jilka, J. M., Radke, G. A., and Roche, T. E. (1986) J. Biol. Chem. 261, 6515-6523). We determined the catalytic requirements for reduction and acetylation of the lipoyl moiety in protein X and linked those changes in protein X to regulatory effects on kinase activity. Using fractions prepared by resolution and proteolytic treatments, we evaluated which subunits are required for regulatory effects on kinase activity. With X-KcKb fraction (treated to remove the mercurial agent used in its preparation), we found that the resolved pyruvate dehydrogenase component, the isolated inner domain of E2 (lacking the lipoyl-bearing region of E2), and the dihydrolipoyl dehydrogenase component directly utilize protein X as a substrate. The resulting reduction and acetylation of protein X occurs in association with enhancement of kinase activity. Following tryptic cleavage of E2 and protein X into subdomains, full acetylation of the lipoyl-bearing subdomains of these proteins is retained along with the capacity of acetylating substrates to stimulate kinase activity. All kinase-containing fractions, including those in which the Kb subunit was digested, were inhibited by pyruvate or ADP, alone, and synergistically by the combination suggesting that pyruvate and ADP bind to Kc. Our results suggest that the Kb subunit of the kinase does not contribute to the observed regulatory effects. A dynamic role of protein X in attenuating kinase activity based on changes in the mitochondrial redox and acetylating potentials is considered.
 ...
PMID:The catalytic requirements for reduction and acetylation of protein X and the related regulation of various forms of resolved pyruvate dehydrogenase kinase. 361 Oct 60

Studies were conducted on four pyruvate dehydrogenase kinase-containing fractions: purified pyruvate dehydrogenase complex, the dihydrolipoyl transacetylase-protein X-kinase subcomplex (E2.X.K), a kinase fraction (K fraction) prepared from the E2.X.K subcomplex, and a kinase fraction generated by limited trypsin-digestion of E2.X.K. We characterized the gel electrophoresis properties of dissociated subunits (one-dimensional and two-dimensional), the catalytic and ATP binding properties of kinase-containing fractions, and the subunit requirements for kinase binding to and being activated by the transacetylase-protein X subcomplex (E2.X). A significant portion of protein X was retained with the transacetylase core following release of virtually all the kinase. The K fraction had four major bands separated by sodium dodecyl sulfate-slab gel electrophoresis which corresponded to the dihydrolipoyl dehydrogenase, protein X, the trypsin-resistant catalytic subunit of the kinase and a chymotrypsin-resistant subunit which had a high pI and comigrated in one-dimensional systems with the chymotrypsin-sensitive alpha-subunit of the pyruvate dehydrogenase component. While purified kidney complex contained only about three molecules of kinase (determined by [14C]ATP binding), one molecule of E2.X subcomplex activated a large number (greater than 15) molecules of kinase associated with the protein X-containing K fraction. Sephadex G-200 chromatography of the K fraction in the presence of dithiothreitol led to coelution of protein X and kinase subunits. Limited trypsin digestion converted the transacetylase into subdomains and cleaved protein X and the high pI subunit of the kinase. Under those conditions, the intact catalytic subunit of the kinase did not bind to the large inner domain of the transacetylase but could be activated by untreated E2.X subcomplex. Thus, binding of the catalytic subunit of the kinase and its activation by E2.X required either protein X or the lipoyl-bearing outer domain of the transacetylase. In combination, our results suggest that protein X serves to anchor the kinase to the core of the complex.
 ...
PMID:Properties of the pyruvate dehydrogenase kinase bound to and separated from the dihydrolipoyl transacetylase-protein X subcomplex and evidence for binding of the kinase to protein X. 370 Apr 4

The activity of pyruvate dehydrogenase kinase in extracts of mitochondria from rat hepatocytes cultured for 21 h in medium 199 was increased 2.5-fold by the presence of 55 nM-glucagon and 1 mM-sodium n-octanoate in the culture medium. The change was comparable with that induced in vivo by 48 h starvation. The potential contribution of branched-chain complex to estimates of PDH-complex activity in rat liver mitochondria has been defined.
 ...
PMID:Modulation of pyruvate dehydrogenase kinase activity in cultured hepatocytes by glucagon and n-octanoate. 370 45

Pyruvate inhibited pyruvate dehydrogenase kinase activity in mitochondria from adipose tissue, heart, brain and kidney of fed rats. Starvation for 24 h led to increased kinase activity in mitochondria from adipose tissue and heart but not from brain or kidney and to reduction of pyruvate inhibition of the enzyme from adipose tissue, heart and brain. Insulin injection into starved animals rapidly restored pyruvate inhibition without alteration of kinase activity in adipose tissue and heart mitochondria. Induction of streptozotocin diabetes resulted in loss of pyruvate inhibition of the kinase in heart mitochondria at 48 h but not at 24 h whereas a significant increase of kinase activity was seen at 24 h. It is concluded that the mechanisms which control fluctuations of pyruvate sensitivity of the kinase are different from the mechanisms which control fluctuations of the uninhibited kinase activity.
 ...
PMID:Pyruvate inhibition of pyruvate dehydrogenase kinase is a physiological variable. 388 4

The regulatory effects of alpha-ketoisovalerate on purified bovine heart pyruvate dehydrogenase complex and endogenous pyruvate dehydrogenase kinase were investigated. Incubation of pyruvate dehydrogenase complex with 0.125 to 10 mM alpha-ketoisovalerate caused an initial lag in enzymatic activity, followed by a more linear but inhibited rate of NADH production. Incubation with 0.0125 or 0.05 mM alpha-ketoisovalerate caused pyruvate dehydrogenase inhibition, but did not cause the initial lag in pyruvate dehydrogenase activity. Gel electrophoresis and fluorography demonstrated the incorporation of acyl groups from alpha-keto[2-14C]isovalerate into the dihydrolipoyl transacetylase component of the enzyme complex. Acylation was prevented by pyruvate and by arsenite plus NADH. Endogenous pyruvate dehydrogenase kinase activity was stimulated specifically by K+, in contrast to previous reports, and kinase stimulation by K+ correlated with pyruvate dehydrogenase inactivation. Maximum kinase activity in the presence of K+ was inhibited 62% by 0.1 mM thiamin pyrophosphate, but was inhibited only 27% in the presence of 0.1 mM thiamin pyrophosphate and 0.1 mM alpha-ketoisovalerate. Pyruvate did not affect kinase inhibition by thiamin pyrophosphate at either 0.05 or 2 mM. The present study demonstrates that alpha-ketoisovalerate acylates heart pyruvate dehydrogenase complex and suggests that acylation prevents thiamin pyrophosphate-mediated kinase inhibition.
 ...
PMID:Effects of alpha-ketoisovalerate on bovine heart pyruvate dehydrogenase complex and pyruvate dehydrogenase kinase. 394 Oct 88

The activity of the pyruvate dehydrogenase kinase, which phosphorylates and thereby inactivates the pyruvate dehydrogenase complex, was stimulated by malonyl-CoA. Treatment with [2-14C]malonyl-CoA resulted in acylation of sites in the complex. Both acylation and activation of kinase activity increased in a time-dependent manner with a parallel increase in those activities when the malonyl-CoA:CoA ratio was varied. Protein-bound acyl groups were labilized by performic acid treatment indicating their attachment to protein at thiol residues; however, the product released was volatile, which is not characteristic of malonic acid. While malonyl-CoA was initially free of acetyl-CoA, stimulation of kinase activity and acylation of sites in the complex by malonyl-CoA were shown to be contingent upon enzyme-catalyzed decarboxylation. Decarboxylation appeared to be catalyzed by a trace contaminant present in highly purified preparations of both the pyruvate and 2-oxoglutarate dehydrogenase complexes. Under conditions in which both free CoA was removed (by conversion to succinyl-CoA) and then, after various periods, free acetyl-CoA was removed (by enzymic conversion to acetyl phosphate), both acetylation of sites in the complex and activation of kinase activity increased in a time-dependent manner. Concomitantly there was a decrease in the concentration dependence for activation of the kinase by malonyl-CoA. Our results strongly support the conclusion that activation of kinase activity is associated with acylation of sites in the complex, and that, in the case of malonyl-CoA, those processes depend on enzyme-catalyzed decarboxylation.
 ...
PMID:Mechanism of activation of bovine kidney pyruvate dehydrogenase a kinase by malonyl-CoA and enzyme-catalyzed decarboxylation of malonyl-CoA. 401 76

Activity of the pyruvate dehydrogenase complex determines the rate of glucose oxidation in animals including man. The complex is regulated by reversible phosphorylation, phosphorylation resulting in inactivation. Activity is therefore dependent upon the activities of pyruvate dehydrogenase kinase and phosphatase. Activity of the complex is reduced in diabetes and starvation as a result of insulin deficiency. The mechanism involves activation of pyruvate dehydrogenase kinase by short-term effects of products of fatty acid oxidation and by longer term effects involving specific protein synthesis; in hepatocytes the signals may include lipid fuels and glucagon. Activity of the branched chain ketoacid dehydrogenase complex determines the rate of degradation of branched chain aminoacids which is adjusted according to dietary supply. The complex is regulated by reversible phosphorylation, phosphorylation being inactivating. In liver and kidney, but not in muscles a protein activator (free E1 component) may reactivate phosphorylated complex without dephosphorylation and facilitate hepatic oxidation of branched chain ketoacids. Metabolic adjustments induced by diet and diabetes include loss of activator protein, loss of total complex activity in liver but not muscles, and enhanced inactivation by phosphorylation in liver.
 ...
PMID:alpha-Ketoacid dehydrogenase complexes and respiratory fuel utilisation in diabetes. 405 46

The oxidative decarboxylation and subsequent production of glucose from alpha-ketobutyrate were studied using perfused livers from fasted rats. The production of 14CO2 from alpha-keto-[1-14C]butyrate increased monotonically while the production of glucose from alpha-ketobutyrate was biphasic as the perfusate concentration of alpha-ketobutyrate was increased. The biphasic gluconeogenic response using alpha-ketobutyrate as the gluconeogenic precursor was similar to that observed with propionate. The decarboxylation of alpha-ketobutyrate was found to be exquisitely sensitive to the effects of the monocarboxylate transport inhibitor, alpha-cyanocinnamate. Infusion of beta-hydroxybutyrate caused a substantial inhibition of alpha-ketobutyrate decarboxylation while dichloroacetate, a pyruvate dehydrogenase kinase inhibitor, did not stimulate the metabolism of alpha-ketobutyrate but was inhibitory. The effects of alpha-ketobutyrate infusion on pyruvate decarboxylation were tested and it was found that at low perfusate pyruvate concentrations (ca. 0.25 mM) increasing alpha-ketobutyrate led to increasing inhibition of pyruvate decarboxylation, while at high perfusate pyruvate concentrations (ca. 2.5 mM) an initial inhibition was apparent which did not increase substantially with increasing alpha-ketobutyrate concentrations. The results obtained indicate that the regulation of alpha-ketobutyrate metabolism by oxidative decarboxylation differs significantly from that of pyruvate. In addition, while the rate of gluconeogenesis using alpha-ketobutyrate as a precursor was remarkably similar to that using propionate as a gluconeogenic precursor, the effects of alpha-ketobutyrate on the oxidative decarboxylation of pyruvate were qualitatively different from the effects of propionate on pyruvate metabolism.
 ...
PMID:alpha-Ketobutyrate metabolism in perfused rat liver: regulation of alpha-ketobutyrate decarboxylation and effects of alpha-ketobutyrate on pyruvate dehydrogenase. 406 89

This paper reports the discovery that the activity of the multienzyme pyruvate dehydrogenase complex from beef kidney mitochondria is regulated by a phosphorylation-dephosphorylation reaction sequence. The site of this regulation is the pyruvate dehydrogenase component of the complex. Phosphorylation and concomitant inactivation of pyruvate dehydrogenase are catalyzed by an ATP-specific kinase (i.e., a pyruvate dehydrogenase kinase), and dephosphorylation and concomitant reactivation are catalyzed by a phosphatase (i.e., a pyruvate dehydrogenase phosphatase). The kinase and the phosphatase appear to be regulatory subunits of the pyruvate dehydrogenase complex.
 ...
PMID:Alpha-keto acid dehydrogenase complexes. X. Regulation of the activity of the pyruvate dehydrogenase complex from beef kidney mitochondria by phosphorylation and dephosphorylation. 430 45


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>