Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.1 (protein kinase)
 81,284 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Lithium ion, like insulin, activated adipocyte glycogen synthase with or without glucose in the medium. However, the effect of lithium ion was much greater than that of insulin under both conditions. The lithium-activated glycogen synthase was stable to both Sephadex chromatography and ethanol precipitation of the enzyme, indicating that the effect of lithium ion on glycogen synthase was through covalent modification of the enzyme. Glycogen synthase was significantly activated by lithium ion under conditions where concentrations of cellular ATP were unaffected. The effect of lithium ion on glycogen synthase was rapid and observed at concentrations as low as 1 to 3 mM, reaching a maximum at the concentration of 40 mM. It was thus the most sensitive of all the effects studied (see previous paper). Insulin further stimulated glycogen synthase at low concentrations but not at maximal concentration of lithium ion. Lithium-activated glycogen synthase was inhibited by both epinephrine and dibutyryl cyclic AMP, but was not affected by the removal of extracellular Ca++. Interestingly, lithium ion had no detectable effect on basal pyruvate dehydrogenase as well as on epinephrine-stimulated phosphorylase. The failure of lithium ion to thus mimic insulin actions on pyruvate dehydrogenase and on phosphorylase suggests that the action of lithium ion on glycogen synthase is quite specific and may be mediated by stimulating a phosphatase or by inhibiting a protein kinase acting specifically on glycogen synthase.
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PMID:'Insulin-like' effects of lithium ion on isolated rat adipocytes. II. Specific activation of glycogen synthase. 641 71

Synaptic membranes from rat brain contain several calcium-requiring protein kinase (PK) activities with different substrate specificities: (a) an activity (CaH-PK) effective at high concentrations of Ca2+ ion in the absence of Mg2+ (active on class F substrates); (b) a (Ca + Mg)-PK activity that is mediated by Ca2+ ion in the presence of Mg2+ (active on class B substrates); (c) (Ca-CaM)-PK activities that exhibit simultaneous requirements for both Ca2+ ion and CaM (for class C and D substrates). Also described are three activities (d-f) that do not require Ca2+ ion: (d) a Mg-PK activity in which the presence of Ca2+ causes the inhibition of phosphorylation (active on class A substrates); (e) an activity affecting a diverse group of substrates (class E substrates), the phosphorylation of which occurs in the presence of Mg2+ ion alone (Mg-PK activity) and is unaffected by the addition of Ca2+ ion and CaM, the substrates of which show different responses to several types of inhibitors; and, finally, (f) the previously well characterized cAMP-dependent PK activities. Several of the substrates of these kinases have been identified in a fairly unambiguous manner: among them are P43 (class A), as the alpha subunit of pyruvate dehydrogenase; P54 (class B), as the presynaptic protein B50; and the doublet P75-P80, as proteins IA and IB of Ueda and Greengard. The most interesting activity is that requiring both Ca2+ and CaM. The half-maximal stimulation (K0.5) for Ca2+ in the presence of CaM was found to be 1.0 microM Ca2+F in untreated membranes. There is little change in this value on prior EGTA extraction of the membranes, which removes the bulk of its Ca2+ and reduces its residual CaM by greater than or equal to 50%. The apparent K0.5 for CaM in the presence of excess Ca2+ ion was found to equal 0.4 microgram per reaction mixture (8 micrograms/ml) or 1.35 micrograms per reaction mixture (27 micrograms/ml), for the untreated and EGTA-treated membranes, respectively.
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PMID:Calcium-stimulated protein phosphorylation in synaptic membranes. 655 Jun 23

A calmodulin-activated protein kinase has been identified in bovine anterior pituitary membranes. This enzyme phosphorylated one endogenous substrate of subunit molecular weight 53,000 in the membranes. Phosphorylation of this protein was rapid, was half-maximal at 2.5 microM calcium in the presence of saturating concentrations of calmodulin (CaM), and was inhibited by trifluoperazine and thioridazine. A second protein was phosphorylated by an endogenous protein kinase in anterior pituitary membranes. Phosphorylation of this 42,000 Mr protein was reduced by calcium, was independent of exogenously added CaM, and was increased by trifluoperazine or thioridazine. The 42,000 Mr protein may be the alpha-subunit of pyruvate dehydrogenase. Calcium-dependent protein phosphorylation was also observed in intact cells; the largest increases were seen in proteins of Mr 42,000, 21,000 and 17,000.
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PMID:Calcium-dependent protein phosphorylation in bovine anterior pituitary membranes and intact cells. 671 13

The phosphorylation of the alpha-subunit of mitochondrial pyruvate dehydrogenase may be involved in the development of long-term potentiation in the hippocampus. Study of this hypothesis is hampered by variability in the incorporation of 32P into pyruvate dehydrogenase of hippocampal subcellular preparations, in vitro. 32P from [gamma-32P]ATP was incorporated into pyruvate dehydrogenase present in mitochondria and in a membrane-enriched synaptic particulate fraction from hippocampus. However, the presence of the synaptic fraction decreased isotopic labeling of the mitochondrial protein. This effect was not due to inhibition of the protein kinase or activation of a protein phosphatase, but the rate of ATP hydrolysis was found to be higher in the synaptic fraction than in the mitochondria (34 nmol/mg protein/min vs 14 nmol/mg protein/min). These data raise a variety of questions about the interpretation of the in vitro phosphorylation assay. It is concluded that variability in in vitro labeling can be minimized if the effect of ATP hydrolysis is diminished by use of a higher concentration of ATP. In addition, these data indicate that quantitative comparisons of the in vitro phosphorylation of diverse subcellular preparations must take into account differential rates of ATP hydrolysis.
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PMID:Factors affecting the phosphorylation of a 41,000 dalton protein of hippocampal subcellular fractions. 673 99

Purified rat adipocyte plasma membranes incubated with insulin produce a soluble chemical mediator that stimulates pyruvate dehydrogenase when added to isolated mitochondria, or glycogen synthase when added to cell homogenates. The mediator appears to be a peptide and has been characterized by conventional chromatographic methods including gel filtration, ion exchange, and hydroxylapatite chromatography. These studies reveal that an insulin-dependent bioactive component, which is small and negatively charged at pH 7.4, can be eluted from Dowex 1 x 4 by 0.3-0.4 N NaCl or from hydroxylapatite by 0.05-0.15 M potassium phosphate. The mediator has also been partially purified by high-pressure liquid chromatography. A molecular sieving matrix produces a peak of insulin-dependent bioactivity that corresponds to a peak of absorbance at 210 nm (apparent Mr of 2000) and is increased by insulin. Reversed-phase high-pressure liquid chromatography indicates that the insulin-dependent bioactivity is of a hydrophilic nature. Previous studies showed that release of mediator from plasma membranes in response to insulin was blocked by inhibitors of serine proteases and esters of arginine. In addition, bioactivity of the insulin-treated plasma membrane supernatant could be destroyed by protease treatment. In the present experiments, pretreatment of intact adipocytes with serine protease inhibitors blocked the action of subsequently added insulin on several intracellular enzyme systems. These observations have been summarized in the following working model of one mode of insulin action. The binding of insulin to its receptor activates a membrane protease or alters an endogenous membrane substrate, resulting in the increased release into the cell of a small peptide fragment by proteolytic cleavage. The released peptide is proposed to modulate several cellular enzymes such as pyruvate dehydrogenase and glycogen synthase by interacting with phosphoprotein phosphatase or protein kinase activities, or both.
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PMID:Production by plasma membranes of a chemical mediator of insulin action. 681 28

Our understanding of the molecular basis of insulin actin remains incomplete, but important new insights have been achieved recently. All available evidence to date indicates that intracellular signalling by the hormone results from its initial interaction with specific cell surface receptors. Insulin receptors from all tissues studied to date appear to be minimally composed of two Mr 125,000 subunits denoted as alpha and two Mr 90,000 subunits denoted as beta. The beta subunit is extremely sensitive to proteolytic cleavage near the center of its amino acid chain. The four subunits are linked together by disulfide bonds to give a symmetrical configuration with a stoichiometry of (alpha-s-s-beta)-s-s-(alpha-s-s-beta). This structure is remarkably similar to the general subunit composition of immunoglobulin G molecules and provides a structural basis for the postulate that this minimum insulin receptor structure may be divalent for binding hormone. A second area of recent progress involves the successful generation of a soluble factor or factors by insulin that are capable of modulating the activity of insulin-sensitive enzymes such as pyruvate dehydrogenase, glycogen synthase and cyclic adenosine monophosphate (cyclic AMP)-dependent protein kinase in cell-free systems. Indirect evidence indicates that the putative mediator or mediators of insulin action exhibits properties expected of a low molecular weight peptide, including destruction by proteases. The data available are consistent with the hypothesis that insulin-receptor interaction leads to the activation of a membrane protease that catalyzes the release of a peptide mediator or mediators of insulin action.
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PMID:Insulin action. 700 87

Rat liver mitoplasts (inner mitochondrial membrane and matrix) contain protein kinase activity. This activity increases twofold on addition of Triton X-100. The activity observed in absence of Triton X-100 is probably exposed on the outer surface of mitoplasts, since it is sensitive to trypsin treatment. Most of the remaining protein kinase is bound to the membrane fraction, presumably on the inside of (or else hidden in) the inner mitochondrial membrane. Only a small part of the kinase activity is found in the mitochondrial matrix. A phosphoprotein band, partly resolved into a doublet, was observed on electrophoresis in SDS-polyacrylamide gels after endogeneous phosphorylation of mitoplasts, inner mitochondrial membrane or matrix. When isolated fractions are phosphorylated approximately 75% of the phosphoprotein is found in the matrix, and the remainder in the inner membrane. The phosphorylation of the doublet is inhibited by inhibitors to pyruvate dehydrogenase kinase, suggesting that it represents the phosphorylated subunit of pyruvate dehydrogenase.
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PMID:Localization of protein kinase activity and phosphoproteins in mitoplasts from rat liver. 733 41

Evidence for a reversible process resulting in stable activated and inactivated states of the mitochondrial branched chain alpha-keto acid dehydrogenase complex in isolated perfused rat heart is presented. The inactivation process is mediated by pyruvate infusion, while activation (up to 18-fold) is facilitated by branched chain alpha-keto acid substrates. The low activity state of the branched chain complex characteristic of freshly excised rat hearts could be maintained by infusion of either pyruvate or glucose. Activation of the complex in the perfused rat heart was achieved slowly by substrate-free perfusion, while rapid activation was accomplished by infusion of branched chain alpha-keto acids. The fully activated enzyme complex resulting from branched chain alpha-keto acid infusion subsequently could be inactivated maximally by infusion of pyruvate alone or intermediate degrees of inactivation could be produced by certain ratios of co-infused pyruvate and branched chain alpha-keto acid. alpha-Ketoisocaproate was an order of magnitude more effective than alpha-keto isovalerate either in preventing inactivation or in stimulating the opposing activation process when co-infused with pyruvate. The mitochondrial pyruvate transport inhibitor, alpha-cyanocinnamate, effectively prevented inactivation of the complex by infused pyruvate. Differential changes in the activation states of the branched chain alpha-keto acid dehydrogenase and pyruvate dehydrogenase complexes were evident when the two complexes were compared in apparently similar flux-inhibited (via octanoate infusion) and flux-stimulated (via dichloroacetate infusion) metabolic conditions. The differential effect of pyruvate concentration on the activity states of the two complexes was also well-defined. The results of the present study suggest distinct systems for the regulation of the activity of the two multienzyme complexes of interest. While our results argue neither for nor against an inactivation of the branched chain alpha-keto acid dehydrogenase complex by a protein kinase, the regulatory properties of such an intramitochondrial protein kinase may not be similar to the pyruvate dehydrogenase kinase. The mechanistic nature of the suggested novel regulatory system concerned with the pyruvate-mediated inactivation of the branched chain alpha-keto acid activation cannot be inferred at the present time.
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PMID:Studies on the activation and inactivation of the branched chain alpha-keto acid dehydrogenase in the perfused rat heart. 743 Jan 1

We have investigated the signalling pathways involved in the stimulation of glycogen and fatty acid synthesis by insulin in rat fat cells using wortmannin, an inhibitor of phosphatidylinositol 3-kinase, and rapamycin, which blocks activation of p70 ribosomal S6 protein kinase (p70S6K). Insulin produced a decrease in the activity of glycogen synthase kinase-3 which is likely to be important in the observed stimulation of glycogen synthase. Both of these actions were found to be sensitive to inhibition by wortmannin. Activation of three processes is involved in the stimulation of fatty acid synthesis from glucose by insulin, namely glucose uptake, acetyl-CoA carboxylase and pyruvate dehydrogenase. Whereas wortmannin largely abolished the effects of insulin on glucose utilization and acetyl-CoA carboxylase activity, it was without effect on the stimulation of pyruvate dehydrogenase. Although epidermal growth factor stimulated mitogen-activated protein kinase to a greater extent than insulin, it was unable to mimic the effect of insulin on glycogen synthase, glycogen synthase kinase-3, glucose utilization, acetyl-CoA carboxylase or pyruvate dehydrogenase. Rapamycin also failed to have any appreciable effect on stimulation of these parameters by insulin, although it did block the effect of insulin on p70S6K. We conclude that the activity of phosphatidylinositol 3-kinase is required for the effects of insulin on glycogen synthesis, glucose uptake and acetyl-Co-AN carboxylase, but is not involved in signalling to pyruvate dehydrogenase. Activation of mitogen-activated protein kinase or p70S6K, however, does not appear to be sufficient to bring about the stimulation of fatty acid or glycogen synthesis. Altogether is seems likely that at least four distinct signalling pathways are involved in the effects of insulin on rat fat cells.
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PMID:Multiple signalling pathways involved in the stimulation of fatty acid and glycogen synthesis by insulin in rat epididymal fat cells. 748 1

Two heat-stable protein inhibitors of protein phosphatase 2A (PP2A), tentatively designated I1PP2A and I2PP2A, have been purified to apparent homogeneity from extracts of bovine kidney. The purified preparations of I1PP2A exhibited an apparent M(r) approximately 30,000 and 250,000 as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and gel permeation chromatography on Sephacryl S-300, respectively. In contrast, the purified preparations of I2PP2A exhibited an apparent M(r) approximately 20,000 and 80,000 as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and gel permeation chromatography on Sephacryl S-200, respectively. The purified preparations of I1PP2A and I2PP2A inhibited PP2A with 32P-labeled myelin basic protein, 32P-labeled histone H1, 32P-labeled pyruvate dehydrogenase complex, 32P-labeled phosphorylase, and protamine kinase as substrates. By contrast, I1PP2A and I2PP2A exhibited little effect, if any, on the activity of PP2A with 32P-labeled casein, and did not prevent the autodephosphorylation of PP2A in incubations with the autophosphorylation-activated protein kinase [Guo, H., & Damuni, Z. (1993) Proc. Natl. Acad. Sci. U.S.A. 90, 2500-2504]. The purified preparations of I1PP2A and I2PP2A had little effect, if any, on the activities of protein phosphatase 1, protein phosphatase 2B, protein phosphatase 2C, and pyruvate dehydrogenase phosphatase. With 32P-labeled MBP as a substrate, kinetic analysis according to Henderson showed that I1PP2A and I2PP2A were noncompetitive and displayed a Ki of about 30 and 25 nM, respectively. Following cleavage with Staphylococcus aureus V8 protease, I1PP2A and I2PP2A displayed distinct peptide patterns, indicating that these inhibitor proteins are the products of distinct genes. The N-terminal amino acid sequences of the purified preparations indicate that I1PP2A and I2PP2A are novel proteins.
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PMID:Purification and characterization of two potent heat-stable protein inhibitors of protein phosphatase 2A from bovine kidney. 753 97


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