Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: 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)

Pyruvate kinase (ATP:pyruvate 2-O-phosphotransferase, EC 2.7.1.40) type L was partly purified from rat kidney. During the last two purification steps, the incorporation of [32P]phosphate into protein on incubation with [32P]ATP and cyclic 3',5'-AMP-dependent protein kinase was found to parallel the pyruvate kinase activity. After phosphorylation of the enzyme, a major radioactive band with a molecular weight of 57 000 was found on polyacrylamide gel electrophoresis [32P]Phosphorylserine was isolated from the kidney pyruvate kinase. Immunological identity was found between the liver and kidney pyruvate kinases type L. By autoradiography of high-voltage electropherograms after partial acid hydrolysis of the phosphorylated rat liver and kidney pyruvate kinases type L, identical results were obtained. The affinity for phosphoenolpyruvate was found to be decreased by phosphorylation of the enzyme with a change in the apparent Km from 0.15 mM to 0.35 mM. After incubation of the phosphorylated kidney pyruvate kinase with phosphatase the phosphoenolpyruvate saturation curve was found to be identical to that for the unphosphorylated enzyme. Thus, the activity of the rat kidney pyruvate kinase type L is with all probability regulated by a reversible phosphorylation-dephosphorylation reaction, thereby indicating that hormonal regulation of gluconeogenesis via cyclic AMP may be of importance in the renal cortex.
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PMID:Phosphorylation of rat kidney pyruvate kinase type L by cyclic 3',5'-AMP-dependent protein kinase. 20 41

Inhibitor-1 is a protein which inhibits phosphorylase phosphatase only when it has been phosphorylated by cyclic-AMP-dependent protein kinase [Huang, F. L. and Glinsmann, W. H. (1976) Eur. J. Biochem. 70, 419--426]. Inhibitor-1 was purified by a heat treatment at 90 degrees C, precipitation with ammonium sulphate, chromatography on DEAE-cellulose, gel filtration on Sephadex G-100, and finally rechromatography of the phosphorylated protein on DEAE-cellulose, The protein was purified 4000-fold and 1.5 mg per 1000 g muscle was obtained in seven days corresponding to an overall yield of 15-20%. The purified protein was in a state approaching homogeneity as judged by the criteria of polyacrylamide-gel electrophoresis and ultracentrifugal analysis. The concentration of inhibitor-1 in vivo was calculated to be 1.5 micron, which is at least as high as the concentration of phosphorylase phosphatase. The amino acid composition of inhibitor-1 showed several unusual features. Glutamic acid and proline accounted for nearly one third of the residues, tyrosine, tryptophan and cysteine were absent, and the content of aromatic amino acids was very low. The molecular weight measured by sedimentation equilibrium centrifugation was 19200 and by amino acid analysis was 20800. These values were lower than the mol. wt 26000 determined empirically by gel electrophoresis in the presence of sodium dodecyl sulphate, and much lower than the apparent molecular weight of 60000 estimated by gel filtration on Sephadex G-100. The gel filtration behaviour, stability to heating at 100 degrees C and amino acid composition suggest that inhibitor-1 may possess little ordered structure. The phosphorylated from of inhibitor-1 contained close to one molecule of covalently bound phosphate per mole of protein, which is consistent with the previous finding of a unique decapeptide sequence at the site of phosphorylation, Ile-Arg-Arg-Arg-Arg-Pro-Thr(P)-Pro-Ala-Thr- [Cohen, P., Rylatt, D. B. and Nimmo, G. A. (1977) FEBS Lett. 76, 182-186].the phosphorylated form of inhibitor-1 inhibited phosphorylase phosphatase activity (0.02U) by 50% at a concentration of only 7.0 nM in the standard assay, but the phosphorylated decapeptide was 1000-2000 times less effective as an inhibitor.
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PMID:The regulation of glycogen metabolism. Purification and characterisation of protein phosphatase inhibitor-1 from rabbit skeletal muscle. 20 44

Sterol ester hydrolase (cholesterol esterase, E.C. 3.1.1.13) of bovine adrenal cortex has been extensively purified by ammonium sulfate fractionation, acid precipitation, hydroxylapatite chromatography, and Sephadex G-200 chromatography. During the purification sequence, the hydrolase activity was purified free of endogenous protein kinase. With this purified preparation, activation by cyclic AMP and ATP-Mg2+ did not occur unless exogenous protein kinase was included in the activating system. Using [gamma-32P]ATP, the transfer of the terminal phosphate to the enzyme protein was demonstrated by three separate experimental approaches. With pooled fractions from Sephadex G-200 chromatography, significant binding of 32P by the enzyme protein was observed only in the presence of exogenous protein kinase. Time course studies disclosed a close concurrence between the extent of activation of the purified enzyme by cyclic AMP-dependent protein kinase and the level of 32P transfer from [gamma-32P]ATP to the enzyme protein. Finally, assays carried out during Sephadex G-200 chromatography showed a correspondence in the peaks for activated sterol ester hydrolase and for 32P binding by protein. The data confirm that activation of adrenal sterol ester hydrolase by cyclic AMP and ATP-Mg2+ involves protein kinase-catalyzed phosphorylation of the enzyme protein.
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PMID:Protein kinase-mediated phosphorylation of a purified sterol ester hydrolase from bovine adrenal cortex. 20 11

A chromatin associated protein kinase was used to add 3 moles of phosphate to seryl side chains of 1 mole of histone H1. The DNA binding properties of this in vitro phosphorylated H1 were compared with those of unmodified H1. Considerably more radioactive superhelical DNA was retained on nitrocellulose filters at 20mM-40mM NaCl by phosphorylated H1 than by unmodified H1. However, zone velocity sedimentation analysis of histone-DNA complexes indicated that similar amounts of phosphorylated and unmodified H1 are bound to DNA. It is therefore concluded that phosphorylated H1 binds distributively to many or all DNA molecules available (depending on the histone/DNA ratio) while unmodified H1 binds cooperatively to a fraction of the DNA molecules in the reaction mixture.
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PMID:Binding of phosphorylated histone H1 to DNA. 20 6

The catalytic subunit of rabbit skeletal muscle cyclic AMP-dependent protein kinase I can catalyze self-phosphorylation. The autophosphorylation reaction uses ATP as the phosphoryl donor, requires Mg2+, and is inhibited by polyarginine. Prior treatment of the catalytic subunit with Escherichia coli alkaline phosphatase in the presence of bovine serum albumin greatly enhances the autophosphorylation of the subunit. The protein-bound phosphate is stable in acid but labile in base. Incubation of the 32P-labeled phosphoenzyme with histones led neither to the phosphorylation of histones nor to a loss of radioactivity from the phosphoenzyme. The results suggest that the phosphoenzyme does not represent an intermediate of the phosphotransferase reaction.
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PMID:Autophosphorylation of rabbit skeletal muscle cyclic AMP-dependent protein kinase I catalytic subunit. 21 18

Turkey gizzard smooth muscle light chain kinase was purified by affinity chromatography on calcium dependent regulator weight of 125,000 +/- 5,000 in sodium dodecyl sulfate-polyacrylamide gel electrophoresis. When myosin light chain kinase is incubated with the catalytic subunit of cyclic AMP-dependent protein kinase, 1 mol of phosphate is incorporated per mol of myosin kinase. Brief tryptic digestion of the 32P-labeled myosin kinase liberates a single radioactive peptide with a molecular weight of approximately 22,000. Phosphorylation of myosin kinase results in a 2-fold decrease in the rate at which the enzyme phosphorylates the 20,000-dalton light chain of smooth muscle myosin. These results suggest that cyclic AMP has a direct effect on actin-myosin interaction in smooth muscle.
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PMID:Phosphorylation of smooth muscle myosin light chain kinase by the catalytic subunit of adenosine 3': 5'-monophosphate-dependent protein kinase. 21 32

Based on the chemotactic activity of approximately 50 different adenosine 3',5'-cyclic-monophosphate (cyclic AMP) derivatives with substitutions at the phosphate, ribose and adenine moieties, a model for the cyclic AMP-chemoreceptor interaction in Dictyostelium discoideum is proposed. In this model the cyclic AMP molecule is bound to the receptor by three hydrogen bonds at, respectively, the 3'-oxygen of the ribose and the 6-amino and the 7-nitrogen of the base, and possibly by one ionic interaction of the negatively charged phosphate group. The conformation of the adenine moiety is in the anti range and binds additionally to the receptor by hydrophobic interactions betueen its pi-electron system and a corresponding acceptor at the active site. Although this receptor clearly differs from that involved in protein kinase activation in higher organisms, the existence of striking similarities suggests a basic mechanism for cyclic AMP interaction conserved during evolution.
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PMID:A model for cyclic AMP-chemoreceptor interaction in Dictyostelium discoideum. 21 55

Avian sarcoma virus (ASV) induces sarcomas in animals and transforms fibroblasts to a neoplastic state in cell culture. A single viral gene (src) is responsible for both the induction and maintenance of neoplastic transformation. Recent work has identified a protein with a molecular weight of 60,000 daltons that is apparently encoded in src and may be the effector molecule for the gene (Brugge and Erikson, 1977; Purchio et al, 1978). The putative product of src can be immunoprecipitated by antisera obtained from rabbits bearing tumors induced by ASV. We have used this approach to isolate the protein to characterize further its genetic origins and possible function. Our rabbit tumor antisera precipitated a protein with a molecular weight of 60,000 daltons; according to serological, biochemical and genetic criteria, this protein is encoded in src. We found that this protein is phosphorylated and therefore denoted it pp60. Phosphorylation of pp60 could be accomplished in vitro with extracts of ASV-infected cells. A temperature-sensitive conditional mutation in src had no demonstrable effect on either the production or stability of pp60 in the infected cell, but phosphorylation of the protein was temperature-sensitive. Since the mutant src is not expressed at the restrictive temperature, our findings raise the possibility that phosphorylation of pp60 is required for its function as the putative effector of src. Immunoprecipitates prepared with extracts of ASV-infected cells and the rabbit tumor antisera contained a protein kinase activity that catalyzed phosphorylation of the heavy chains of immunoglobulin molecules, using either ATP or GTP as phosphate donor. The kinase activity immunoprecipitated in parallel with pp60 was obtained only from cells that contained a functioning product of src and could not be precipitated with antisera directed against structural proteins of ASV. A temperature-sensitive conditional mutation in src caused the kinase activity to be thermally inactivated in vitro far more rapidly than the activity from cells infected with wild-type virus. We conclude that both the protein kinase and pp60 are encoded in src, and that the enzymatic activity may be an intrinsic property of pp60. Phosphorylation of pp60 in cellular extracts was inhibited by calcium ion, whereas the immunoprecipitable kinase activity was not, suggesting that the kinase responsible for pp60 phosphorylation may be distinct from that encoded in src. Collett and Erikson (1978) have also identified a protein kinase activity associated with pp60. These findings raise the possibility that phosphorylation of specific cellular targets might account for transformation of the host cell by src.
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PMID:Evidence that the transforming gene of avian sarcoma virus encodes a protein kinase associated with a phosphoprotein. 21 42

Chinese hamster ovary cells exhibit several characteristic morphological and physiological responses upon treatment with agents which increase the intracellular level of adenosine 3':5'-phosphate (cyclic AMP). To better understand the mechanism of these cyclic AMP-mediated responses, we separated two cyclic AMP-dependent protein kinases (ATP:protein phosphotransferase, EC 2.7.1.37) (protein kinase I and protein kinase II) from the cytosol of Chinese hamster ovary cells by DEAE-cellulose chromatography and studied their properties. Protein kinase I is eluted at a lower salt concentration than protein kinase II and is stimulable to 10 times its basal catalytic activity, while protein kinase II is stimulable only 2-fold. Both kinases are completely dissociated by cyclic AMP and inhibited by specific cyclic AMP-dependent protein kinase inhibitor. They have similar Km values for magnesium (approximately 1 mM), cyclic AMP (approximately 60 nM), and ATP (approximately 0.1 mM), and the dissociation constant (Kdis) for cyclic AMP (approximately 13 nM) is the same for both enzymes. However, they appear to have different substrate preferences and cyclic AMP-binding properties in that cyclic AMP bound to protein kinase II exchanges readily with free cyclic AMP, while that bound to protein kinase I is not exchangeable. The native enzymes have different sedimentation coefficients (6.4 S for protein kinase I and 4.8 S for protein kinase II), whereas those of the activated enzymes are the same (2.9--3.0 S). It appears that the two cyclic AMP-dependent protein kinases which differ from each other in their regulatory subunits may play different roles in the mediation of cyclic AMP action in Chinese hamster ovary cells.
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PMID:Characterization of two adenosine 3':5'-phosphate-dependent protein kinase species from Chinese hamster ovary cells. 21 11

32P phosphorylation of plasma membranes from human blood platelets, under conditions that closely resemble physiological ones (endogeneous phosphate donors and intact platelets in homologous plasma), result in the incorporation of the label mainly in a membrane glycoprotein of apparently high molecular weight (greater than 400 000). Dibutyryl cyclic AMP, an inhibitor of platelet aggregation, specifically increases the degree of phosphorylation of this glycoprotein. Moreover, it has been found that prostaglandin E1 one of the most potent inhibitors of platelet aggregation which also increases phosphorylation of the same glycoprotein, is significantly more effective than cyclic AMP. Cyclic GMP does not have any apparent effect on platelet aggregation. However, incubation of platelet-rich plasma with both cyclic GMP and cyclic AMP results in a partial recovery of the platelet responsiveness towards ADP-induced aggregation. Coincidently, the degree of phosphorylation of the high molecular weight glycoprotein under these conditions, although still higher than in controls (no nucleotides added), is significantly decreased as compared with cyclic AMP-treated cells. Furthermore, cyclic GMP inhibits the cyclic AMP-dependent protein kinase activity in isolated platelet plasma membranes. These results suggest a central role for this membrane phosphoglycoprotein in the triggering of platelet aggregation and, furthermore, suggest that modulation of its degree of phosphorylation may be exerted through some cyclic AMP/cyclic GMP relationship, which in the basal state might be critical for platelet responsiveness.
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PMID:Modulation of platelet responsiveness through selective phosphorylation of plasma membrane proteins. 21 26


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