Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.11 (AMPK)
 12,425 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

During meiotic maturation of Xenopus laevis stage 6 oocytes into unfertilized eggs, 40S ribosomal protein S6 undergoes multiple phosphorylation. Extracts prepared from unfertilized eggs are up to 10-fold more efficient in phosphorylating S6 than those prepared from immature oocytes. When analyzed by DEAE chromatography the S6 kinase activity elutes as a single peak. If extracts from unfertilized eggs are prepared in the absence of beta-glycerol phosphate, a putative phosphatase inhibitor, there is a severe reduction in recovered S6 kinase activity. Under optimal conditions, incubation of unfertilized egg extracts with 40S ribosomes in the presence of ATP leads to the average incorporation of 3.5 mol of phosphate/mol of S6. Prior incubation of these extracts with the cAMP-dependent protein kinase inhibitor does not inhibit S6 phosphorylation indicating that another kinase is responsible. Analysis of the in vitro phosphorylated peptides demonstrates that they migrate to the equivalent position of those observed previously in vivo and in vitro. More strikingly, if each of the increasingly phosphorylated derivatives of S6 is analyzed independently, it is found that the phosphopeptides appear in a specific order.
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PMID:Activation of ribosomal protein S6 phosphorylation during meiotic maturation of Xenopus laevis oocytes: in vitro ordered appearance of S6 phosphopeptides. 370 23

A tyrosine protein kinase activity has been partially purified from calf thymus using the phosphorylation of the tyrosine-containing peptide angiotensin I as an assay. Detergent extracts of calf thymus possessed only low levels of specific peptide phosphorylating activity when assayed at low ionic strength. The inclusion of NaCl at a concentration of 2 M stimulated endogenous tyrosine protein kinase activity, while the activity of other endogenous kinases was inhibited. This sensitivity to NaCl was retained following partial purification of the enzyme. The phosphorylation of other substrates such as casein or the R-R-SRC peptide (Arg-Arg-Leu-Ile-Glu-Asp-Ala-Glu-Tyr-Ala-Ala-Arg-Gly) by the tyrosine protein kinase was less sensitive to NaCl. Phosphorylation of the PK-1 peptide (Leu-Arg-Arg-Ala-Ser-Leu-Gly) by the purified catalytic subunit of cAMP-dependent protein kinase was inhibited by NaCl. The effect of NaCl on angiotensin I phosphorylation could be mimicked by KCl or sodium acetate. The principal effect of NaCl was to increase the Vmax of the enzyme for the phosphorylation of angiotensin I. At low ionic strength, Mn2+ and Co2+ were the preferred required divalent cations. At elevated NaCl concentrations Mg2+ was preferred, with half-maximal activation occurring at 35 mM Mg2+. By conducting peptide phosphorylation assays in the presence of elevated levels of Mg2+ and NaCl, tyrosine protein kinase activity can readily be detected in extracts from cell lines that express low levels of the enzyme.
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PMID:Properties of a tyrosine protein kinase from calf thymus. Response to ionic strength and divalent cations. 387 56

Protein kinase capable of phosphorylating 40S ribosomal protein S6 on serine residues has been detected in chicken embryo fibroblasts. This activity appears to be regulated in direct response to expression of pp60v-src in chicken embryo fibroblasts infected with a temperature-sensitive transformation mutant of Rous sarcoma virus. Partially purified S6 kinase was highly specific for S6 in 40S ribosomal subunits. The S6 kinase was not inhibited by calcium or by the heat-stable inhibitor of cAMP-dependent protein kinase, nor was it activated by phosphatidylserine, diacylglycerol, and calcium. Thus, it is distinct from protein kinase C and cAMP-dependent protein kinase, which are capable of phosphorylating S6 in vitro. The tumor-promoter phorbol 12-myristate 13-acetate also stimulated ribosomal protein S6 kinase activity in serum-starved chicken embryo fibroblasts, whereas phorbol, the inactive analog of phorbol 12-myristate 13-acetate, had no effect. S6 kinase activity stimulated by expression of pp60v-src, by phorbol 12-myristate 13-acetate, or by serum growth factors exhibited similar chromatographic properties upon ion-exchange chromatography. These results suggest that a common protein kinase may be activated by three diverse stimuli all involved in regulating cell proliferation.
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PMID:Regulation of a ribosomal protein S6 kinase activity by the Rous sarcoma virus transforming protein, serum, or phorbol ester. 393 63

Tyrosine-specific protein kinase (ATP:protein phosphotransferase, EC 2.7.1.37) activity was measured in normal human nonadherent peripheral blood lymphocytes using synthetic peptide substrates having sequence homologies with either pp60src or c-myc. A high level of tyrosine-specific protein kinase activity was found associated with the cell particulate fraction (100 000 X g pellet). High-pressure liquid chromatography and phosphoamino acid analysis of the synthetic peptide substrates substantiated the phosphorylation of tyrosine residues by the particulate fraction enzyme. The human enzyme was also capable of phosphorylating a synthetic random polymer of 80% glutamic acid and 20% tyrosine. Enzyme activity was half-maximal with 22 microM Mg X ATP and had apparent Km values for the synthetic peptides from 1.9 to 7.1 mM. The enzyme preferred Mg2+ to Mn2+ for optimal activity and was stimulated 2-5-fold by low levels (0.05%) of some ionic as well as non-ionic detergents including deoxycholate, Nonidet P-40 and Triton X-100. The enzyme activity was not stimulated by N6;O2'-dibutyryl cyclic AMP (100 microM), N6;O2'-dibutyryl cyclic GMP (100 microM), Ca2+ (200 microM), insulin (1 microgram/ml) or homogeneous human T-cell growth factor (3 micrograms/ml) under the conditions used. Alkaline-resistant phosphorylation of particulate proteins in vitro revealed protein bands with Mr 59 000 and 54 000 suggesting that there are endogenous substrates for the human lymphocyte tyrosine protein kinase.
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PMID:High tyrosine-specific protein kinase activity in normal human peripheral blood lymphocytes. 403 88

A protein kinase activity fraction was defined in cytosols and membranes of mammary tissue isolated from rats during pregnancy lactation, and weaning. By partial purification on DEAE-cellulose columns, it was shown that this protein kinase activity is cAMP independent and that its preferential substrate is casein and not histone. This protein kinase activity is inhibited by the bioflavonoid quercetin at doses that do not inhibit cAMP-dependent protein kinase activity. The enzyme requires Mg2+ and is inactive in the presence of 10 mM Ca+2; these properties distinguish this activity from casein kinase activity found in the Golgi fraction and involved in milk protein processing. By following the physiological cycle of mammary gland development during pregnancy, lactation, and weaning, we found a close correlation between proliferation, expressed as the DNA content per gland, and quercetin-inhibited cytosolic protein kinase activity. Moreover, changes in this phosphorylating activity preceded the glandular growth changes. There was a less significant correlation between the growth process and protein kinase activity in the membrane fraction. The cytosolic cAMP-dependent protein kinase activity showed (only partial) correlation with growth only during pregnancy. Cytosolic progesterone receptor levels in mammary tissue were used as an estrogenic marker. Tissue growth correlated with progesterone receptor levels during pregnancy, where estrogens are the predominant hormones affecting tissue proliferation. However, no such correlation was found during lactation and weaning, when PRL is the major hormone affecting mammary gland growth. These results suggest that quercetin-inhibitable protein kinase activity is not merely another estrogenic marker, but represents more general regulatory activity which might be connected to growth processes of breast tissue.
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PMID:Protein kinase activity in the rat mammary gland during pregnancy, lactation, and weaning: a correlation with growth but not with progesterone receptor levels. 609 41

The intrinsic cAMP-dependent protein kinase activity of highly purified cardiac sarcolemmal vesicles was characterized. The sarcolemmal protein kinase was specifically activated by cAMP. Binding of cAMP to the kinase was saturable and occurred exclusively to a protein of Mr = 55,000 intrinsic to the vesicles. This binding of cAMP to the sarcolemmal vesicles caused a selective release of catalytic activity from the membranes, which was capable of phosphorylating several endogenous sarcolemmal substrates as well as one additional substrate, which was also identified in purified vesicles of cardiac sarcoplasmic reticulum. Unmasking experiments conducted with the ionophore alamethicin demonstrated that the protein kinase activity and its endogenous sarcolemmal substrates were localized on the inner, cytoplasmic surfaces of the vesicles, and, furthermore, suggested that at least 75% of the vesicles were right side out. The major protein substrates phosphorylated in the sarcolemmal fraction exhibited apparent molecular weights of 21,000 and 8,000, as analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Heating the membranes in the presence of sodium dodecyl sulfate prior to electrophoresis completely converted the 21,000-dalton substrate into the form of higher mobility, suggesting that the two substrates were, in fact, identical proteins. This was supported by the observation that both substrates exhibited identical pI values of approximately 6.7. Although present in the sarcolemmal fraction, these two substrates were not localized exclusively to sarcolemmal membranes. The same two substrates were present in 3-fold higher content in purified cardiac sarcoplasmic reticulum vesicles. Moreover, although phosphorylation of all other sarcolemmal proteins in right side out vesicles by exogenously added protein kinase was increased 4-fold or greater by alamethicin, phosphorylation of the substrates of Mr = 21,000 and 8,000 was not altered appreciably by the ionophore. The results suggest that these two major substrates identified in the sarcolemmal preparations are not intrinsic sarcolemmal proteins.
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PMID:Characterization of the intrinsic cAMP-dependent protein kinase activity and endogenous substrates in highly purified cardiac sarcolemmal vesicles. 612 8

The Rous sarcoma virus (RSV) transforming gene product has been identified and characterized as a phosphoprotein with a molecular weight of 60,000, denoted pp60src. Partially purified pp60src displays a closely associated phosphotransferase activity with the unusual specificity of phosphorylating tyrosine residues in a variety of proteins. That the enzymatic activity observed is actually encoded by the RSV-transforming gene is indicated by the comparison of the pp60src-protein kinase isolated from cells tranformed by a wild-type RSV or by a RSV temperature-sensitive transformation mutant; these experiments revealed that the latter enzyme had a half-life of 3 min at 41 degrees C, whereas that of the wild-type enzyme was 20 min. Evidence is now beginning to accumulate showing that viral pp60src expresses its protein kinase activity in transformed cells as well as in vitro because at least one cellular protein has been identified as a substrate for this activity of pp60src. Although the protein kinase activity associated with pp60src is itself cyclic AMP (cAMP) independent, the molecule contains at least one serine residue that is directly phosphorylated by the cellular cAMP-dependent protein kinase, thus suggesting that the viral transforming gene product may be regulated indirectly by the level of cAMP. The significance of this latter observation must be regarded from the point of view that the RSV src gene is apparently derived from a normal cellular gene that seemingly expresses in normal uninfected cells a phosphoprotein structurally and functionally closely related to pp60src. This celluar protein, found in all vertebrate species tested, also is a substrate for a cAMP-dependent protein kinase of normal cells, and, therefore, may be evolved to function in a regulatory circuit involving cAMP.
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PMID:Molecular events in cells transformed by Rous Sarcoma virus. 625 1

We investigated the action of thyroid hormone on each protein kinase in rat liver cytosol. Kinases were analyzed by polyacrylamide disc gel electrophoresis and isoelectric focusing in polyacrylamide gel. Polyacrylamide disc gel electrophoresis separated cAMP-dependent protein kinase type I (Rf = 0.35), type II (Rf = 0.44), their catalytic subunit (Rf = 0.26), and cAMP-independent protein kinase (Rf = 0.50). Casein kinase was detected at Rf = 0.37. In addition to the catalytic subunit with Rf = 0.26, another catalytic subunit was found at Rf = 0.44 when the cytosol was preincubated with cAMP. The administration of T3 (20 micrograms/100 g BW for 3 days) to hypothyroid rats increased enzyme activities of type I holoenzyme and casein kinase by 48%. Free catalytic subunit, separated from holoenzyme, had the same level of enzyme activity in both groups, suggesting greater endogenous dissociation of type I holoenzyme in hypothyroid rats. When heat-inactivated rat liver cytosol was used as substrate in the assay of protein kinase activity, the peak enzyme active in phosphorylating the cytosol corresponded to the casein kinase peak. Our data indicate that casein kinase is the main enzyme that mediates phosphorylation of endogenous proteins in rat liver cytosol, and that T3 treatment increases the activity of casein kinase and of type I cAMP-dependent protein kinase.
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PMID:Thyroid hormone increases type I adenosine 3', 5'-monophosphate-dependent protein kinase and casein kinase activities in rat liver cytosol: analysis of protein kinases by polyacrylamide disc gel electrophoresis. 629 94

In vitro, hypothalamic HD1 from rat, could strikingly be inhibited by ATP and cyclic AMP. The enzyme inhibition is partially dependent upon Mg2+ and the circumstances favourable for a cAMP-dependent phosphorylation. An almost complete inhibition could be achieved by incubating the homogenate of the hypothalamus under phosphorylating conditions (ATP, cAMP, Mg2+ and IBMX) in the presence of a cAMP-dependent protein kinase (obtained from bovine thymus). Cyclic nucleotides and ATP alone elicit only moderate inhibitions on the hypothalamic HD activity. Neither ATP, nor cAMP, added alone or in combinations, alter the total brain or the hypothalamic HNMT from guinea-pigs or rats in concentrations up to 10(-3) M. Results suggest that hypothalamic HD is regulated through a cAMP-dependent process, probable a direct phosphorylation, via a cAMP-dependent protein kinase.
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PMID:Regulation of histidine decarboxylase activity in rat hypothalamus in vitro by ATP and cyclic AMP: enzyme inactivation under phosphorylating conditions. 632 55

The effects of substrate and cofactors on the phosphorylation of hepatic phenylalanine hydroxylase by cAMP-dependent protein kinase and on dephosphorylation by phosphoprotein phosphatase have been examined. The presence of the natural cofactor (6R)-tetrahydrobiopterin strongly inhibits the activation observed under phosphorylating conditions; in contrast, this activation is enhanced approximately 20 to 50% by phenylalanine. The phosphorylation of the hydroxylase is strongly inhibited (approximately 80%) by (6R)-tetrahydrobiopterin, while phosphorylation is modestly stimulated by phenylalanine. High concentrations of phenylalanine (1 mM), however, can substantially reverse the inhibition of phosphorylation by (6R)-tetrahydrobiopterin. Neither (6R)-tetrahydrobiopterin nor phenylalanine affect the phosphorylation of a synthetic peptide substrate of cAMP-dependent protein kinase. The inhibition is specific for (6R)-tetrahydrobiopterin; the diastereoisomer (6S)-tetrahydrobiopterin has a much smaller effect, and 6-methyltetrahydropterin and 6,7-dimethyltetrahydropterin have no effect. Both phenylalanine and (6R)-tetrahydrobiopterin inhibit to a small extent the dephosphorylation of phosphorylated phenylalanine hydroxylase catalyzed by phosphoprotein phosphatase. Neither phenylalanine nor (6R)-tetrahydrobiopterin inhibit the dephosphorylation of phosphorylated histones by phosphoprotein phosphatase. These results suggest that the phosphorylation state, and thus the activation state, of phenylalanine hydroxylase in vivo may be modulated, in part, by the availability of substrate.
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PMID:Ligand effects on the phosphorylation state of hepatic phenylalanine hydroxylase. 669 76


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