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)

One ribosomal protein kinase activity and 3 soluble protein kinase activities have been identified in plasma cell tumors by DEAE-cellulose chromatography. We have shown phosphorylation in vivo and in vitro of a protein fraction from the ribosomal KCl wash which we have termed 'PPx fraction'. Phosphorylation of this protein fraction has been obtained in vitro with the ribosome-associated protein kinase. We have determined for the ribosomal protein kinase the following characteristics. 1. It is an Mg2+-dependent enzyme that transfers the gamma-phosphate from ATP into phosphoseryl and phosphothreonyl residues of the substrate. 2. It has a wide substrate specificity. Like the soluble protein kinases it catalyses the phosphorylation of several proteins like histone, phosvitin, casein and ribosomal proteins but it differs from the main soluble kinases (I, II) by the fact that it catalyses specifically the phosphorylation at least of one of the ribosomal KCl wash proteins. On dodecylsulfate-polyacrylamide gels this protein has a molecular weight of approximately 90000 and it is released from ribosomes under conditions commonly employed for extraction of initiation factors. 3. The ribosome-associated protein kinase is not stimulated by the addition of cyclic adenosine 3':5'-monophosphate. 4. KCl has no effect, NaCl has a weak effect on the phosphorylation, Mn2+ and Ca2+ are inhibitors. 5. ADP has been found to be a competitive inhibitor. 6. The maximum velocity of the ribosomal protein-kinase-catalysed reaction is 0.65 nmol of 32P incorporated in the KCl wash protein per min and per mg protein. 7. The apparent Km for the ribosomal KCl wash protein as substrate is 0.71 mg/ml and the Km for ATP is 94 muM. 8. The molecular weight of the ribosomal protein kinase, estimated by electrophoresis in polyacrylamide-dodecylsulfate gels, is 60000 and corresponds probably to a catalytic subunit.
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PMID:A plasmocytoma ribosome-associated protein kinase which phosphorylates a specific protein of the ribosomal KCl wash. 124 81

Subcellular fractionation of oviduct tissue from estrogen-treated chicks indicated that the bulk of the protein kinase activity of this tissue is located in the cytoplasmic and nuclear fractions, DEAE-cellulose chromatography of cytosol revealed a major peak of cAMP stimulatable activity eluting at 0.2 M KCl. This peak was further characterized and found to exhibit properties consistent with cytoplasmic cAMP dependent protein kinases isolated from other tissues; it had a Km for ATP of 2 X 10(-5) M, preferred basic proteins such as histones, as substrate, and had a M of 165 000. Addition of 10(-6) M cAMP caused the holoenzyme to dissociate into cAMP binding regulatory subunit and a protein kinase catalytic subunit. Extraction of purified oviduct nuclei with 0.3 M KCl released greater than 80% of the kinase activity in this fraction. Upon elution from phospho-cellulose, the nuclear extract was resolved into two equal peaks of kinase activity (designated I and II). Peak I had a sedimentation coefficient of 3S and a Km for ATP of 13 muM. while peak II had a sedimentation coefficient of 6S and a Km for ATP of 9 muM. Both enzymes preferred alpha-casein as a substrate over phosvitin or whole histone, although they exhibited different salt-activity profiles. The cytoplasmic and nuclear enzymes were well separated on phospho-cellulose and this resin was used to quantitate the amount of cAMP dependent histone kinase activity in the nucleus and the amount of casein kinase activity in the cytosol. Protein kinase activity in nuclei from estrogen-stimulated chicks was found to be 40% greater than hormone-withdrawn animals. This increase in activity was not due to translocation of the cytoplasmic protein kinase in response to hormone, but to an increase in nuclear (casein) kinase activity. During the course of this work, we observed small but significant amounts of cAMP binding activity very tightly bound to the nuclear fraction. Solubilization of the binding activity by sonication in high salt allowed comparison studies to be performed which indicated that the nuclear binding protein is identical with the cytoplasmic cAMP binding regulatory subunit. The possible role of the nuclear binding activity is discussed.
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PMID:Protein kinases of the chick oviduct: a study of the cytoplasmic and nuclear enzymes. 126 2

Two protein kinases active on casein and phosvitin were partially purified from the soluble fraction of ejaculated bovine spermatozoa. They were operationally termed casein kinase A and B based on the order of their elution from a phosphocellulose column. CK-A showed an approximate molecular mass of 38 kDa, and it phosphorylated serine residues of casein and phosvitin utilizing ATP as a phosphate donor (Km 19 microM). Enzyme activity was maximal in the presence of 10 mM MgCl2, whereas it decreased in the presence of spermine, polylysine, quercetin, and NaCl (20-250 mM). CK-B seemed to have a monomeric structure of about 41 kDa; it underwent autophosphorylation and cross-reacted with polyclonal antibodies raised against recombinant alpha, but not beta, subunit of human type 2 casein kinase. It phosphorylated both serine and threonine residues of casein and phosvitin, utilizing ATP (Km 12 microM) but not GTP as a phosphate donor. Threonine was more affected in the phosphorylated phosvitin than in the partially dephosphorylated substrate. CK-B was active toward the synthetic peptide Ser-(Glu)5 and calmodulin (in the latter case, in the presence of polylysine), and it was activated by spermine, polylysine, MgCl2 (30 mM), and NaCl (20-400 mM). The activity of the enzymes was not affected by cAMP, or the heat-stable inhibitor of the cAMP-dependent protein kinase, or calcium.
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PMID:Purification and characterization of two casein kinases from ejaculated bovine spermatozoa. 129 85

Ca(2+)-dependent protein kinase (CDPK) was purified 900-fold from the soluble fraction of Dunaliella tertiolecta cells by ammonium sulfate precipitation, DEAE-Toyopearl, phenyl-Sepharose, and hydroxylapatite column chromatography. The CDPK was activated by micromolar concentration of Ca2+ and required neither calmodulin nor phospholipids for its activation. The enzyme phosphorylated casein, myosin light chain, and histone type III-S (histone H-1), but did not phosphorylate protamine and phosvitin. The Km values for ATP and casein were 11 microM and 300 micrograms/ml, respectively. Phosphorylation of casein was inhibited by calmodulin antagonists, calmidazolium, trifluoperazine, and compound 48/80, but not affected by calmodulin. CDPK bound to phenyl-Sepharose in the presence of Ca2+ and was eluted by ethylene glycol bis(beta-aminoethyl ether) N,N'-tetraacetic acid (EGTA). This suggests that hydrophobicity of the enzyme was increased by Ca2+. CDPK was also bound to the microsomes isolated from Dunaliella cells in the presence of micromolar concentration of Ca2+ and released in the presence of EGTA, suggesting the possibility of in vivo Ca(2+)-dependent association of the enzyme. The enzyme phosphorylated many proteins in the microsomes but few in the cytosol, if at all.
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PMID:Ca(2+)-dependent protein kinase from the halotolerant green alga Dunaliella tertiolecta: partial purification and Ca(2+)-dependent association of the enzyme to the microsomes. 131 89

1. A protein kinase type II was purified from calf thymus chromatin using ammonium sulphate fractionation, ion exchange chromatography on DEAE and phosphocellulose and affinity chromatography on phosvitin- and casein-sepharose columns. 2. The enzyme moves as a single band in non-denaturing gel electrophoresis at pH 8.3, which coincides with the enzyme activity assayed on gel slices. 3. Sodium dodecyl sulphate gel electrophoresis shows three separate polypeptide chains having M(r) of 40,000, 38,000 and 25,000, respectively. The native M(r) was about 130,000, as measured by HPLC on Superose 12 column, suggesting a subunit structure of alpha, alpha', beta 2 type. The enzyme incubated with [gamma 32P]ATP or [gamma 32P]GTP as phosphoryl donors undergoes autophosphorylation in the M(r) = 25,000 subunit. 4. The enzyme phosphorylates casein (Km = 7 microM) and phosvitin (Km = 5 microM) but not histones and was strongly deactivated by Zn2+ ions (I50 = 0.05 mM) and heparin (I50 = 0.1 micrograms/ml). 5. The enzyme seems to be the major phosphorylating system present in the 0.35 M NaCl chromatin extract of calf thymus. The RNA polymerase II from calf thymus and RNA polymerase from E. coli are both phosphorylated by protein kinase NII. The effect of phosphorylation, which causes a remarkable increase of DNA transcription rate, was studied in vitro and extensively discussed.
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PMID:Protein kinase NII from calf thymus chromatin. Isolation, characterization and some functional properties. 145 14

We have purified and characterised an apparently novel nuclear 42-kDa casein kinase from epithelial cells of Chironomus tentans which comigrates with a phosphoprotein associated with transcriptionally active salivary gland genes. The protein kinase promotes phosphorylation of casein and phosvitin, using either ATP or GTP as phosphate donors, and undergoes autophosphorylation. The casein kinase activity of the 42-kDa protein is sensitive to heparin, 5,6-dichloro-1-beta-D-ribofuranosylbezimidazole (DRB), spermine and spermidine indicating that it is a novel enzyme with similar but not identical properties to casein kinase II or nuclear protein kinase NII.
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PMID:A novel nuclear 42-kDa casein kinase identified in Chironomus tentans. 146 64

Protein kinases induced by Bombyx mori nuclear polyhedrosis virus in pupae of the silkworm B. mori were examined by activity gel analysis using phosvitin as a protein substrate. The method involved PAGE of the soluble fraction from pupae under native conditions and in the presence of SDS, followed by in situ renaturation of proteins and recovery of protein kinase activity in the intact gel. A novel protein kinase able to phosphorylate phosvitin was detected in the infected pupae from 2 days post-infection. This enzyme was not present in uninfected silkworms at any stage of the pupal period. The novel kinase activity was found by SDS-PAGE to be associated with a single polypeptide with an apparent M(r) of 50K. However, on electrophoresis under native conditions its activity was associated with a set of polypeptides with similar but not identical electrophoretic mobilities. Microheterogeneity of the catalytically active polypeptides suggests that the virus-induced protein kinase undergoes post-translational modification during the course of infection.
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PMID:Induction of a novel protein kinase in pupae of the silkworm Bombyx mori after infection with nuclear polyhedrosis virus. 146 62

The casein kinase I (CKI) family consists of widely distributed monomeric Ser/Thr protein kinases that have a preference for acidic substrates. Four mammalian isoforms are known. A full length cDNA encoding the CKI alpha isoform was cloned from a rabbit skeletal muscle cDNA library and was utilized to construct a bacterial expression vector. Active CKI alpha was expressed in Escherichia coli as a polypeptide of Mr 36,000. The protein kinase phosphorylated casein, phosvitin and a specific peptide substrate (D4). The enzyme was inhibited by the isoquinolinesulfonamide CKI-7, half-maximally at 70 microM. Heparin inhibited phosphorylation of the D4 peptide or phosvitin by CKI alpha. Polylysine activated when the D4 peptide was the substrate but had no effect on phosvitin phosphorylation. It is becoming clear that the individual CKI isoforms have different kinetic properties and hence could have quite distinct cellular functions.
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PMID:Recombinant rabbit muscle casein kinase I alpha is inhibited by heparin and activated by polylysine. 147 67

Extracellular protein kinase activity is demonstrated in intact cultured porcine aortic endothelial cells and is characterised. When cells were incubated with [gamma-32P]ATP (1 microM) a major cell surface protein, corresponding to 115 kDa, and at least four serum proteins (19, 21, 55 and 126 kDa) became phosphorylated. Protein kinase activity is released by intact endothelial cells, which is not due to cell damage, as judged by various cell viability parameters (e.g., release of marker enzymes, trypan blue exclusion). The activity of the protein kinase released amounted to 170 fmol/min per mg endothelial cell protein with phosvitin as substrate, which represents 9% of the total cellular phosvitin protein kinase activity. Repetitive incubation of endothelial cells substantially decreased phosvitin-kinase release. Exo-protein kinase is not influenced by cAMP and cGMP but is effectively inhibited by heparin (EC50, 0.3 microgram/ml). The findings clearly demonstrate: (1) exo-protein kinase is released by intact porcine aortic endothelial cells; (2) substrates of this enzyme are endothelial surface proteins and serum proteins.
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PMID:Exo-protein kinase release from intact cultured aortic endothelial cells. 150 3

Ecto-protein kinases have been detected as physiological constituents of cells. One feature of ecto-phosvitin/casein kinase (ecto-PK) is its release from the surface in a soluble form when cells are incubated with exogenous substrate protein. This is interesting in view of the fact that some ecto-enzymes are anchored to the plasma membrane via glycosylphosphatidylinositol (GPI). Such enzymes are known to be released from the surface through cleavage by a phospholipase activity. We therefore investigated whether bacterial phospholipase C (PI-PLC) was able to release ecto-PK from intact HeLa cells. The data show that whereas alkaline phosphatase, known to be GPI-anchored, was solubilized, the ecto-PK was neither released nor affected in its activity. Another effect of treatment of cells with phospholipases was the formation of diacylglycerol or phosphatidic acid which, however, did not occur when cells were incubated with phosvitin, the condition which induces ecto-PK release. These results coherently indicate that cellular phospholipases are not involved in the release mechanism of ecto-PK. Also, the presence of various protease inhibitors did not affect ecto-PK release. Cross-linking of cell-surface proteins by bifunctional agents of the succinimidyl-type suggest a protein-protein interaction responsible for membrane anchoring of the ecto-PK.
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PMID:Ecto-protein kinase release differs from cleavage by phospholipases of a glycosyl-phosphatidylinositol membrane anchor. 153 99


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