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

---

Query: EC:2.7.11.1 (protein kinase)
81,284 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A molecular cDNA clone (P1 KIN) was isolated that encodes the human RNA-dependent P1/eIF-2 alpha protein kinase. The complete cDNA sequence of the P1 KIN cDNA was determined; the longest open reading frame (ORF) encoded a 551 amino acid protein with a deduced molecular weight of 62055 Da. Transcripts prepared from the P1 KIN cDNA by transcription in vitro with T7 RNA polymerase programmed the cell-free synthesis of a protein indistinguishable by immunoprecipitation and immunoblot gel analyses from the authentic 67-kDa P1 protein synthesized in human U cells treated with interferon (IFN). Furthermore, by use of a sensitive primer extension assay with T7 DNA polymerase, the major site of translation initiation within the deduced ORF of the P1 KIN cDNA was directly identified. Northern RNA gel-blot analysis revealed that the P1 KIN cDNA strongly hybridized to two IFN-induced mRNAs present in both human amnion U cells and HeLa cells; their sizes were 2.5 and 6 kb. Both transcripts were efficiently induced by IFN-alpha, but poorly by IFN-gamma. Polyclonal antibody was prepared against the product of the P1 KIN cDNA expressed in Escherichia coli. In Western blot analysis the antibody recognized a 67-kDa protein induced in human cells by IFN-alpha and, in addition, a 90-kDa protein whose level was not greatly altered by IFN treatment. The IFN-induced 67-kDa protein was found associated with the ribosomal salt-wash fraction of IFN-treated human cells, whereas the 90-kDa protein was predominantly in the S100 soluble fraction. The time course for the induction by IFN-alpha of RNA-dependent protein P1 kinase activity measured by immunoprecipitation was comparable to the time course for protein P1 induction measured by Western immunoblot analysis. The amino acid sequence of P1/eIF-2 alpha protein kinase deduced from the cDNA was 62% identical with the 518-residue murine TIK kinase and contained, within the carboxy-terminal half of the protein, the motifs commonly conserved among protein-serine/threonine kinases. The amino-terminal half of the P1 protein did not possess conserved kinase motifs, but did show extensive homology with vaccinia virus-predicted protein E3L.
...
PMID:Mechanism of interferon action: cDNA structure, expression, and regulation of the interferon-induced, RNA-dependent P1/eIF-2 alpha protein kinase from human cells. 137 53

The work described in this report suggests the existence of two biochemically distinguishable forms of the interferon-inducible, double-stranded RNA-dependent protein kinase. Kinase isolated from the cytosolic fraction (S-100) and the ribosome salt wash fraction of interferon-treated cells differed in their chromatographic properties. S-100 kinase eluted from a gel filtration column with M(r) = 140,000-160,000 and was predominantly anionic in nature, whereas ribosomal kinase eluted with M(r) = 66,000 and was predominantly cationic in nature. Purified preparations of S-100 kinase contained the M(r) = 66,000 subunit, P1, as the only polypeptide present in stoichiometric amounts, and thus the S-100 kinase appears to be a dimer of P1 subunits. Dimerization of the S-100 kinase was dependent on the phosphorylation state of the enzyme. Kinase isolated from S-100 was partially phosphorylated. Dephosphorylation of the S-100 kinase by treatment with alkaline phosphatase resulted in a monomeric form of the enzyme with biochemical characteristics similar to that of the ribosome salt wash kinase.
...
PMID:Cytosolic double-stranded RNA-dependent protein kinase is likely a dimer of partially phosphorylated Mr = 66,000 subunits. 137 30

The light-activated protein kinase C inhibitor, calphostin C, is shown to inhibit the ability of IL-3-dependent 32D cells to reduce the tetrazolium salt, MTT. To determine whether this inhibition was mediated through mitochondria which have been implicated in MTT reduction, isolated mitochondria were treated with calphostin C in the presence of various substrates for mitochondrial electron transport and EDTA (to exclude PKC involvement). Calphostin C extensively inhibited succinate-dependent MTT reduction (IC50 = 110nM) but had little effect on either NADH- or NADPH-dependent MTT reduction. An alternative protein kinase C inhibitor, H7, did not affect succinate-dependent mitochondrial MTT reduction, and the protein kinase A inhibitor, KT5720, had little effect on either cellular or mitochondrial MTT reduction. These results show that in addition to its role as a PKC inhibitor, calphostin C is also a potent inhibitor of succinate-dependent mitochondrial electron transport.
...
PMID:The protein kinase C inhibitor, calphostin C, inhibits succinate-dependent mitochondrial reduction of MTT by a mechanism that does not involve protein kinase C. 137 66

1. Solitary horizontal cells were isolated from catfish retinas and their membrane current was recorded with a whole-cell voltage clamp. Reducing the extracellular Ca2+ concentration produced a current that could be suppressed by dopamine. This Ca(2+)- and dopamine-sensitive current is hereafter termed I gamma. The voltage dependence, cytoplasmic regulation, and permeability of the I gamma channel suggest that it is half of a gap-junction channel. 2. I gamma was voltage and time dependent. In the steady state, the current-voltage relation displayed outward rectification at voltages more depolarized than 0 mV and a negative resistance region at voltages more hyperpolarized than -15 mV. The reversal potential was 3.3 +/- 1.5 mV when NaCl was the predominant extracellular salt and potassium-D-aspartate was the predominant intracellular salt. 3. The size of I gamma depended on the extracellular Ca2+ concentration. I gamma was maximal at external Ca2+ concentrations below 10 microM, half-maximal at 220 microM-Ca2+, and reduced to less than 4% of its maximum amplitude at external Ca2+ concentrations above 1 mM. Increasing the extracellular Ca2+ concentration reduced the amplitude of I gamma without changing the shape of the current-voltage relation or the kinetics of inactivation. Thus, rectification does not result from a voltage-dependent block by extracellular Ca2+. 4. Patches of cell membrane were voltage clamped in both the cell-attached and excised-patch configurations. In the cell-attached configuration, the addition of dopamine to the solution outside the patch pipette blocked the opening of channels within the membrane patch. Thus, dopamine closes I gamma channels by initiating an intracellular messenger cascade. In the excised-patch configuration, a maximum conductance of 145 pS was measured while Cs+ and tetraethylammonium+ (TEA+) were the only monovalent cations on both sides of the membrane. 5. The ability of dopamine to suppress I gamma was blocked by introducing an inhibitor of the cyclic AMP-dependent protein kinase, PKI5-24, into the cytoplasm. Thus, the action of dopamine is mediated by a pathway that includes the activation of a cyclic AMP-dependent kinase. 6. I gamma was suppressed by nitroprusside, an agent which activates guanylate cyclase and increases the intracellular cyclic GMP concentration. The effect of nitroprusside was not altered by the intracellular application of PKI5-24. Thus, nitroprusside suppresses I gamma through a pathway that does not include the activation of a cyclic AMP-dependent kinase.(ABSTRACT TRUNCATED AT 400 WORDS)
...
PMID:Hemi-gap-junction channels in solitary horizontal cells of the catfish retina. 138 84

Components of centrosomes are those among cellular proteins that are phosphorylated at the transition from interphase to mitosis. Using an anti-phosphoprotein antibody (CHO3) directed against isolated mitotic CHO spindles, we identified a 225-kDa centrosomal phosphocomponent in mitotic CHO cells and in cleaving sea urchin eggs. The 225-kDa protein is tightly attached to the centrosome, which allowed us to separate it from other spindle-associated factors by high salt extraction. Phosphorylation of the 225-kDa protein occurred during mitosis. This was shown by isotope labeling on gels as well as by visualization of thiophosphorylated centrosomes with an anti-thiophosphoprotein antibody (M. Cyert, T. Scherson, and M. W. Kirschner, 1988, Dev. Biol. 129, 209) after preincubation with ATP-gamma-S in vivo and in vitro. Mitotic spindles isolated from CHO cells retained their ability to phosphorylate the centrosomal component, whereas sea urchin spindles did not, possibly due to loss or inactivation of protein kinase(s) during spindle isolation. The enzyme associated with isolated CHO spindles was extractable by high salt treatment and was capable of phosphorylating many spindle components, including the 225-kDa centrosomal protein of CHO cells and sea urchin embryos. Such high salt extracts contain protein kinases, including cell cycle control protein kinase p34cdc2, suggesting that the enzyme responsible for centrosomal phosphorylation could be p34cdc2 or other downstream mitotic kinases activated by the action of p34cdc2.
...
PMID:Phosphorylation of a 225-kDa centrosomal component in mitotic CHO cells and sea urchin eggs. 139 87

Mitotic spindles isolated from prometaphase-arrested mammalian cells contain associated protein kinases that are extracted by high salt treatment. Their fractionation by ion-exchange chromatography reveals three major peaks of protein kinase activity that phosphorylate brain microtubule-associated proteins and differ in their substrate specificity. One of them has been identified as a casein kinaseII-like enzyme. A mitotic spindle-associated 325 kDa protein related to brain MAP1B is a major substrate for this casein kinase II-like enzyme. Another mitotic spindle protein kinase has been tentatively identified as a proline-directed protein kinase.
...
PMID:Protein kinases associated with isolated mitotic spindles from mammalian cells: identification of a casein kinase II-like enzyme. 140 49

Membrane organella are transported bidirectionally in cells, and the axonal transport system has provided an ideal model system for studying this bidirectional transport. Kinesin and cytoplasmic dynein were identified as candidates for the motor molecules of fast axonal transport, which transport organella along microtubules anterogradely and retrogradely. However, the mechanism that controls this bidirectional transport is unknown. Our previous work revealed that kinesin in axons was associated abundantly with anterogradely transported membranous organella, most of which are believed to be precursors of synaptic vesicles and axonal plasma membranes, while the fractions bound to retrogradely transported ones were very small (Hirokawa, N., Sato-Yoshitake, R., Kobayashi, N., Pfister, K. K., Bloom, G. S., and Brady, S. T. (1991) J. Cell Biol. 114, 295-302). Here we demonstrated in vitro that the binding of kinesin to synaptic vesicles was concentration-dependent and saturable and could be released by high salt concentration. When kinesin was phosphorylated by cAMP-dependent protein kinase, its binding to symaptic vesicles was significantly reduced. By motility assay and by statistical analysis using electron microscopy, we further revealed that synaptic vesicles preincubated with phosphorylated kinesin associated less frequently with microtubules than synaptic vesicles preincubated with unphosphorylated kinesin. The phosphorylation of kinesin should therefore play an essential role in regulating the direction of fast axonal transport by inhibiting its binding to membrane organella, thus releasing it from membrane organella at nerve terminals.
...
PMID:The phosphorylation of kinesin regulates its binding to synaptic vesicles. 142 30

To study the phosphorylation of one of the G-box binding factors from Arabidopsis (GBF1), we have obtained large amounts of this protein by expression in Escherichia coli. Bacterial GBF1 was shown to be phosphorylated very efficiently by nuclear extracts from broccoli. The phosphorylation activity was partially purified by chromatography on heparin-Sepharose and DEAE-cellulose and was characterized. It showed the essential features of casein kinase II activity: utilization of GTP in addition to ATP as a phosphate donor, strong inhibition by heparin, preference for acidic protein substrates, salt-induced binding to phosphocellulose, and salt-dependent deaggregation. The very low Km value for GBF1 (220 nM compared to approximately 10 microM for casein) was in the range observed for identified physiological substrates of casein kinase II. Phosphorylation of GBF1 resulted in stimulation of the G-box binding activity and formation of a slower migrating protein-DNA complex. The conditions of this stimulatory reaction fully corresponded to the properties of casein kinase II, in particular its dependence on the known phosphate donors. The DNA binding activity of the endogenous plant GBF was shown to be reduced by treatment with calf alkaline phosphatase; this reduction was diminished by addition of fluoride and phosphate or incubation in the presence of casein kinase II and ATP.
...
PMID:DNA binding activity of the Arabidopsis G-box binding factor GBF1 is stimulated by phosphorylation by casein kinase II from broccoli. 152 62

We found that a preparation of the 90-kDa heat shock protein, HSP90, purified to apparent homogeneity, contains a serine/threonine kinase which phosphorylates HSP90. The protein kinase was identified as casein kinase II (CKII) according to its properties. The protein kinase was separable from HSP90 by adsorption to heparin-Sepharose or phosphocellulose. CKII was coimmunoprecipitated with HSP90 by anti-HSP90 antibodies from cell extracts. Sucrose density gradient centrifugation analysis revealed that an addition of anti-HSP90 antibodies to cell extracts induces a shift of the sedimentation peak of CKII toward the bottom of a centrifuge tube. These results suggest that CKII is associated with HSP90 in cell lysates at low salt conditions. Furthermore, the CKII.HSP90 complex was reconstituted from purified HSP90-free CKII and CKII-free HSP90. In a buffer at low ionic strength, CKII forms large aggregates, but HSP90 dissociates the aggregates. Finally, we found that HSP90 activates CKII; an addition of HSP90 to CKII dramatically increased phosphorylation of exogenous substrates as well as the CKII beta subunit. Taken altogether, these observations suggest that CKII is structurally and functionally active when it forms a complex with HSP90.
...
PMID:The 90-kDa heat shock protein, HSP90, binds and protects casein kinase II from self-aggregation and enhances its kinase activity. 155 11

Synapsin IIa belongs to a family of neuron-specific phosphoproteins called synapsins, which are associated with synaptic vesicles in presynaptic nerve terminals. In order to examine the biochemical properties of synapsin IIa, and ultimately its physiological function, purified protein is required. Since attempts to purify significant quantities of synapsin IIa, an isoform of the synapsins, from mammalian brain have proven difficult, we undertook the production of recombinant synapsin IIa by utilizing the baculovirus expression system. Rat synapsin IIa cDNA was introduced into the baculovirus genome via homologous recombination, and the recombinant baculovirus was purified. Spodoptera frugiperda (Sf9) cells infected with this virus expressed synapsin IIa as 5% of the total cellular protein. The recombinant protein was extracted from the particulate fraction of the infected Sf9 cells with salt and a nonionic detergent and purified by immunoaffinity chromatography. The purified synapsin IIa was phosphorylated by the catalytic subunit of cAMP-dependent protein kinase to a stoichiometry of 0.8 mol of phosphate/mol of protein. Metabolic labeling with [32P]Pi demonstrated synapsin IIa phosphorylation in infected Sf9 cells. Using a homogenate of uninfected Sf9 cells, a cAMP-dependent protein kinase activity which can phosphorylate synapsin IIa was detected. Limited proteolysis of recombinant synapsin IIa phosphorylated in vitro and in vivo resulted in identical phosphopeptide maps. Further, synapsin IIa, like synapsin I, binds with high affinity in a saturable manner to synaptic vesicles purified from rat cortex.
...
PMID:Synapsin IIa: expression in insect cells, purification, and characterization. 156 72


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

---