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

Arachidonic acid metabolism in resident rat alveolar macrophages and in those activated with complete Freund's adjuvant (CFA) was studied. Adult Sprague-Dawley rats were injected with 0.05 ml CFA, and macrophages were harvested 10 days later. Macrophages were labeled overnight with carbon 14-labeled arachidonic acid, washed, and then stimulated with calcium ionophore A23187 (IoA), phorbol myristate acetate (PMA), or zymosan for 30 minutes. Prostaglandins, thromboxane, and leukotrienes were extracted from the medium and analyzed by radioimmunoassay or radio high-pressure liquid chromatography. Cell lipids were analyzed by radio thin-layer chromatography. Medium and cell beta-glucuronidase activity and protein kinase C activity of the membrane fraction were also assayed. We found (1) lower leukotriene B4 (LTB4) production in stimulated resident macrophages when compared with resident macrophages after IoA stimulation--the suppressed LTB4 production was reversed by PMA; (2) unchanged or higher LTB4 production in activated macrophages when compared with resident macrophages after zymosan stimulation; (3) inhibition of zymosan-stimulated LTB4 production by staurosporine, a protein kinase C inhibitor, in both groups; and (4) lower diacylglycerol (DAG) production in activated macrophages when compared with resident macrophages after IoA stimulation, but not after zymosan stimulation. These results suggest that the reduced response of activated macrophages to IoA is due to decreased production of an endogenous protein kinase C activator. This hypothesis was further supported by the observation that protein kinase C activation in response to IoA was lower in activated macrophages than in resident macrophages. In contrast, zymosan stimulation resulted in higher protein kinase C activation in activated macrophages when compared with resident cells. We hypothesize that protein kinase activation is necessary for leukotriene production and that the preserved ability of zymosan to activate PKC via DAG accounts for the high leukotriene production in zymosan-activated macrophages. We also found that stimulated thromboxane production was higher in activated than resident cells, regardless of the stimulus, and that thromboxane production was not affected by staurosporine. Thus alterations of eicosanoid metabolism in immunologically activated macrophages depend on the stimulus used and the type of eicosanoid examined. Furthermore, leukotriene biosynthesis in rat alveolar macrophages may be regulated by protein kinase C.
J Lab Clin Med 1992 Sep
PMID:Production of leukotrienes and thromboxane by resident and activated rat alveolar macrophages: a possible role of protein kinase C. 132 31

HPV types associated with genital disease are termed "high-risk" or "low-risk" viruses according to their prevalence in cancers. Two viral genes, E6 and E7, are invariably expressed in cervical carcinomas. The E7 gene product has been found to bind the retinoblastoma tumor suppressor protein and to be phosphorylated by casein kinase II. Although present in both high- and low-risk E7 proteins, these activities are diminished in the low-risk HPV-6 E7 polypeptide. To better understand the oncogenic potential of the HPV-6 E7 protein, we replaced four of its amino acids with HPV-16 E7 residues present in the analogous region of the N-terminal half of the protein. Replacement of the arginine at position 4 of the HPV-6 E7 protein with an aspartate present in HPV-16 E7 slowed the mobility of the protein when expressed in vivo. Replacement of the glycine at position 22 with an aspartate resulted in higher affinity for retinoblastoma protein binding. Replacement of valine residues at positions 30 and 37 with asparagine and aspartate, respectively, resulted in higher levels of casein kinase II phosphorylation. The substitution at position 22 was the only mutation that exhibited increased transforming activity, suggesting a correlation between the HPV E7 protein affinity for the retinoblastoma tumor suppressor protein and its ability to transform established cells. Our results show that subtle changes in sequence may result in marked differences in biological activity of HPV oncogenes.
Proc Natl Acad Sci U S A 1992 Sep 01
PMID:Single amino acid substitutions in "low-risk" human papillomavirus (HPV) type 6 E7 protein enhance features characteristic of the "high-risk" HPV E7 oncoproteins. 132 43

Halothane directly relaxes airway smooth muscle. To determine the direct inhibitory mechanisms of halothane on canine tracheal smooth muscle contraction, the effects of this anesthetic on the levels of several intracellular second messengers were investigated by measuring intracellular Ca2+ concentration ([Ca2+]i), Ca2+/phospholipid-dependent protein kinase (PKC) translocation, and intracellular cyclic adenosine monophosphate concentration ([cAMP]i). When carbachol (1 microM) was used to increase [Ca2+]i to the same concentration as that induced by high-K+ (72.7 mM), the carbachol-induced contraction was more than twice as great, indicating that carbachol enhances the sensitivity of contractile elements to Ca2+ or activates a Ca(2+)-independent mechanism. Similarly, 12-deoxyphorbol 13-isobutylate, a potent PKC activator, markedly potentiated high-K(+)-induced muscle contraction without an increase of [Ca2+]i. The addition of halothane (0.33, 0.75, 1.15, and 1.47 mM) decreased [Ca2+]i and the muscle tension induced by carbachol. However, the decrease of muscle tension was more marked than that of [Ca2+]i at the higher concentrations. Although [Ca2+]i in the presence of verapamil and carbachol was not affected by halothane, the anesthetic markedly decreased muscle force by decreasing the "Ca2+ sensitization" or the Ca(2+)-independent enhancement of tension observed with carbachol. Halothane (0.75 and 1.47 mM) significantly released the membrane-associated PKC to cytosol, which decreased PKC activity. [cAMP]i of the smooth muscle stimulated by carbachol was moderately but significantly increased by halothane. However, when equivalent relaxation was induced with forskolin, which acts via adenylate cyclase activation, a much higher [cAMP]i was observed, which suggests that halothane acts via an additional pathway.(ABSTRACT TRUNCATED AT 250 WORDS)
Anesthesiology 1992 Sep
PMID:Direct inhibitory mechanisms of halothane on canine tracheal smooth muscle contraction. 132 45

Two isozymes of cGMP-dependent protein kinase (cGMP kinase) have been identified. Polyclonal antibodies were developed which recognize both isozymes or specifically the I alpha and I beta isoform. The specificity of these antibodies was verified by using the recombinant or purified I alpha and I beta isozymes. The antibodies cross-reacted with the purified isozymes of cGMP kinase from bovine tracheal smooth muscle. The tissue concentration of cGMP kinase was determined by ELISA. High concentrations (greater than 10 pmol/g wet tissue) were present in bovine lung, rumen, trachea, aorta, uterus and stomach. The tissue distribution of the isozymes I alpha and I beta was investigated by immunoblots using crude extracts of the different tissues. The I beta-specific antibody yielded strong signals with extracts of trachea, aorta, stomach and uterus, whereas heart, cerebellum and lung apparently contain mainly the I alpha isozyme.
Eur J Biochem 1992 Sep 01
PMID:Detection of cGMP dependent protein kinase isozymes by specific antibodies. 132 10

We examined the binding interactions of the methylation-dependent chemotaxis receptors Tsr and Tar with the chemotaxis-specific protein kinase CheA and the coupling factor CheW. Receptor directly bound CheW, but receptor-CheA binding was dependent upon the presence of CheW. These observations in combination with our previous identification of a CheW-CheA complex suggest that CheW physically links the kinase to the receptor. The ternary complex of receptor, CheW, and CheA is both kinetically and thermodynamically stable at physiological concentrations. Stability is not significantly altered by changes associated with attractant or repellent binding to the receptor. Such binding greatly modulates the kinase activity of CheA. Our results demonstrate that modulation of the kinase activity does not require association-dissociation of the ternary complex. This suggests that the receptor signal is transduced through conformational changes in the ternary complex rather than through changes in the association of the kinase CheA with receptor and/or CheW.
Cell 1992 Sep 18
PMID:Assembly of an MCP receptor, CheW, and kinase CheA complex in the bacterial chemotaxis signal transduction pathway. 132 8

We have measured the activity of the n type K+ channel present in human (Jurkat) T lymphocytes using the patch clamp technique in the whole-cell configuration. We report that protein kinase A (PKA) and protein kinase C (PKC) modulate, in a dual manner, the K+ conductance in these cells. Activation of PKA decreases the amplitude of the current, as previously reported (Bastin, B., Payet, M. D., and Dupuis, G. (1990) Cell. Immunol. 128, 385-399), and this is also the case for 12-O-tetradecanoylphorbol-13-acetate-dependent activation of PKC. In contrast, inhibitors of PKC (H7, staurosporine, polymixin B, and anti-PKC antibody) increase the current amplitude. Of importance, down-regulation of PKC or its inhibition prevented the PKA-dependent inhibition of the K+ channels. Addition of alkaline phosphatase via the patch pipette increased the K+ conductance under basal conditions and reversed the inhibition produced by PKA. The dual modulation of K+ channels in Jurkat T cells is in agreement with the presence of consensus sequences in the primary structure of the n type K+ channel.
J Biol Chem 1992 Sep 15
PMID:Dual regulation of the n type K+ channel in Jurkat T lymphocytes by protein kinases A and C. 132 19

Considerable structural similarities are present in a region of approximately 270 amino acids in most known cyclic nucleotide phosphodiesterase (PDE) sequences, opening the possibility that this region encodes the catalytic domain of the enzyme. To test this hypothesis, the structure of a high affinity cAMP PDE (cAMP-PDE) was analyzed by deletion mutations and site-directed mutagenesis. A ratPDE3 cDNA was mutated using a strategy based on fragment amplification by polymerase chain reaction. The effect of the introduced mutations was determined by expressing wild type and mutated proteins in prokaryotic and eukaryotic cells. The level of expression of the PDE protein was monitored by immunoblot analysis using two specific cAMP-PDE polyclonal antibodies and by measuring the PDE activity. After removal of a 99-amino acid region at the carboxyl terminus flanking the conserved domain, the protein retains its catalytic activity even though its Km and velocity were changed. Internal deletions at the amino terminus of this PDE showed that the enzyme activity was increased when a 97-amino acid fragment (from Tyr49 to Lys145) was removed. Further deletions within the amino terminus produced inactive proteins. Within the domain that appears essential for catalysis, 1 threonine and 2 serine residues are conserved in all PDEs. Substitutions of the invariant threonine (Thr349) present in the most conserved region with alanine, proline, or serine yielded proteins of the correct size and a level of expression comparable to the wild type PDE. However, in both expression systems used, proteins were completely devoid of the ability to hydrolyze cyclic nucleotides, except when the threonine was substituted with a serine. Conversely, mutations of 2 other conserved serine residues (Ser305 and Ser398) present in the catalytic domain either had no effect or produced changes only in Km and Vmax, but did not abolish catalytic activity. In addition, 2 histidine residues (His278 and His311) present in proximity to Thr349 appeared to be essential for the structure of the catalytic domain, since any substitution performed in these residues yielded an inactive enzyme. Mutations of a serine residue (Ser295) in the region homologous to the cAMP binding site of the regulatory subunit of the cAMP-dependent protein kinase demonstrated that this region does not have the same function in the two proteins. These data provide direct evidence that a 37-kDa domain, which in part corresponds to the region of conservation in all PDEs, contains the catalytic domain, and that threonine and histidine residues are probably involved in catalysis and/or are essential for the conformation of an active enzyme.
J Biol Chem 1992 Sep 15
PMID:Characterization of the structure of a low Km, rolipram-sensitive cAMP phosphodiesterase. Mapping of the catalytic domain. 132 38

The lamin B receptor (LBR) is an integral protein of the inner nuclear membrane that interacts with lamin B in vitro. If contains a 204-amino acid nucleoplasmic amino-terminal domain and a hydrophobic carboxyl-terminal domain with eight putative transmembrane segments. We found cell cycle-dependent phosphorylation of LBR using phosphoamino acid analysis and phosphopeptide mapping of in vivo 32P-labeled LBR immunoprecipitated from chicken cells in interphase and arrested in mitosis. LBR was phosphorylated only on serine residues in interphase and on serine and threonine residues in mitosis. Some serine residues phosphorylated in interphase were not phosphorylated in mitosis. To identify a threonine residue specifically phosphorylated in mitosis and the responsible protein kinase, wild-type and mutant LBR nucleoplasmic domain fusion proteins were phosphorylated in vitro by p34cdc2-type protein kinase. Comparisons of phosphopeptide maps to those of in vivo 32P-labeled mitotic LBR showed that Thr188 is likely to be phosphorylated by this enzyme during mitosis. These phosphorylation/dephosphorylation events may be responsible for some of the changes in the interaction between the nuclear lamina and the inner nuclear membrane that occur during mitosis.
J Biol Chem 1992 Sep 25
PMID:The lamin B receptor of the inner nuclear membrane undergoes mitosis-specific phosphorylation and is a substrate for p34cdc2-type protein kinase. 132 41

Mitogen-activated protein (MAP) kinases are 42- and 44-kD serine-threonine protein kinases that are activated by tyrosine and threonine phosphorylation in cells stimulated with mitogens and growth factors. MAP kinase and the protein kinase that activates it (MAP kinase kinase) were constitutively activated in NIH 3T3 cells infected with viruses containing either of two oncogenic forms (p35EC12, p3722W) of the c-Raf-1 protein kinase. The v-Raf proteins purified from cells infected with EC12 or 22W viruses activated MAP kinase kinase from skeletal muscle in vitro. Furthermore, a bacterially expressed v-Raf fusion protein (glutathione S-transferase-p3722W) also activated MAP kinase kinase in vitro. These findings suggest that one function of c-Raf-1 in mitogenic signaling is to phosphorylate and activate MAP kinase kinase.
Science 1992 Sep 04
PMID:Activation of mitogen-activated protein kinase kinase by v-Raf in NIH 3T3 cells and in vitro. 138 11

The UL13 open reading frame of herpes simplex virus type 1 (HSV-1) has been expressed in insect cells by a recombinant baculovirus and in Escherichia coli. In the latter case, the UL13 gene was fused to the gene for glutathione S-transferase (GST) to allow high-level expression of an 80-kDa GST-UL13 fusion protein. Antibody raised against the fusion protein reacted specifically with the 55-kDa UL13 gene product expressed by the recombinant baculovirus. This antibody also recognized a late phosphoprotein in HSV-1-infected cell lysates and a component of purified HSV-1 virions, both with the same electrophoretic mobility as the baculovirus-expressed protein. The virion component was efficiently phosphorylated in vitro by a virion-associated protein kinase. Using the same antibody, the probable homolog of the UL13 gene product was identified in HSV-2-infected cells and purified virions.
Virology 1992 Sep
PMID:Herpes simplex virus type 1 gene UL13 encodes a phosphoprotein that is a component of the virion. 132 2


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