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)

Despite the extensive literature on the biological actions of Ca2+ and calmodulin, very little is known about their involvement in nuclear functions, e.g., regulation of specific gene expression. To date, the only genes other than prolactin and growth hormone shown to be regulated by perturbations in cell Ca2+ are those coding for two glucose-regulated proteins. However, there is a growing body of indirect evidence for nuclear functions of Ca2+ and calmodulin, and we suspect that other examples of Ca2+-regulated genes will emerge. We have described in this chapter several different experimental approaches which we have employed to examine first whether prolactin gene expression is regulated by changes in cell Ca2+ content, and then to begin searching for the components of the mechanism by which Ca2+ exerts its effects on the prolactin gene. The tentative identification of 56-kDa nuclear matrix protein as both a calmodulin-binding protein and a substrate of a Ca2+-calmodulin-dependent protein kinase suggests that NMP 56 may be a subunit of a multifunctional Ca2+-calmodulin-protein kinase. This enzyme was recently detected in the nuclear matrix fraction of neuronal nuclei, and was shown to phosphorylate a chromatin protein similar to high mobility group protein 17 (HMG 17). Since HMG 17 is associated with actively transcribed chromatin, its phosphorylation in GH3 cells might play a role in the Ca2+-calmodulin-dependent regulation of prolactin gene expression by hormones and growth factors.
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PMID:Ca2+/calmodulin regulation of prolactin gene expression. 358 41

Phospholamban, the cardiac sarcoplasmic reticulum proteolipid, is phosphorylated by cAMP-dependent protein kinase, by Ca2+/phospholipid-dependent protein kinase, and by an endogenous Ca2+/calmodulin-dependent protein kinase, the identity of which remains to be defined. The aim of this study was therefore to characterize the latter kinase, called phospholamban kinase. Phospholamban kinase was purified approximately 42-fold with a yield of 11%. The purified fraction exhibits a specific activity of 6.5 nmol of phosphate incorporated into exogenous phospholamban per minute per milligram of protein. Phospholamban kinase appears to be a high molecular weight enzyme and presents a broad substrate specificity, synapsin-1, glycogen synthase, and smooth muscle myosin regulatory light chain being the best substrates. Phospholamban kinase phosphorylates synapsin-1 on a Mr 30 000 peptide. The enzyme exhibits an optimum pH of 8.6, a Km for ATP of 9 microM, and a requirement for Mg2+ ions. These data suggest that phospholamban kinase might be an isoenzyme of the multifunctional Ca2+/calmodulin-dependent protein kinase. Consequently we have searched for Mr 50 000-60 000 phosphorylatable subunits among cardiac sarcoplasmic reticulum proteins. A Mr 56 000 protein was found to be phosphorylated in the presence of Ca2+/calmodulin. Such phosphorylation alters the electrophoretic migration velocity of the protein. In addition, this protein that binds calmodulin was always found to be present in fractions containing phospholamban kinase activity. This Mr 56 000 protein is therefore a good candidate for being a subunit of phospholamban kinase. However, the Mr 56 000 calmodulin-binding protein and the Mr 53 000 intrinsic glycoprotein which binds ATP are two distinct entities.
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PMID:Characterization and partial purification of cardiac sarcoplasmic reticulum phospholamban kinase. 373 Mar 67

Calmodulin was isolated and purified to homogeneity from dog pancreas. Highly purified subcellular fractions were prepared from dog pancreas by zonal sucrose-density ultracentrifugation and assayed for their ability to bind 125I-calmodulin in vitro. Proteins contained in these fractions were also examined for binding of 125I-calmodulin after their separation by polyacrylamide-gel electrophoresis in SDS. Calmodulin-binding proteins were detected in all subcellular fractions except the zymogen granule and zymogen-granule membrane fractions. One calmodulin-binding protein (Mr 240,000), observed in a washed smooth-microsomal fraction, has properties similar to those of alpha-fodrin. The postribosomal-supernatant fraction contained three prominent calmodulin-binding proteins, with apparent Mr values of 62,000, 50,000 and 40,000. Calmodulin-binding proteins, prepared from a postmicrosomal-supernatant fraction by Ca2+-dependent affinity chromatography on immobilized calmodulin, exhibited calmodulin-dependent phosphodiesterase, protein phosphatase and protein kinase activities. In the presence of Ca2+ and calmodulin, phosphorylation of smooth-muscle myosin light chain and brain synapsin and autophosphorylation of a Mr-50,000 protein were observed. Analysis of the protein composition of the preparation by SDS/polyacrylamide-gel electrophoresis revealed a major protein of Mr 50,000 which bound 125I-calmodulin. This protein shares characteristics with the calmodulin-dependent multifunctional protein kinase (kinase II) recently observed to have a widespread distribution. The possible role of calmodulin-binding proteins and calmodulin-regulated enzymes in the regulation of exocrine pancreatic protein synthesis and secretion is discussed.
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PMID:Calmodulin-binding proteins and calmodulin-regulated enzymes in dog pancreas. 382 65

The association of regulatory subunits (RII) of Type II cAMP-dependent protein kinase from bovine cerebral cortex (RII-B) and bovine cardiac and skeletal muscle (RII-H) with specific binding proteins in bovine brain cytosol and purified brain microtubules was demonstrated using a solid phase binding assay. RII-binding proteins present in bovine cerebral cortex were immobilized on nitrocellulose filters after separation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Incubation of the filters with 32P-labeled regulatory subunits showed that both RII-B and RII-H interact with the 75,000-dalton calmodulin-binding protein (P75) and microtubule-associated protein 2 (MAP-2). However, significant differences in binding affinities and capacities were observed. RII-B displayed a higher affinity for P75 compared to RII-H while RII-H preferentially bound to MAP-2. Quantitation of radioactive RII bound to MAP-2 showed that MAP-2 bound 4-6 times more RII-H than RII-B. The differential binding affinities and capacities of RII-H and RII-B for MAP-2 were not affected by autophosphorylation since both phospho and dephospho forms of RII displayed the same binding characteristics. Competitive binding studies suggest that RII-H and RII-B bind to the same sites on MAP-2. The biochemical basis for the differential binding of RII-B and RII-H to the same sites of MAP-2 is unknown. However, other high affinity RII-binding proteins present in cerebral cortex (i.e. P75) might affect the affinity of RII-B for MAP-2. 32P-RI did not bind to P75 nor MAP-2 under the conditions used.
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PMID:Differential binding of the regulatory subunits (RII) of cAMP-dependent protein kinase II from bovine brain and muscle to RII-binding proteins. 394 17

Subcellular fractions prepared from rodent forebrain at different postnatal ages were examined for calmodulin-binding proteins using [125I]calmodulin and a gel overlay technique. Synaptic junction (SJ) fractions from newborn brain, which display purity comparable to adult SJ fractions, contain low but detectable amounts of 60 and 50 kdalton calmodulin-binding polypeptides; the latter being the major postsynaptic density protein. These polypeptides have recently been shown to be the calmodulin-binding protein subunits of calmodulin-dependent protein kinase II (CaM-kinase II). CaM-kinase II polypeptides represented the predominent calmodulin-binding proteins in nearly every subcellular fraction examined, regardless of postnatal age. Large increases were observed in the CaM-kinase II content of every subcellular fraction throughout postnatal development. During development, a striking shift in the subcellular distribution of CaM-kinase Ii was observed. Over 4 times as much CaM-kinase II was cytosolic relative to particulate in newborn brain while this ratio was completely reversed in adult brain. Large age-dependent increases in particulate-associated CaM-kinase II were observed in highly purified synaptic plasma membrane (5-fold) and SJ (14-fold) fractions. The CaM-kinase II content of SJ fractions increased approximately 70% between days 24 and 90, a period in development that follows the most active stages of synapse formation in situ. In adult brain, approximately 60% of CaM-kinase II in crude synaptosomal fractions (P2-INT) was recovered in SJ fractions. The CaM-kinase II in SPM fractions from all developmental ages resists solubilization in Triton X-100 and greater than 90% is recovered in SJ fractions. These studies indicate that during brain development the accumulation of SJ-associated CaM-kinase II represents an important process in the molecular and enzymatic maturation of CNS postsynaptic structures.
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PMID:Changes in the subcellular distribution of calmodulin-kinase II during brain development. 398 14

Myosin light chain kinase and a fraction of type II cAMP-dependent protein kinase have been partially purified from bovine brain by affinity chromatography on calmodulin-Sepharose. The myosin kinase was purified approximately 3700-fold and has an estimated molecular weight of 130,000 +/- 10,000 by sodium dodecyl sulfate gel electrophoresis. A fraction of soluble cAMP-dependent protein kinase also bound to calmodulin-Sepharose and was purified 2300-fold. A fraction of this cAMP-dependent protein kinase after purification by glycerol gradient centrifugation was shown to contain the two subunits of calcineurin, a major calmodulin-binding protein in brain, and the two subunits of type II cAMP-dependent protein kinase in a ratio of 1:1:2:2. Its sedimentation coefficient was 8.1 S and 9.0 S when centrifuged in the absence or presence of calmodulin, suggesting the formation of a complex between calmodulin and protein kinase. Our results suggest the possibility that calcineurin may be involved in the interaction between the protein kinase and calmodulin. Furthermore, our studies imply that the regulatory subunit of the cAMP-dependent protein kinase, but not the catalytic subunit, is the site of interaction with calmodulin since the catalytic subunit of protein kinase was partially resolved from the complex by cAMP.
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PMID:Interaction of calmodulin with myosin light chain kinase and cAMP-dependent protein kinase in bovine brain. 626 40

The localization of cGMP, cGMP-dependent protein kinase, calmodulin and the calmodulin-binding protein calcineurin in Paramecium tetrauelia cells has been examined with immunocytochemical methods. These molecules appeared to be localized to a large extent in the cilia of this protozoan. To ascertain that antibodies had access to all cellular compartments we have used three different preparations for immunocytochemistry: (i) with 'whole cell' preparations immunofluorescent staining for the four molecules was mainly visible in the cilia; (ii) in 'deciliated' Paramecium, staining for cGMP and calmodulin was found in regular patterns on the cell surface most likely representing kinetosomes; (iii) using 'sectioned cells', additional cytoplasmic calmodulin appeared to be associated with glycogen particles as evidenced by the disappearance of the granular staining pattern after preincubation with alpha-amylase. In contrast, cGMP, cGMP-dependent protein kinase and calcineurin fluorescence was only very weak and diffuse in cell bodies. No nuclear fluorescence was detectable after staining with any of the antibodies. Because of the colocalization of cGMP, cGMP-dependent protein kinase, a guanylate cyclase-calmodulin-complex, and calcineurin in cilia from Paramecium, an involvement of these components in the regulation of ciliary activity is discussed.
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PMID:Immunocytochemical localization of cyclic GMP, cGMP-dependent protein kinase, calmodulin and calcineurin in Paramecium tetraurelia. 632 Nov 86

A simple and rapid procedure for the purification of the native form of chicken gizzard myosin light-chain kinase (Mr 136000) is described which eliminates problems of proteolysis previously encountered. During this procedure, a calmodulin-binding protein of Mr 141000, which previously co-purified with the myosin light-chain kinase, is removed and shown to be a distinct protein on the basis of lack of kinase activity, different chymotryptic peptide maps, lack of cross-reactivity with a monoclonal antibody to turkey gizzard myosin light-chain kinase, and lack of phosphorylation by the purified catalytic subunit of cyclic AMP-dependent protein kinase. This Mr-141000 calmodulin-binding protein is identified as caldesmon on the basis of Ca2+-dependent interaction with calmodulin, subunit Mr, Ca2+-independent interaction with skeletal-muscle F-actin, Ca2+-dependent competition between calmodulin and F-actin for caldesmon, and tissue content.
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PMID:Isolation of the native form of chicken gizzard myosin light-chain kinase. 632 48

A calmodulin-binding protein is present in extracts of the macrophage-like mouse cell line J774 and in extracts of thioglycollate-stimulated mouse peritoneal macrophages; it is deficient in variants of J774 resistant to trifluoperazine and in resident peritoneal macrophages. The calmodulin-binding protein [CaMBP (J7)0.5] was purified from J774 and resolved from endogenous cyclic nucleotide phosphodiesterase and protein kinase activities. The protein has an apparent native Mr of 125,000-150,000 and binds calmodulin in a calcium-dependent manner with a Kd of 20 nM. It inhibits the ability of calmodulin to activate phosphodiesterase. Its sedimentation constant in glycerol gradients containing calmodulin was dependent upon the relative concentrations of calmodulin and the calmodulin-binding protein.
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PMID:Characterization of a calmodulin-binding protein that is deficient in trifluoperazine-resistant variants of the macrophage-like cell line J774. 657 95

Previous work from our laboratory has demonstrated that neurohumoral stimulation of the exocrine pancreas is associated with the phosphorylation of the Mr 29,000 ribosomal protein S6. In a cell-free system using pancreatic postmicrosomal supernatant as the kinase donor, we found that the following co-factors stimulate the phosphorylation of the Mr 29,000 ribosomal protein: calcium with calmodulin, calcium with phosphatidyl serine, and cAMP. These findings suggest that the pancreas contains a calcium-calmodulin-dependent protein kinase (CaM-PK) that can phosphorylate the Mr 29,000 ribosomal protein. A CaM-PK activity was partially purified sequentially by ion exchange, gel filtration, and calmodulin-affinity chromatography. Phosphorylation of the Mr 29,000 ribosomal protein by the partially purified CaM-PK was dependent on the presence of both calcium and calmodulin and not on the other co-factors. The CaM-PK fraction contained a phosphoprotein of Mr 51,000 whose phosphorylation was also dependent on calcium and calmodulin. When 125I-calmodulin-binding proteins from the CaM-PK fraction were identified using electrophoretic transfers of SDS-polyacrylamide gels, a single Mr 51,000 protein was labeled. The preparation enriched in CaM-PK activity contained an Mr 51,000 protein that underwent phosphorylation in a calcium-calmodulin-dependent manner and an Mr 51,000 calmodulin-binding protein. It is therefore possible that the CaM-PK may comprise a calmodulin-binding phosphoprotein component of Mr 51,000.
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PMID:Calmodulin-stimulated protein kinase activity from rat pancreas. 661 94


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