Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.13 (protein kinase C)
 49,245 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The Ca2+ ionophore A23187 (0.2-5 microM) stimulates the phosphorylation of the substrates of protein kinase C (40,000 dalton protein) and myosin light chain kinase (20,000 dalton protein) in the presence or absence of cyclooxygenase inhibitors. In the presence of cyclooxygenase inhibitors or millimolar Ca2+ there is no stimulation of phospholipase C by A23187. Fingerprints of the 32P-labeled 40,000 dalton protein isolated from platelets that have been stimulated with A23187, thrombin, phorbol 12,13-dibutyrate and 1,2-didecanoylglycerol were identical. Higher concentrations of A23187 (1-5 microM) induced the loss of polyphosphoinositides through phosphomonoesterase activity.
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PMID:Ionophore A23187 stimulates phosphorylation of the 40,000 dalton protein in human platelets without phospholipase C activation. 301 50

Calcium initiates smooth muscle contraction by binding to calmodulin and activating the enzyme myosin light chain kinase. The activated form of myosin light chain kinase phosphorylates myosin on the 20,000-dalton light chain and contractile activity ensues. Calcium may also enhance smooth muscle contractile activity by binding directly to myosin, the main component of the thick filament. Recent studies raise the possibility that the calcium-calmodulin complex may also modulate smooth muscle contractile activity by removing the inhibition imposed by caldesmon, a protein that is bound to the thin (i.e., actin-containing) filaments of smooth muscle. In vitro studies have demonstrated that the calcium-activated, phospholipid-dependent kinase, protein kinase C, can phosphorylate smooth muscle myosin at a different site than does myosin light chain kinase and down-regulate its actin-activated magnesium adenosine triphosphatase activity. This raises the possibility that protein kinase C phosphorylation of myosin may play a role in modulating vascular contractile activity in vivo.
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PMID:Effects of calcium on vascular smooth muscle contraction. 302 18

Rat tissue levels of Ca2+ . calmodulin-dependent protein kinase II (protein kinase II) and Ca2+ . phospholipid-dependent protein kinase (protein kinase C) were selectively assayed using the synthetic peptide syntide-2 as substrate. The sequence of syntide-2 (pro-leu-ala-arg-thr-leu-ser-val-ala-gly-leu-pro-gly-lys-lys) is homologous to phosphorylation site 2 in glycogen synthase. The relative Vmax/Km ratios of the known Ca2+-dependent protein kinases for syntide-2 were determined to be as follows: protein kinase II, 100; protein kinase C, 22; phosphorylase kinase, 2; myosin light chain kinase, 0.005. Levels of protein kinase II were highest in cerebrum (3.36 units/g tissue) and spleen (0.85 units/g) and lowest in testis (0.05 units/g) and kidney (0.04 units/g). Protein kinase II activity was localized predominantly in the 100,000g particulate fraction of cerebrum and testis, in the supernatant fraction of heart, liver, adrenal, and kidney, and about equally distributed between particulate and supernatant in spleen and lung. Likewise, protein kinase C activity was highest in cerebrum (0.56 units/g) and spleen (0.47 units/g), and the majority of activity was present in the cytosolic fraction for all tissues measured except for cerebrum and testis in which the kinase activity was equal in both fractions. Finally, the ratios of protein kinase II to protein kinase C were different in various rat tissues and between particulate and supernatant fractions. These results suggest somewhat different functions for these two Ca2+-regulated, multifunctional protein kinases.
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PMID:Calcium . calmodulin-dependent protein kinase II and calcium . phospholipid-dependent protein kinase activities in rat tissues assayed with a synthetic peptide. 302 65

Control of the contraction/relaxation cycle in vascular smooth muscle is regulated by Ca2+ and the cyclic nucleotides, cAMP and cGMP. For the most part, the effectors of these intracellular messengers are the protein kinases. Four major protein kinases (myosin light chain kinase, protein kinase C, cAMP dependent protein kinase, and cGMP dependent protein kinase) have been identified in vascular smooth muscle. Substantial biochemical and physiological evidence exists supporting the involvement of Ca2+/calmodulin-mediated activation of myosin light chain kinase and phosphorylation of the 20,000 dalton P-light chain of myosin in the regulation of vascular contractile activity. However, alternative hypotheses exist which suggest that additional Ca2+ dependent regulatory mechanisms reside at other contractile protein sites. Calcium also activates protein kinase C, which requires phospholipid and diacylglycerol as co-factors instead of calmodulin. Protein kinase C also phosphorylates smooth muscle myosin P-light chain; however, phosphorylation occurs at a different site on the P-light chain and represses ATPase activity which has been stimulated by myosin light chain kinase-catalyzed phosphorylation. The precise physiological role of protein kinase C in modulating vascular smooth muscle contractile activity remains to be elucidated. Relaxation of vascular smooth muscle by some different relaxants is linked to either cAMP or cGMP formation. Correlative evidence also links activation of cAMP dependent protein kinase with relaxation. Two isozymes of cAMP dependent protein kinase exist in arterial smooth muscle; potential specific roles for each isozyme have not been elucidated. Mechanistically, relaxation mediated by both cyclic nucleotide-regulated protein kinases most likely involves primary effects on Ca2+ ion flux regulation rather than direct effects on contractile protein interactions. Activation of cGMP dependent protein kinase may be important in mediating the relaxant effects of endothelium derived relaxant factor or atrial natriuretic factor. Direct pharmacological modulation of smooth muscle vascular protein kinase activity represents an approach towards developing novel vasodilator agents. Various classes of agents, including phenothiazine antipsychotics, antidepressants, naphthalene sulfonamides, and certain lipophilic Ca2+ antagonists, inhibit myosin light chain kinase activity primarily by competition with the enzyme for Ca2+-calmodulin. However, additional inhibition via binding to the myosin P-light chain may also occur with some of these agents.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Regulation of contractile activity in vascular smooth muscle by protein kinases. 302 13

Smooth muscle heavy meromyosin (HMM) is phosphorylated by the Ca2+-activated phospholipid-dependent protein kinase, i.e. protein kinase C, at three sites on each 20,000-dalton light chain. Phosphorylation of three sites also is observed with isolated 20,000-dalton light chain and HMM subfragment 1. The phosphorylation sites are serine 1, serine 2, and threonine 9. Threonine is phosphorylated most rapidly followed by either serine 1 or 2. Phosphorylation of the third site occurs only on prolonged incubation. Phosphorylation is a random process. HMM phosphorylated at two sites per light chain by protein kinase C can be dephosphorylated, as shown using two phosphatase preparations. Increasing levels of phosphorylation of HMM by protein kinase C causes a progressive inhibition of the subsequent rate of phosphorylation of serine 19 by myosin light chain kinase and causes a progressive inhibition of actin-activated ATPase activity of HMM, prephosphorylated by myosin light chain kinase. Inhibition of ATPase activity is due to a decreased affinity of HMM for actin rather than a change in Vmax. Previous results with HMM and protein kinase C (Nishikawa, M., Sellers, J. R., Adelstein, R. S., and Hidaka, H. (1984) J. Biol. Chem. 259, 8808-8814) examined effects induced by phosphorylation of the threonine residues. Our results confirm these and consider also the influence of higher levels of phosphorylation by protein kinase C.
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PMID:Phosphorylation of the 20,000-dalton light chain of smooth muscle myosin by the calcium-activated, phospholipid-dependent protein kinase. Phosphorylation sites and effects of phosphorylation. 303 66

The cellular and molecular mechanisms underlying smooth muscle contraction are reviewed in the light of recent studies of smooth muscle ultrastructure and of the role of polyphosphoinositide turnover and protein kinase C function in smooth muscle contraction. A new model of smooth muscle contraction is proposed that differs radically from accepted views, particularly the latch bridge hypothesis, in terms of both Ca2+ messenger function and the molecular events underlying this process. A coordinate fibrillar domain model of contraction is proposed in which the initial and sustained phases of contraction are mediated by different cellular and molecular events. The initial phase of response is mediated by a rise in [Ca2+]c and the resulting calmodulin-dependent activation of both myosin light chain kinase and the dissociation of caldesmon from the actin-caldesmon-tropomyosin-myosin fibrillar domain. These events lead to an interaction between actin and the phosphorylated light chains of myosin just as in previous models. However, this initial phase is followed by a sustained phase in which a rise in [Ca2+]sm stimulates the plasma membrane-associated, Ca2+-sensitive form of protein kinase C that results in the phosphorylation of both structural and regulatory components of the filamin-actin-desmin fibrillar domain. These events underlie the tonic phase of contraction.
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PMID:Protein kinase C in the regulation of smooth muscle contraction. 304 May 4

At relatively high concentrations of myosin light chain kinase, a second site on the 20,000-dalton light chain of smooth muscle myosin is phosphorylated (Ikebe, M., and Hartshorne, D. J. (1985) J. Biol. Chem. 260, 10027-10031). In this communication the site is identified and kinetics associated with its phosphorylation and dephosphorylation are described. The doubly phosphorylated 20,000-dalton light chain from turkey gizzard myosin was hydrolyzed with alpha-chymotrypsin and the phosphorylated peptide was isolated by reverse phase chromatography. Following amino acid analyses and partial sequence determinations the second site of phosphorylation is shown to be threonine 18. This site is distinct from the threonine residue phosphorylated by protein kinase C. The time courses of phosphorylation of serine 19 and threonine 18 in isolated light chains follow a single exponential indicating a random process, although the phosphorylation rates differ considerably. The values of kcat/Km for serine 19 and threonine 18 for isolated light chains are 550 and 0.2 min-1 microM-1, respectively. With intact myosin, phosphorylation of serine 19 is biphasic; kcat/Km values are 22.5 and 7.5 min-1 microM-1 for the fast and slow phases, respectively. In contrast, phosphorylation of threonine 18 in intact myosin is a random, but markedly slower process, kcat/Km = 0.44 min-1 microM-1. Dephosphorylation of doubly phosphorylated myosin (approximately 4 mol of phosphate/mol of myosin) and isolated light chains (approximately 2 mol of phosphate/mol of light chain) follows a random process and dephosphorylation of the serine 19 and threonine 18 sites occurs at similar rates.
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PMID:Identification, phosphorylation, and dephosphorylation of a second site for myosin light chain kinase on the 20,000-dalton light chain of smooth muscle myosin. 307 56

Inositol 1,4,5-trisphosphate induces aggregation and the release of [3H]5-hydroxytryptamine from human platelets rendered permeable with saponin. This action of inositol 1,4,5-trisphosphate is associated with a significant formation of thromboxane B2, activation of phospholipase C, and phosphorylation of 20,000- and 40,000-dalton proteins, which are the substrates for myosin light chain kinase and protein kinase C, respectively. All of these responses are blocked by the cyclooxygenase inhibitors indomethacin and aspirin and the dual cyclooxygenase and lipoxygenase inhibitor 3-amino-1-[m-(trifluoromethyl)phenyl]-2-pyrazoline (BW 755C). These data indicate that platelet activation by inositol 1,4,5-trisphosphate is initiated by the mobilization of Ca2+, which leads to phospholipase A2 activation. The thromboxanes and endoperoxides that are subsequently generated then induce activation via cell surface receptors.
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PMID:Inositol 1,4,5-trisphosphate induces aggregation and release of 5-hydroxytryptamine from saponin-permeabilized human platelets. 308 84

Membrane assembly of the C5b-9 proteins on gel-filtered human platelets has been shown to initiate the nonlytic release of alpha-granule contents and expression of membrane prothrombinase sites, suggesting cellular activation by these ostensibly cytolytic plasma proteins (Wiedmer, T., Esmon, C. T., and Sims, P. J. (1986) J. Biol. Chem. 261, 14587-14592). We now examine the mechanism of the C5b-9-induced release reaction. The release of alpha-granule contents upon C5b-9 assembly is accompanied by expression of alpha-granule membrane glycoprotein 140 on the platelet surface, confirming that the complement-mediated release reaction occurs by secretory fusion of the alpha-granule with the plasma membrane. C5b-9 binding initiates the phosphorylation of both 40- and 20-kDa platelet proteins, indicative of activation of protein kinase C and myosin light chain kinase, respectively. Activation of cellular protein kinases under these conditions was not accompanied by the formation of inositol phosphates and was found to strictly depend upon extracellular Ca2+, suggesting that the platelet's secretory response to the C5b-9 proteins is triggered directly by the influx of Ca2+ across the plasma membrane. measurement of intracellular Ca2+ confirmed that elevation of this ion in the cytosol was strictly dependent upon increased plasma membrane permeability due to C5b-9 assembly and was not accompanied by mobilization of this ion from internal storage pools. The C5b-9-mediated secretory response was blocked by sphingosine, a potent inhibitor of protein kinase C, but was unaffected by the cyclooxygenase inhibitor indomethacin, suggesting that feedback (receptor-linked) by thromboxane is not required for platelet activation after C5b-9 insertion.
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PMID:Complement C5b-9-stimulated platelet secretion is associated with a Ca2+-initiated activation of cellular protein kinases. 311 83

K-252a, (8R*,9S*,11S*)-(-)-9-hydroxy-9-methoxycarbonyl-8-methyl-2,3,9,10-tetr ahy dro-8,11-epoxy-1H,8H,11H-2,7b,11a-triazadi benzo[a,g]cycloocta[c,d,e]triden-1-one, an indole carbazol compound isolated from microbial origin, potently inhibits protein kinase C in partially purified enzyme and intact platelets. We examined the effects of this compound on platelet-activating factor [1-O-alkyl-alpha-acetyl-sn-glycero-phosphocholine (AGEPC)] induced protein phosphorylation, serotonin release and a rise in intracellular free calcium using washed rabbit platelets. In Ca2+-containing medium (1 mM CaCl2), AGEPC at 10(-10) and 10(-9) M markedly phosphorylated two proteins having molecular weights of 40,000 daltons (40 K protein) and 20,000 daltons (20 K protein) and evoked a marked rise in cytosolic free calcium. K-252a at 3 and 10 microM caused a concentration-dependent inhibition in the 20 K protein phosphorylation but caused only slight inhibition in the 40 K protein phosphorylation. K-252a inhibited the basal phosphorylation of 20 K protein obtained in non-stimulated platelets, and caused no significant alteration in the rise of intracellular free calcium evoked by AGEPC. It can be considered, from this evidence, that K-252a may act directly on myosin light chain kinase, resulting in the inhibition of 20 K protein phosphorylation. In Ca2+-free medium [1 mM ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA)], AGEPC at 10(-8) M predominantly phosphorylated 40K protein, although phosphorylation of 20K protein and cytosolic free calcium were increased slightly. K-252a at 1-10 microM caused a concentration-dependent inhibition in the 40K protein phosphorylation. These results indicate that K-252a functions as an inhibitor of both protein kinase C and myosin light chain kinase in rabbit platelets. In AGEPC-stimulated platelets, the inhibition of 20K protein phosphorylation in Ca2+-containing medium and of 40K protein phosphorylation in Ca2+-free medium was closely correlated with the inhibition of serotonin release by K-252a. These results strongly suggest that the phosphorylation of these two proteins may be a prerequisite for serotonin release in AGEPC-stimulated platelets.
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PMID:Parallel inhibition of platelet-activating factor-induced protein phosphorylation and serotonin release by K-252a, a new inhibitor of protein kinases, in rabbit platelets. 312 96


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