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

Phosphorylation of beta-connectin (titin 2), an elastic protein of chicken breast muscle, occurred in the presence of [gamma-32P] ATP, 0.2 mM CaCl2 and 25 mM phosphate buffer, pH 7.0. Addition of 3 mM MgCl2 did not affect the phosphorylation. However, Ca2+ ions were required for the phosphorylation and EGTA inhibited it even if MgCl2 were present. Myosin light chain kinase (gizzard MLCK), cAMP dependent protein kinase (A kinase), and protein kinase C (C kinase) did not phosphorylate beta-connectin in vitro under optimal conditions. Thus it appears that beta-connectin, possibly containing a domain homologous with MLCK, has an autophosphorylating action.
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PMID:Autophosphorylation of beta-connectin (titin 2) in vitro. 154 1

We prepared anti-platelet 20-kDa myosin light chain (MLC-20) antibody and demonstrated diphosphorylation of MLC-20 in platelets ex vivo in the initial phase of activation by thrombin. Our results are as follows. (1) By Western blotting, using anti-MLC-20 antibody, both mono- and diphosphorylated myosin were seen in the initial phase of aggregation of platelets by thrombin. The peak of the diphosphorylation was later than that of monophosphorylation and the degree of both mono- and diphosphorylation reduced in the process of aggregation. (2) ML-7 (a synthetic inhibitor of MLCK) inhibited both mono- and diphosphorylation of myosin and also blocked aggregation of thrombin-activated platelets. However, H-7 (an inhibitor of protein kinase C) had little effect on either the (di)phosphorylation of myosin or the aggregation of thrombin-activated platelets. (3) Arg-Gly-Asp-Ser (RGDS) peptide, a synthetic anti-adhesive peptide, inhibited aggregation of thrombin-activated platelets in a dose-dependent manner (100-200 microM). However, it had little effect on either mono- or diphosphorylation of myosin in the process of the platelet aggregation stimulated by thrombin. From these results, we conclude that mono- and diphosphorylation of myosin by MLCK play a role in the initial phase of activation of thrombin-stimulated platelets in vivo and that mono- and diphosphorylation of myosin by MLCK precedes the secondary signal mediated by GPIIb/IIIa.
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PMID:Diphosphorylation of platelet myosin ex vivo in the initial phase of activation by thrombin. 164 15

Synthetic peptides corresponding to the autoinhibitory domains of calcium/calmodulin-dependent protein kinase II (CaMK-(281-309)), smooth muscle myosin light chain kinase (MLCK-(480-501)), and protein kinase C (PKC-(19-36)) as well as a peptide derived from the heat-stable inhibitor of cAMP-dependent protein kinase (PKI-tide) were tested for their inhibitory specificities. The inhibitory potencies of the four peptides were determined for each of the four protein kinases using both peptide substrates (at approximate Km concentrations) and protein substrates (at concentrations less than Km). In agreement with previous studies PKI-tide was a specific and potent inhibitor of only cAMP kinase, and none of the other inhibitory peptides gave significant inhibition of cAMP kinase at concentrations of less than 100 microM. With synthetic peptide substrates, PKC-(19-36) strongly inhibited native PKC (IC50 less than 1 microM) but also significantly inhibited autophosphorylated CaMK-II (IC50 = 30 microM) and proteolytically activated MLCK (IC50 = 35 microM). MLCK-(480-501) potently inhibited MLCK (IC50 = 0.25 microM) and also strongly inhibited both PKC and CaMK-II (IC50 = 1.4 and 1.7 microM, respectively). CaMK-(281-309) inhibited autophosphorylated CaMK-II, PKC, and proteolyzed MLCK almost equally (IC50 = 10, 38, and 48 microM, respectively). Qualitatively similar results were obtained with protein substrates. These studies validate the use of PKI-tide as a specific inhibitor of cAMP kinase in intact cell studies and suggest that PKC-(19-36) can also be used but only within a narrow concentration range. However, the autoinhibitory domain peptides from MLCK and CaMK-II are not sufficiently specific to be used in similar investigations.
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PMID:Specificities of autoinhibitory domain peptides for four protein kinases. Implications for intact cell studies of protein kinase function. 215 65

Thymus myosin, light chains and a synthetic peptide (S-S-K-R-A-K-A-K-T-T-K-K-R-P-Q-R-A-T-S-N-V-F-S) corresponding to the N-terminal sequence of smooth muscle myosin light chains were compared as substrates for calcium/calmodulin-dependent protein kinase (MLCK), calcium/phospholipid-dependent protein kinase (PKC), and a MgATP-activated protein kinase (H4PK) from lymphoid cells. All protein kinases catalyzed phosphorylation of the substrates although H4PK showed higher affinity for isolated light chains and the peptide. Phosphoamino acid analysis and analysis of thermolysin peptides established that PKC catalyzed phosphorylation of threonine-9 or 10. In addition, PKC and H4PK catalyzed phosphorylation at serine-19, the MLCK site. Collectively the data support the hypothesis that myosin filament assembly in nonmuscle cells may be regulated by a variety of calcium-dependent and calcium-independent protein kinases.
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PMID:Nonmuscle myosin phosphorylation sites for calcium-dependent and calcium-independent protein kinases. 308 Sep 87

The role of different protein kinases in the process of T cell activation has been studied using several inhibitors. The model we adopted was the activation of PBMC by monoclonal antibody OKT3. The results obtained confirm that PKC and PTK are involved. Thus, the inhibitors H-7, staurosporine, and genistein exerted a dose-dependent inhibition of CD2 up-regulation, CD25 expression, IL-2 production, and cellular proliferation. On the other hand, our data indicate that PKA is not involved since the inhibitor HA1004 was ineffective. W-7, an inhibitor of Ca(2+)-CaM protein kinases, inhibited OKT3-induced modulation of cell-surface markers and PBMC proliferation, whereas a slight increase in IL-2 release was detected at the highest dose used (20 microM). Using the MLCK inhibitor ML-9, we extended our studies to the myosin light chain kinase, which influences the organization of the cytoskeleton. ML-9-inhibited PBMC activation in terms of modulation of cell-surface markers and proliferation but stimulated IL-2 production. Similar results were obtained using the cytoskeleton disruptors demecolcine and cytochalasin B. Taken together the data described herein indicate that T cell activation is a complex event in which, aside from classical signal transduction-associated kinases PKC and PTK, at least two other kinases, Ca(2+)-CaM kinases and MLCK, seem to be involved, the latter probably through correct assembly of the cytoskeleton.
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PMID:Involvement of multiple protein kinases in CD3-mediated activation of human T lymphocytes. 790 41

The activation of human platelets is inhibited by two intracellular pathways regulated by either cGMP- or cAMP-elevating agents. There is considerable evidence that the inhibitory effects of cGMP and cAMP are mediated by the cGMP-PK and cAMP-PK, respectively, in human platelets. The cGI-PDE is an additional target for cGMP, and the cGMP-mediated elevation of cAMP levels contributes to the well known synergism between cAMP- and cGMP-elevating platelet inhibitors. Stimulation of both cAMP-PK and cGMP-PK prevents the agonist-induced activation of MLCK and PKC and inhibits the agonist-induced calcium mobilization from intracellular stores without any major effect on the ADP-regulated cation channel. These studies suggest that the inhibition of an early event of platelet activation, e.g. activation of PLC, is an effect common to both cGMP-PK and cAMP-PK stimulation. A common substrate of both cGMP-PK and cAMP-PK, the 46/50 kDa protein VASP, has been recently identified as a novel microfilament- and focal contact-associated protein whose phosphorylation correlates very well with platelet inhibition. Future investigations will have to identify the precise molecular mechanism of cyclic nucleotide inhibition of Ca2+ discharge from intracellular stores and whether cGMP-PK- and cAMP-PK-mediated VASP phosphorylation is an important component of this effect of cyclic nucleotides in human platelets.
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PMID:Role of cyclic nucleotide-dependent protein kinases and their common substrate VASP in the regulation of human platelets. 820 91

The function of the uterine smooth muscle in gestation and parturition is affected by a variety of hormones and biomolecules, some of which alter the intracellular levels of cAMP and Ca2+. Since the activity of smooth muscle MLCK has been shown to be modulated by phosphorylation, the effect of this modification of pregnant sheep myometrium (psm) MLCK by the catalytic subunit of cAMP-dependent protein kinase (PKA) and protein kinase C (PKC) was studied. In contrast to other smooth muscle MLCK reported, PKA incorporates 2.0-2.2 moles phosphate into a mole of psm MLCK both in the presence and absence of Ca(2+)-calmodulin. Modification of serine residues inhibited the activity of the enzyme. PKC also incorporated 2.0-2.1 moles of phosphate per mole psmMLCK under both conditions but had no effect on the MLCK activity. Sequential phosphorylation by PKC and PKA incorporated 3.8-4.1 moles phosphate suggesting that the amino acid residues modified by the two kinases are different. Phosphoamino acid analysis of the MLCK revealed that PKC phosphorylated serine and threonine residues. The double reciprocal plots of the enzyme activity and calmodulin concentrations showed that the Vmax of the reaction is not altered by phosphorylation by PKA but the calmodulin concentration require for half-maximal activation is increased about 4-fold. Only 10 out of 17 monoclonal antibodies to various regions of the turkey gizzard MLCK cross-reacted with psmMLCK suggesting structural differences between these enzymes. Comparison of the deduced amino acid sequence of the cDNA encoding the C-terminal half of the psmMLCK molecule showed that while cgMLCK and psmMLCK are highly homologous, a number of nonconservative substitutions are present, particularly near the PKA phosphorylation site B (S828).
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PMID:Phosphorylation and partial sequence of pregnant sheep myometrium myosin light chain kinase. 856 50

We studied enzymatic activities in sea urchin egg extracts that phosphorylate myosin regulatory light chain (MRLC) from chicken gizzard smooth muscle. The activity in the presence of EGTA showed cell cycle-dependent changes similar to that of histone H1 kinase, namely, it peaked shortly before cleavage, while that in the presence of Ca2+ ions did not show significant change during division cycle. Phosphopeptide mapping revealed that both the sites phosphorylatable by smooth muscle myosin light chain kinase (MLCK sites) and the sites phosphorylatable by protein kinase C (PKC sites) were phosphorylated in the presence or absence of Ca2+ ions. By analyses using an inhibitor of cdc2 kinase, butyrolactone-I, and ion exchange column chromatography, at least three kinases were detected as kinases that phosphorylate MRLC in vitro. These kinases phosphorylated distinct sites on MRLC. The first one, which phosphorylated the PKC sites, was identified as cdc2 kinase. The second one phosphorylated the MLCK sites in the absence of Ca2+ ions. The third one phosphorylated unknown sites. Possible implication of these activities in regulation of cytokinesis is discussed.
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PMID:Cell cycle-dependent phosphorylation of smooth muscle myosin light chain in sea urchin egg extracts. 879 90

We present here the nucleotide sequence for a cDNA clone encoding p34cdc2 from sea urchin, Hemicentrotus pulcherrimus. The obtained cDNA comprised 301 amino acid residues that contained the PSTAIRE domain to be important for binding to cyclins. Amino acid sequence similarity between this clone and other eukaryotic cdc2 sequences averaged approximately 72%. Using p13suc1-conjugated Sepharose 4B and a selective inhibitor of p34cdc2 kinase, butyrolactone I, it was first suggested that p34cdc2 kinase is involved in the phosphorylation of MRLC at both MLCK site and two PKC sites.
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PMID:Identification of p34cdc2 kinase from sea urchin Hemicentrotus pulcherrimus and its involvement in the phosphorylation of myosin II regulatory light chain in the metaphase extract. 937 Mar 2

The role of protein kinase C (PKC) in sustained contraction was examined in intestinal circular and longitudinal muscle cells. Initial contraction induced by agonists (CCK-8 and neuromedin C) was abolished by 1) inhibitors of Ca(2+) mobilization (neomycin and dimethyleicosadienoic acid), 2) calmidazolium, and 3) myosin light chain (MLC) kinase (MLCK) inhibitor KT-5926. In contrast, sustained contraction was not affected by these inhibitors but was abolished by 1) the PKC inhibitors chelerythrine and calphostin C, 2) PKC-epsilon antibody, and 3) a pseudosubstrate PKC-epsilon inhibitor. GDPbetaS abolished both initial and sustained contraction, whereas a Galpha(q/11) antibody inhibited only initial contraction, implying that sustained contraction was dependent on activation of a distinct G protein. Sustained contraction induced by epidermal growth factor was inhibited by calphostin C, PKC-alpha,beta,gamma antibody, and a pseudosubstrate PKC-alpha inhibitor. Ca(2+) (0.4 microM) induced an initial contraction in permeabilized muscle cells that was blocked by calmodulin and MLCK inhibitors and a sustained contraction that was blocked by calphostin C and a PKC-alpha,beta,gamma antibody. Thus initial contraction induced by Ca(2+), agonists, and growth factors is mediated by MLCK, whereas sustained contraction is mediated by specific Ca(2+)-dependent and -independent PKC isozymes. G protein-coupled receptors are linked to PKC activation via distinct G proteins.
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PMID:Sustained muscle contraction induced by agonists, growth factors, and Ca(2+) mediated by distinct PKC isozymes. 1089 64


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