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)

Human myelin basic protein (MBP) was fractionated into several of its charge isomers (components). Of these, the secondary structures of four isomers before and after phosphorylation have been studied by circular dichroism (CD). None of the four showed any alpha-helical structure. All of the components showed varying amounts of beta-structure, random structure, and turns. Component 1 (C-1), the most cationic of the components, showed 13%; component 2 (C-2) had 19%; C-3, 17%; and C-4, 24% of beta-structure. Each of the four components was phosphorylated with protein kinase C, from human brain. The extent of phosphorylation varied considerably from 2.8 +/- 0.6 mol of PO4/mol of protein in C-1 to 5.2 +/- 0.8 mol of PO4/mol of protein in C-4. The effect of phosphorylation on the secondary structure was to induce beta-structure in all the components. The largest change in beta-structure was in C-1 and the least in C-4. The surprising result is that although the components were phosphorylated to different extents, the amount of beta-structure in all four components increased to a final proportion of 35-40%. Treatment of phosphorylated C-1 with acid phosphatase removed 50% of the total radioactivity. Although the remainder represented approximately 1 mol of PO4/mol of protein, the proportion of beta-structure was unaltered. We concluded that a single phosphorylation site identified as residues 5-13 represented a critical size for stabilization of beta-structure of MBP in solution and that phosphorylation at the other sites had little influence on secondary structure.
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PMID:Secondary structure of charge isomers of myelin basic protein before and after phosphorylation. 247 58

We have previously shown that cyclic AMP (cAMP) inhibits the protein kinase C (PKC)-mediated phosphorylation of myelin basic protein (MBP) in cultured oligodendrocytes (OLGs). Recently, it has been demonstrated that the long chain base sphingosine inhibits PKC by competing PKC effectors (diacylglycerol and phorbol esters) for a binding site on the kinase (Hannun and Bell: Science 235: 670-674, 1987). In this report we define further the mechanism by which cAMP inhibits MBP phosphorylation by comparing the effects of cAMP with that of galactosylsphingosine (psychosine), a potential catabolite of galactocerebroside, the major OLG glycosphingolipid. We identify the consequences of psychosine treatment and PKC down-regulation on OLG morphology and electrophysiology and discuss their relevance. Our results in intact ovine oligodendrocytes are consistent with a mechanism in which cAMP inhibits MBP phosphorylation by interfering with the release of diacylglycerol (DAG) from phosphatidylinositol. First, the effects of cAMP on MBP phosphorylation are reversed with exogenous TPA; and second, cAMP inhibits the incorporation of 1-[14C]arachidonate into DAG and specifically inhibits the turnover (as judged by 32PO4 3-incorporation) of phosphatidylinositol. Psychosine inhibits MBP phosphorylation, and its action can be reversed by TPA suggesting a mechanism of inhibition similar to that described for other systems. In addition, psychosine has profound effects on OLG morphology; it disintegrates OLG processes while leaving the cell soma intact. Stable hyperpolarized resting potentials were obtained following psychosine treatment, but there was a 66% decrease in membrane capacitance indicating a significant decrement in membrane surface area. The morphological changes induced by psychosine are reversible and can be eliminated by removing the drug but not by the addition of TPA. Whether inhibition of PKC by psychosine plays any role in process dissolution remains an unanswered question. However, current evidence suggests that a PKC-independent mechanism may be at play. This investigation in conjunction with our previous work emphasizes a role for the interregulation of protein kinase A (PKA) and PKC in the control of OLG somal vs. myelin components. This may have significant implications for central nervous system myelin assembly.
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PMID:Phosphorylation of myelin basic protein in intact oligodendrocytes: inhibition by galactosylsphingosine and cyclic AMP. 247 66

Rabbit myelin basic protein (MBP) was phosphorylated by a ganglioside-stimulated protein kinase to a stoichiometry of 1.4 and 2.1 mol phosphate/mol MBP in the presence and absence of GTlb, respectively. Two-dimensional peptide mapping analyses revealed that two of the sites of phosphorylation were distinct from those catalyzed by cAMP-dependent protein kinase or protein kinase C. Phosphorylation of one of these sites by ganglioside-stimulated protein kinase was inhibited by GTlb, suggesting that the inhibitory effect of gangliosides on MBP phosphorylation may be substrate-directed. Although ganglioside-stimulated protein kinase did not phosphorylate MBP at a domain containing residues 82-117, a synthetic peptide Arg-Phe-Ser-Trp-Gly-Ala-Glu-Gly-Gln-Lys corresponding to residues 111-120 was phosphorylated by the kinase in a ganglioside-stimulated manner. These findings suggest that the conformation of MBP may be important in determining its phosphorylatability.
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PMID:Phosphorylation of myelin basic protein and peptides by ganglioside-stimulated protein kinase. 248 Jan 29

Compound 1,2-isopropylidene-3-decanoyl-sn-glycerol (IpOCOC9) augments the phosphorylation in vitro of histone III-S and myelin basic protein (MBP) by a partially purified Ca2(+)- and phospholipid-dependent protein kinase activity (protein kinase C) from human polymorphonuclear leukocytes. IpOCOC9 can substitute for either Ca2+ and phosphatidylserine or for phorbol ester. The related compound decanoid acid cyclopentyl methylester (DACPME) is less effective than IpOCOC9 in this respect. These data lend support to the notion that the secretagogue activity of IpOCOC9 with respect to human basophil histamine release and neutrophil superoxide radical generation is due to protein kinase C activation.
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PMID:Activation of human neutrophil protein kinase C in vitro by 1,2-isopropylidene-3-decanoyl-sn-glycerol (IpOCOC9). 248 37

We have previously found that treatment of quiescent mammalian fibroblast cells with several mitogenic factors activates in common a Ca2+-sensitive serine/threonine-specific protein kinase activity toward microtubule-associated protein 2 (MAP2) [Hoshi, M., Nishida, E. and Sakai, H. (1988) J. Biol. Chem. 263, 5396-5401]. Here, we characterized the mitogen-activated MAP2 kinase activity in rat 3Y1 cells. The activated kinase activity was detected in the cytosolic fraction but not in the membrane fraction. The inhibitory effect of Ca2+ on the kinase activity was reversible. Kinetic analyses revealed that the apparent Km values of the kinase activity for MAP2 and ATP were 1.6 microM and 30 microM, respectively. Free Ca2+ at 4 microM decreased apparent Vmax values for MAP2 and ATP without changing the apparent Km values. The MAP2 kinase had an apparent molecular mass of about 40 kDa as determined by gel filtration and by sucrose density gradient centrifugation. Myelin basic protein as well as MAP2 could serve as good substrates for this kinase, but 40S ribosomal protein S6, casein, histone, phosphorylase b, protamine, tubulin, actin and tau could not. These properties of the enzyme indicate that the Ca2+-sensitive MAP2 kinase may be a previously unidentified enzyme. Down-regulation of protein kinase C by prolonged phorbol ester treatment abolished the MAP2 kinase activation by phorbol ester, but did not prevent the MAP2 kinase activation by epidermal growth factor (EGF) or fresh serum. This suggests that the Ca2+-sensitive MAP2 kinase could be activated through protein-kinase-C-dependent and -independent pathways. Activation of the MAP2 kinase occurred shortly after the addition of EGF or phorbol ester even in the presence of protein synthesis inhibitors (cycloheximide, puromycin and emetin). Moreover, treatment of the EGF- or phorbol-ester-activated MAP2 kinase with acid phosphatase inactivated the kinase activity. Thus, the MAP2 kinase may be activated through phosphorylation.
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PMID:Characterization of a mitogen-activated, Ca2+-sensitive microtubule-associated protein-2 kinase. 255 90

Myelin basic protein, an 80-kilodalton (kDa) protein in rat oligodendrocytes, and an 80-kDa basic protein in neuroblastoma x neonatal Chinese hamster brain explant hybrids were phosphorylated extensively when the cells were treated with either phorbol esters (TPA) or diacylglycerols (e.g., oleyoyl-acetylglycerol). TPA-stimulated phosphorylation was inhibited by pre-incubation with 50 microM psychosine (galactosyl-sphingosine), confirming that it is mediated through the phospholipid-dependent protein kinase C (PK-C). Surprisingly, phosphorylation of these proteins was inhibited by incubation of cells with agents which result in activation of cyclic-AMP-dependent protein kinase (dibutyryl cyclic AMP or forskolin). In contrast, phosphorylation of other nonbasic proteins, for example, the oligodendrocyte-specific 2',3'-cyclic nucleotide phosphohydrolase, was stimulated under these conditions (Vartanian et al.: Proceedings of the National Academy of Sciences of the United States of America 85:939, 1988). The possible role of cyclic AMP in activating specific phosphatases or restricting the availability of diacylglycerol for PK-C activation is discussed.
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PMID:Differential regulation of basic protein phosphorylation by calcium phospholipid and cyclic-AMP-dependent protein kinases. 277 6

Calf spleen profilin is shown to be an in vitro substrate of purified human placental protein kinase C (PKC), with an apparent Km of 4 microM. Phosphatidylinositol bisphosphate (PIP2) was an effective activator of the profilin phosphorylation by PKC and caused a maximum 13-fold increase of Vmax with a half maximal effect at 40 micrograms/ml. The action of PIP2 was not mimicked by phosphatidylserine, phosphatidic acid or phosphatidylinositol, whereas phosphatidylinositol monophosphate was slightly stimulatory. By contrast, protein kinase C-dependent phosphorylation of histone type III-S, myelin basic protein or lipocortin-I was not affected by PIP. It is suggested that PIP2 modifies the nature of the profilin-PKC interactions.
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PMID:Protein kinase C-dependent phosphorylation of profilin is specifically stimulated by phosphatidylinositol bisphosphate (PIP2). 282 77

A synthetic peptide ArgThrProProProSerGly with sequence similar to the threonine sites of phosphorylation in both myelin basic protein and simian virus 40 T antigen could be phosphorylated in vitro by a purified rat brain Ca2+-activated and phospholipid-dependent protein kinase, protein kinase C. The apparent Km and Vm values of this heptapeptide for the enzyme were determined to be 240 microM and 60 nmol/min/mg, respectively. Up to 0.8 mol 32P could be incorporated into the peptide, mainly at the threonine residue. Substitution of the L-threonine residue in the heptapeptide by its D-enantiomer abolished the phosphorylatability of the peptide by protein kinase C. However, this (D)Thr-containing peptide could act as a competitive inhibitor for the kinase with an apparent Ki value of approximately 320 microM. These findings suggest that a triprolyl sequence may act as a recognition site for protein kinase C.
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PMID:Substrate specificity of rat brain calcium-activated and phospholipid-dependent protein kinase. 308 Oct 2

Biochemical characteristics of three rat brain protein kinase C isozymes, types I, II, and III, were compared with respect to their protein kinase and phorbol ester-binding activities. All three isozymes appeared to be alike in their phorbol ester-binding activities as evidenced by their similar Kd for phorbol 12,13-dibutyrate and requirements for Ca2+ and phospholipids. However, differences with respect to the effector-mediated stimulation of protein kinase activity were detectable among these isozymes. The type I enzyme could be stimulated by cardiolipin to a greater extent than those of the type II and III enzymes. In the presence of cardiolipin, the concentrations of dioleoylglycerol or phorbol 12,13-dibutyrate required for half-maximal activation (A1/2) of the type I enzyme were nearly an order of magnitude lower than those for the type II and III enzymes. In the presence of phosphatidylserine, differences in the A1/2 of dioleoylglycerol and phorbol 12,13-dibutyrate for the three isozymes of protein kinase C were less significant than those measured in the presence of cardiolipin. Nevertheless, the A1/2 of these two activators for the type I enzyme were lower than those for the type II and III enzymes. At high levels of phosphatidylserine (greater than 15 mol %), binding of phorbol 12,13-dibutyrate to the type I enzyme evoked a corresponding stimulation of the kinase activity, whereas binding of this phorbol ester to the type II and III enzymes produced a lesser degree of kinase stimulation. For all three isozymes, the concentrations of phosphatidylserine required for half-maximum [3H]phorbol 12,13-dibutyrate binding were almost an order of magnitude less than those for kinase stimulation. Consequently, neither isozyme exhibited a significant kinase activity at lower levels of phosphatidylserine (less than 5 mol %) and phorbol 12,13-dibutyrate (50 nM), a condition sufficient to promote near maximal phorbol ester binding. In addition to their different responses to the various activators, the three protein kinase C isozymes also have different Km values for protein substrates. The type I enzyme appeared to have lower Km values for histone IIIS, myelin basic protein, poly(lysine, serine) (3:1) polymer, and protamine than those for the type II and III enzymes. These results documented that the three protein kinase C isozymes were distinguishable in their biochemical properties. In particular, the type I enzyme, which is a brain-specific isozyme, is distinct from the type II and III enzymes, both have a widespread distribution among different tissues.
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PMID:Biochemical characterization of rat brain protein kinase C isozymes. 317 May 68

Cultured ovine oligodendrocytes (OLGs) express a number of voltage-dependent potassium currents after they attach to a substratum and as they begin to develop processes. At 24-48 hours following plating, an outward potassium current can be identified that represents a composite response of a rapidly inactivating component and a steady-state or noninactivating component. After 4-7 days in culture, OLGs also develop an inward rectifier current. We studied the effects of forskolin and phorbol 12-myristate 13-acetate (PMA) on OLG outward currents. These compounds are known to alter the myelinogenic metabolism of OLGs. PMA, an activator of protein kinase C (PK-C), has been shown to enhance myelin basic protein phosphorylation while forskolin acting on adenylate cyclase, and thereby increasing cAMP, inhibits it. Both forskolin and PMA increase the phosphorylation of 2'3'-cyclic nucleotide phosphodiesterase, an OLG/myelin protein. We found that forskolin decreased the steady-state outward current at 120 mV by 10% at 100 nM, and by 72% at 25 microM from a holding potential of -80 mV. The time course of inactivation of the peak currents was decreased, affecting both the fast and slow time constants. There was no significant change in the steady-state parameters of current activation and inactivation. The effect of forskolin was attenuated when the adenylate cyclase inhibitor adenosine (2 mM) was present in the intracellular/pipette filling solution. The results of PMA experiments were similar to those obtained with forskolin. Whereas the amplitude of the currents in the presence of PMA was reduced by 28% at 1.5 nM and 60% and 600 nM, the decay phase of the peak currents was less affected. The PMA effect could still be seen when the intracellular Ca2+ was reduced to less than or equal to 10 nM with 5 mM BAPTA, but was inhibited when the cells were pre-exposed to 50 microM psychosine, a PK-C inhibitor. It is postulated that the potassium currents in OLG can be physiologically modulated by two distinct second-messenger systems, perhaps converging at the level of a common phosphorylated enzyme or regulatory protein.
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PMID:Forskolin and phorbol esters decrease the same K+ conductance in cultured oligodendrocytes. 321 67


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