Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.17 (CaMKII)
 4,029 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The 63-kDa subunit, but not the 60-kDa subunit, of brain calmodulin-dependent cyclic nucleotide phosphodiesterase was phosphorylated in vitro by the autophosphorylated form of Ca2+/calmodulin-dependent protein kinase II. When calmodulin was bound to the phosphodiesterase, 1.33 +/- 0.20 mol of phosphate was incorporated per mol of the 63-kDa subunit within 5 min with no significant effect on enzyme activity. Phosphorylation in the presence of low concentrations of calmodulin resulted in a phosphorylation stoichiometry of 2.11 +/- 0.21 and increased about 6-fold the concentration of calmodulin necessary for half-maximal activation of the phosphodiesterase. Peptide mapping analyses of complete tryptic digests of the 63-kDa subunit revealed two major (P1, P4) and two minor (P2, P3) 32P-peptides. Calmodulin-binding to the phosphodiesterase almost completely inhibited phosphorylation of P1 and P2 with reduced phosphorylation rates of P3 and P4, suggesting the affinity change of the enzyme for calmodulin may be caused by phosphorylation of P1 and/or P2. When Ca2+/calmodulin-dependent protein kinase II was added without prior autophosphorylation, there was no phosphorylation of the 63-kDa phosphodiesterase subunit or of the kinase itself in the presence of a low concentration of calmodulin, and with excess calmodulin the phosphodiesterase subunit was phosphorylated only at P3 and P4. Thus the 63-kDa subunit of phosphodiesterase has a regulatory phosphorylation site(s) that is phosphorylated by the autophosphorylated form of Ca2+/calmodulin-dependent protein kinase II and blocked by Ca2+/calmodulin binding to the subunit.
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PMID:Regulation of Ca2+/calmodulin-dependent cyclic nucleotide phosphodiesterase by the autophosphorylated form of Ca2+/calmodulin-dependent protein kinase II. 254 85

Felodipine, a dihydropyridine Ca2+ channel blocker, appears to have intracellular sites of action in addition to its ability to attenuate voltage-dependent Ca2+ channels in smooth muscle cells. In vitro, felodipine inhibits several calmodulin-dependent enzymes such as myosin light chain kinase, cyclic nucleotide phosphodiesterase and caldesmon kinase [Walsh MP, Sutherland C and Scott-Woo GC, Biochem Pharmacol 37: 1569-1580, 1988]. Such effects may partially explain the relaxant effects of felodipine and related dihydropyridines on vascular smooth muscle. We have examined the effects of felodipine on the activity of another important enzyme which has been implicated in the regulation of the contractile state of smooth muscle, protein kinase C. We chose to use a physiologically relevant substrate of protein kinase C for these studies, viz. platelet P47 protein, rather than the more commonly used lysine-rich histone which is probably not a physiologically important substrate. Protein kinase C and P47 were purified from human platelets and their important structural and functional properties were characterized. Felodipine and the p-chloro analogue of felodipine enhanced both the rate and extent of P47 phosphorylation by protein kinase C. Half-maximal activation was observed at 9.5 microM felodipine and 8.5 microM p-chloro analogue. Activation by felodipine was dependent upon the presence of phospholipid but did not require diacylglycerol. These observations suggest that the pharmacological actions of felodipine and related dihydropyridines may involve activation of protein kinase C in addition to their known effects on voltage-dependent Ca2+ channels and calmodulin-dependent enzymes.
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PMID:Activation of protein kinase C by the dihydropyridine calcium channel blocker, felodipine. 270 18

Chicken gizzard smooth muscle contains two major calmodulin-binding proteins: caldesmon (11.1 microM; Mr 141 000) and myosin light-chain kinase (4.6 microM; Mr 136 000), both of which are associated with the contractile apparatus. The amino acid composition of caldesmon is distinct from that of myosin light-chain kinase and is characterized by a very high glutamic acid content (25.5%), high contents of lysine (13.6%) and arginine (10.3%), and a low aromatic amino acid content (2.4%). Caldesmon lacked myosin light-chain kinase and phosphatase activities and did not compete with either myosin light-chain kinase or cyclic nucleotide phosphodiesterase (both calmodulin-dependent enzymes) for available calmodulin, suggesting that calmodulin may have distinct binding sites for caldesmon on the one hand and myosin light-chain kinase and cyclic nucleotide phosphodiesterase on the other. Consistent with the lack of effect of caldesmon on myosin phosphorylation, caldesmon did not affect the assembly or disassembly of myosin filaments in vitro. As previously shown [Ngai & Walsh (1984) J. Biol. Chem. 259, 13656-13659], caldesmon can be reversibly phosphorylated. The phosphorylation and dephosphorylation of caldesmon were further characterized and the Ca2+/calmodulin-dependent caldesmon kinase was purified; kinase activity correlated with a protein of subunit Mr 93 000. Caldesmon was not a substrate of myosin light-chain kinase or phosphorylase kinase, both calmodulin-activated protein kinases.
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PMID:Properties of caldesmon isolated from chicken gizzard. 299 32

Calmodulin, the ubiquitous and multifunctional Ca(2+)-binding protein, mediates many of the regulatory effects of Ca2+, including the contractile state of smooth muscle. The principal function of calmodulin in smooth muscle is to activate crossbridge cycling and the development of force in response to a [Ca2+]i transient via the activation of myosin light-chain kinase and phosphorylation of myosin. A distinct calmodulin-dependent kinase, Ca2+/calmodulin-dependent protein kinase II, has been implicated in modulation of smooth-muscle contraction. This kinase phosphorylates myosin light-chain kinase, resulting in an increase in the calmodulin concentration required for half-maximal activation of myosin light-chain kinase, and may account for desensitization of the contractile response to Ca2+. In addition, the thin filament-associated proteins, caldesmon and calponin, which inhibit the actin-activated MgATPase activity of smooth-muscle myosin (the cross-bridge cycling rate), appear to be regulated by calmodulin, either by the direct binding of Ca2+/calmodulin or indirectly by phosphorylation catalysed by Ca2+/calmodulin-dependent protein kinase II. Another level at which calmodulin can regulate smooth-muscle contraction involves proteins which control the movement of Ca2+ across the sarcolemmal and sarcoplasmic reticulum membranes and which are regulated by Ca2+/calmodulin, e.g. the sarcolemmal Ca2+ pump and the ryanodine receptor/Ca2+ release channel, and other proteins which indirectly regulate [Ca2+]i via cyclic nucleotide synthesis and breakdown, e.g. NO synthase and cyclic nucleotide phosphodiesterase. The interplay of such regulatory mechanisms provides the flexibility and adaptability required for the normal functioning of smooth-muscle tissues.
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PMID:Calmodulin and the regulation of smooth muscle contraction. 781 54

The present study was undertaken to determine kinetic and inhibition parameters and the mechanism of S-adenosyl-L-methionine:calmodulin-L-lysine N6-methyltransferase (EC 2.1.1.60, CLNMT), an enzyme for which calmodulin is a substrate. Partially purified CLNMT isolated from rat testes had a Vmax of 540 pmol/min/mg and Km values for mushroom demethylcalmodulin and S-adenosyl-L-methionine of 230 nM and 2.0 microM, respectively. Kinetic analysis indicated a complex Bi Bi sequential kinetic mechanism for CLNMT where S-adenosyl-L-methionine binds initially and is followed by demethylcalmodulin binding. When the effects of 20 different compounds that are either inhibitors of calmodulin-specific or methylation-specific functions were examined, CLNMT displayed a pattern of inhibition which differs from that seen with calmodulin-activated enzymes. The product of calmodulin methylation, fully trimethylated calmodulin, and nonmethylatable VU-3 calmodulin acted as competitive inhibitors of CLNMT, with Ki values of 310 and 400 nM, respectively. Of the 13 compounds tested, which are inhibitors of calmodulin-dependent cyclic nucleotide phosphodiesterase, only the calmodulin-binding domain from Ca2+/calmodulin-dependent kinase II, melittin, and calmidazolium were effective inhibitors of CLNMT and each exhibited a complex pattern of inhibition with Kis values of 21, 50, and 65 nM, respectively. The only potent methylation-specific inhibitor was S-adenosyl-L-homocysteine, which also displayed a complex pattern of inhibition.
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PMID:Calmodulin N-methyltransferase. Kinetics, mechanism, and inhibitors. 866 90

The activation of six target enzymes by calmodulin phosphorylated on Tyr99 (PCaM) and the binding affinities of their respective calmodulin binding domains were tested. The six enzymes were: myosin light chain kinase (MLCK), 3'-5'-cyclic nucleotide phosphodiesterase (PDE), plasma membrane (PM) Ca2+-ATPase, Ca2+-CaM dependent protein phosphatase 2B (calcineurin), neuronal nitric oxide synthase (NOS) and type II Ca2+-calmodulin dependent protein kinase (CaM kinase II). In general, tyrosine phosphorylation led to an increase in the activatory properties of calmodulin (CaM). For plasma membrane (PM) Ca2+-ATPase, PDE and CaM kinase II, the primary effect was a decrease in the concentration at which half maximal velocity was attained (Kact). In contrast, for calcineurin and NOS phosphorylation of CaM significantly increased the Vmax. For MLCK, however, neither Vmax nor Kact were affected by tyrosine phosphorylation. Direct determination by fluorescence techniques of the dissociation constants with synthetic peptides corresponding to the CaM-binding domain of the six analysed enzymes revealed that phosphorylation of Tyr99 on CaM generally increased its affinity for the peptides.
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PMID:Tyrosine phosphorylation modulates the interaction of calmodulin with its target proteins. 1041 41