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:3.6.1.3 (ATPase)
65,361 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The relaxant action of amiloride was investigated in the smooth muscles of guinea pig taenia ceci and chicken gizzard. Amiloride inhibited the contractions induced by high K+ (45.4 mM) and carbachol (10 microM) in the taenia with the concentrations needed to induce 50% inhibition (IC50) of approximately 41 microM. A prolonged incubation period (greater than 1 hr) was necessary to obtain the full inhibition of these contractions. The taenia gradually accumulated amiloride and the tissue/medium ratio exceeded 2.0 after a 120-min incubation period. Amiloride had no effect on the high K+-stimulated 45Ca++ uptake or the ATP content of the taenia. Amiloride inhibited the Ca++-induced contraction of the saponin-treated taenia with an IC50 of 186 microM. Amiloride (10-1000 microM) also inhibited superprecipitation and Mg++-adenosine triphosphatase activity of the gizzard native actomyosin as well as the phosphorylation of myosin light chain. The inhibition of the phosphorylation was antagonized competitively by ATP. Amiloride (1 mM) had no effect on the dephosphorylation of myosin light chain upon removal of Ca++ from reaction medium. Amiloride, at concentrations up to 1 mM, had not effect on calmodulin activity as monitored by the Ca++-calmodulin-activated erythrocyte membrane (Ca++ + Mg++)-adenosine triphosphatase and phosphodiesterase activities. In contrast to this, trifluoperazine inhibited the calmodulin activity at the concentration needed to inhibit the Ca++-induced contraction of the permeabilized taenia and the superprecipitation and the phosphorylation of myosin light chain of gizzard. We conclude that amiloride, unlike trifluoperazine, may inhibit directly the myosin light chain kinase activity to induce muscle relaxation.
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PMID:Inhibition by amiloride of contractile elements in smooth muscle of guinea pig taenia cecum and chicken gizzard. 282 5

The effect of phosphorylation by cyclic GMP-dependent protein kinase (G-kinase) on the activity of the plasmalemmal Ca2+-transport ATPase was studied on isolated plasma membranes and on the ATPase purified from pig erythrocytes and from the smooth muscle of pig stomach and pig aorta. Incubation with G-kinase resulted, in both smooth-muscle preparations, but not in the erythrocyte ATPase, in a higher Ca2+ affinity and in an increase in the maximal rate of Ca2+ uptake. Cyclic AMP-dependent protein kinase (A-kinase) did not exert such an effect. The stimulation of the (Ca2+ + Mg2+)-dependent ATPase activity of the purified Ca2+ pump reconstituted in liposomes depended on the phospholipid used for reconstitution. The stimulation of the (Ca2+ + Mg2+)-ATPase activity by G-kinase was only observed in the presence of phosphatidylinositol (PI). G-kinase, but not A-kinase, stimulated the phosphorylation of PI to phosphatidylinositol phosphate (PIP) in a preparation of (Ca2+ + Mg2+)-ATPase obtained by calmodulin affinity chromatography from smooth muscle, but not in a similar preparation from erythrocytes. Adenosine inhibited both the phosphorylation of PI and the stimulation of the (Ca2+ + Mg2+)-ATPase by G-kinase. In the absence of G-kinase the (Ca2+ + Mg2+)-ATPase was stimulated by the addition of PIP, but not by PI. In contrast with previous results of Furukawa & Nakamura [(1987) J. Biochem (Tokyo) 101, 287-290], no convincing evidence for a phosphorylation of the (Ca2+ + Mg2+)-ATPase was found. Evidence is presented showing that the apparent phosphorylation occurs in a contaminant protein, possibly myosin light-chain kinase. It is proposed that G-kinase stimulates the plasmalemmal Ca2+ pump of smooth-muscle cells indirectly via the phosphorylation of an associated PI kinase.
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PMID:Cyclic GMP-dependent protein kinase stimulates the plasmalemmal Ca2+ pump of smooth muscle via phosphorylation of phosphatidylinositol. 285 Aug 1

Nanomolar concentrations of synthetic peptides corresponding to the calmodulin-binding domain of skeletal muscle myosin light chain kinase were found to inhibit calmodulin activation of seven well-characterized calmodulin-dependent enzymes: brain 61 kDa cyclic nucleotide phosphodiesterase, brain adenylate cyclase, Bordetella pertussis adenylate cyclase, red blood cell membrane Ca++-pump ATPase, brain calmodulin-dependent protein phosphatase (calcineurin), skeletal muscle phosphorylase b kinase, and brain multifunctional Ca++ (calmodulin)-dependent protein kinase. Inhibition could be entirely overcome by the addition of excess calmodulin. Thus, the myosin light chain kinase peptides used in this study may be useful antagonists for studying calmodulin-dependent enzymes and processes.
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PMID:Synthetic peptides based on the calmodulin-binding domain of myosin light chain kinase inhibit activation of other calmodulin-dependent enzymes. 290 35

The effect of four slow Ca2+ channel blockers (felodipine, nifedipine, prenylamine and bepridil) that possess the ability to bind to calmodulin (CaM) section and to inhibit myosin light chain kinase (MLCK) on CaM-regulated Ca2+ pumping ATPase of cardiac sarcolemma (SL) and brain cyclic AMP phosphodiesterase (PDE) was studied. The ability of these drugs to inhibit Ca2+ pumping ATPase correlated with their inhibitory effect on CaM-activated Ca2+-dependent PDE. Nifedipine was unable to inhibit markedly both enzymes. Prenylamine also was a weak inhibitor, which was unexpected because of its CaM binding potency. Felodipine (10-50 microM) and bepridil (50 microM) markedly reduced activities of SL Ca2+ pumping ATPase and PDE. Striking differences were, however, demonstrated when Ca2+ and CaM concentrations, respectively, were increased. Previously it was reported that inhibition of the SL Ca2+ pumping ATPase by the CaM antagonist calmidazolium could be overcome by increasing Ca2+ concentrations (J. M. J. Lamers and J. T. Stinis, Cell Calcium 4, 281-294, 1983). Felodipine (10-50 microM) in the present study, appeared to be equipotent with calmidazolium in reducing Ca2+ pumping ATPase, but increasing Ca2+ up to 12.2 microM could not counteract this effect. Felodipine (2-10 microM) also inhibited brain PDE noncompetitively with respect to CaM contrary to the competitive effectors calmidazolium and bepridil. On the other hand, bepridil (10-20 microM) decreased or increased Ca2+ pumping ATPase activity depending on the Ca2+ concentration (0.29 and 12.2 microM, respectively) used. These findings suggest at least two types of CaM antagonists, which can be discriminated on basis of their inhibition patterns of PDE and heart SL Ca2+ pumping ATPase.
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PMID:Slow calcium channel blockers and calmodulin. Effect of felodipine, nifedipine, prenylamine and bepridil on cardiac sarcolemmal calcium pumping ATPase. 293 41

Thin-filament preparations from four smooth muscle types (gizzard, stomach, trachea, aorta) all activate myosin MgATPase activity, are regulated by Ca2+, and contain actin, tropomyosin and a 120000-140000-Mr protein in the molar proportions 1:1/7:1/26. The 120000-140000-Mr protein from all sources is a potent inhibitor of actomyosin ATPase activity. Peptide-mapping and immunological evidence is presented showing that it is identical with caldesmon. Quantitative immunological data suggest that caldesmon is a component of all the thin filaments and that the thin-filament-bound caldesmon accounts for all the caldesmon in intact tissue. The myosin light-chain kinase content of thin-filament preparations was found to be negligible. We propose that caldesmon-based thin-filament Ca2+ regulation is a physiological mechanism in all smooth muscles.
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PMID:Caldesmon is a Ca2+-regulatory component of native smooth-muscle thin filaments. 293 55

The crude actomyosin precipitate from sea urchin (Arbacia punctulata) egg extracts contains Ca2+-sensitive myosin light chain kinase activity. Activity can be further increased by exogenous calmodulin (CaM). Egg myosin light chain kinase activity is purified from total egg extract by fractionating on three different chromatographic columns: DEAE ion exchange, gel filtration on Sephacryl-300, and Affi-Gel-CaM affinity. The purified egg kinase depends totally on Ca2+ and CaM for activity. Unphosphorylated egg myosin has very little actin-activated ATPase. After phosphorylation of the phosphorylable light chain by either egg kinase or gizzard myosin light chain kinase, the actin-activated ATPase of egg myosin is enhanced several fold. However, the egg kinase bears some unique characteristics which are very different from conventional myosin light chain kinases of differentiated tissues. The purified egg kinase has a native molecular mass of 405 kDa, while on sodium dodecyl sulfate-polyacrylamide electrophoresis it shows a single subunit of 56 kDa. The affinity of egg kinase for CaM (Ka = 0.4 microM) is relatively weaker than that of the gizzard myosin light chain kinase. The egg kinase autophosphorylates in the presence of Ca2+ and CaM and has a rather broad substrate specificity. The possible relationship between this egg Ca2+-CaM-dependent kinase and the Ca2+-CaM-dependent kinases from brain and liver is discussed.
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PMID:Purification and characterization of a sea urchin egg Ca2+-calmodulin-dependent kinase with myosin light chain phosphorylating activity. 293 87

Cross-linked complex of gizzard myosin light chain kinase (MLCK) and calmodulin (CM) was produced by glutaraldehyde treatment of a mixture of these proteins in a high Ca2+ (0.1 mM) solution. Although the specific activity was reduced, this complex showed MLCK activity in a Ca2+-independent manner, different from the original MLCK whose activity was Ca2+-dependent. Chlorpromazine, one of the CM antagonists, was no longer able to inhibit the MLCK activity of this complex. These observations support the previously proposed hypothesis on the regulatory mechanism of MLCK activity via Ca2+. This complex could be regarded as another kind of Ca2+-independent MLCK different from that obtained by chymotryptic digestion of MLCK (Walsh, M.P., Dabrowska, R., Hinkins, S., & Hartshorne, D.J. (1982) Biochemistry 21, 1919-1925). This complex caused superprecipitation of gizzard actomyosin and enhanced actin-activated ATPase of myosin Ca2+-independently.
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PMID:Ca2+-independent gizzard myosin light chain kinase produced by cross-linking of the enzyme with calmodulin using glutaraldehyde. 294 Feb 35

We have previously isolated two Ca2+, calmodulin-dependent protein kinases with molecular weights of 120,000 (120K enzyme) and 640,000 (640K enzyme), respectively, by gel filtration analysis from rat brain. Chicken gizzard myosin light-chain kinase and the 120K enzyme phosphorylated two light chains of brain myosin, whereas the 640K enzyme phosphorylated both the two light chains and the heavy chain. The phosphopeptides of the light chains digested by Staphylococcus aureus V8 protease were similar among chicken gizzard myosin light-chain kinase, the 120K enzyme, and the 640K enzyme. Only the seryl residue in the light chains and the heavy chain was phosphorylated by the enzymes. The phosphorylation of brain myosin by any of these enzymes led to an increase in actin-activated Mg-ATPase activity. The results suggest that brain myosin is regulated by brain Ca2+, calmodulin-dependent protein kinases in a similar but distinct mechanism in comparison with that of smooth muscle myosin.
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PMID:Regulation of the actin-activated Mg-ATPase of brain myosin via phosphorylation by the brain Ca2+, calmodulin-dependent protein kinases. 294 Mar 39

Actomyosin in smooth muscle is in a quiescent state. The mechanism or mechanisms by which Ca2+ activates the actomyosin ATPase is not clear. There is sufficient evidence for the presence of enzyme systems which phosphorylate and dephosphorylate myosin light chains. The activity of the kinase that phosphorylates the myosin is regulated by cAMP-dependent protein kinase. Phosphorylated kinase has decreased affinity for calmodulin and lower activity when compared with unphosphorylated myosin light chain kinase. The activity of myosin light chain kinase is also regulated by calcium-calmodulin. In the presence of Ca2+, myosin is phosphorylated. In the absence of Ca2+, the phosphatase activity becomes dominant; the myosin remains in the unphosphorylated form under this condition. The Mg2+-ATPase of the phosphorylated myosin is activated by actin. The maximal activation of the Mg2+-ATPase by actin requires Ca2+ and tropomyosin, a protein located on the thin filament. Hence, the actin-activation of the Mg2+-ATPase requires Ca2+ even after phosphorylation by the calcium-calmodulin dependent kinase. The regulation of actin-activated ATPase activity by myosin light chain phosphorylation is depicted in the schematic diagram. Caldesmon, an actin-binding protein which also binds to calmodulin in the presence of Ca2+, has been shown to be present in thin-filaments isolated from smooth muscle. This protein inhibits actin-activated myosin ATPase activity. The release from this inhibition requires Ca2+ and calmodulin. The possibility that caldesmon is also involved in the calcium regulation of actomyosin in smooth muscle is presently under investigation in a number of laboratories.
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PMID:Regulation of actomyosin ATPase in smooth muscle. 294 44

Electron microscopy has been used to study the structural changes that occur in the myosin filaments of tarantula striated muscle when they are phosphorylated. Myosin filaments in muscle homogenates maintained in relaxing conditions (ATP, EGTA) are found to have nonphosphorylated regulatory light chains as shown by urea/glycerol gel electrophoresis and [32P]phosphate autoradiography. Negative staining reveals an ordered, helical arrangement of crossbridges in these filaments, in which the heads from axially neighboring myosin molecules appear to interact with each other. When the free Ca2+ concentration in a homogenate is raised to 10(-4) M, or when a Ca2+-insensitive myosin light chain kinase is added at low Ca2+ (10(-8) M), the regulatory light chains of myosin become rapidly phosphorylated. Phosphorylation is accompanied by potentiation of the actin activation of the myosin Mg-ATPase activity and by loss of order of the helical crossbridge arrangement characteristic of the relaxed filament. We suggest that in the relaxed state, when the regulatory light chains are not phosphorylated, the myosin heads are held down on the filament backbone by head-head interactions or by interactions of the heads with the filament backbone. Phosphorylation of the light chains may alter these interactions so that the crossbridges become more loosely associated with the filament backbone giving rise to the observed changes and facilitating crossbridge interaction with actin.
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PMID:Structural changes accompanying phosphorylation of tarantula muscle myosin filaments. 295 83


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