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:3.6.4.1 (myosin ATPase)
1,140 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The flexibility of the tertiary structure around the active site of myosin ATPase [EC 3.6.1.3] was studied using the reactivity of two specific thiol groups, S1 and S2, as a structural probe. The following four maleimide derivatives were used as thiol-directed reagents: N-ethylmaleimide (NEM), N-(4-methoxy-2-benzimidazolyl methyl) maleimide (MBM), N-(p-(2-benzimidazolyl)phenyl)maleimide (BIPM) and N-(4-dimethyl-amino-3,5-dinitrophenyl)maleimide (DDPM). 1. All the maleimide derivatives used activated the Ca2+-ATPase activity and inhibited the EDTA-ATPase activity, like NEM, indicating that they modified S1. The rate of modification of S1 by NEM and BIPM increased with increasing pH, while that by DDPM decreased. BIPM simultaneously modified S1 and S2. 2. S1 showed much higher reactivity toward the maleimides, except for BIPM, than did N-acetylcysteine (N-Ac-Cys) a low molecular-weight model compound. The extremely small pKa value of S1, 6.28, accounted for this high reactivity. In addition, the ATP-induced increase in its reactivity inducated that S1 was in a buried state. Kinetic analysis showed that the teritiary structure around S1 at alkaline pH differed from that at acidic pH. 3. The apparent rate constant of S2-modification with NEM was approximately one seven-hundredth and one four-hundredth of those of S1 and N-Ac-Cys, respectively. Fluorimetric studies using BIPM revealed that S2 in the buried state was exposed upon adding ATP; this was compensated by the burying of some other thiol group(s) (Sp). Non-linearity of the Arrhenius plots of the reaction rate of S2 suggested that the S2 region of myosin had different conformations at high and low temperatures, the transition temperature being 10--15degrees. This non-linearity completely disappeared in the presence of Mg2+-ATP. On the other hand, Arrhenius plots for the thiols reactive to BIPM did not show non-linearity in the presence or absence of ATP.
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PMID:Thiols of myosin. IV. "Abnormal" reactivity of S1 thiol and the conformational changes around S2 thiol. 0 75

The rate of enzymic reaction of ATP, ITP, GTP with myosin is studied in the presence of potassiu, ammonium and calcium ions in H2O--D2O solutions. There is no kinetic isotope effect of ITPase and GTPase reaction in the neutral pH region (VHVD = 1). The value VH/VD for the ATPase reaction in the pH range from 6.5 to 8.5 with all cations studied varies from 1.05 to 1.26. Such changes of myosin enzymic activity in D2O infer that small changes in the interaction of subunits is not the decisive one in the regulation of myosin ATPase. The equality of isotope effects in potassium salts and ammonium solution suggests that a specific effect of ammonium ion as a proton donor affects the ATPase reaction of myosin. The relationship between the value of isotope effect and D2O concentration in solution in non-linear. The shape of concentration curve suggests essential conformational changes of myosin during ATP hydrolysis.
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PMID:[Enzyme activity of myosin activated by different cations in a mixed H2O--D2O solvent]. 3 22

While modification of six lysyl residues causes a near maximal decrease in Ca2+, K+, and actin + Mg2+ -activated myosin ATPase activities in rabbit skeletal muscle myosin, it takes nearly twice this number of modified lysyl groups to cause a similar alteration in canine cardiac myosin where trinitrophenylation is nonspecific. It appears that there are several rapidly reacting lysyl residues in cardiac myosin; the active site of cardiac myosin is protected by ATP after modification of a limited number of these rapidly reacting lysyl groups. In both myosins, after a charge modification of these rapidly reacting lysyl groups, 6 in rabbit skeletal muscle myosin and 10 in canine cardiac myosin, there is a decrease in Ca2+, K+, and actin + Mg2+ -stimulation of myosin but an activation of Mg2+ -stimulated myosin ATPase activity, thus making actin + Mg2+ -stimulated myosin ATPase activity more like activation with K+ or Ca2+ as compared to activation with Mg2+ alone.
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PMID:Comparison of Mg2+ vs Ca2+, K+ and actin-activation of myosin after trinitrophenylation. 4 Dec 96

A myosin was isolated from the clonal rat glial cell strain C-6 and compared with rat skeletal muscle myosin. After cell extracts were subjected to gel filtration chromatography in the presence of KI and magnesium pyrophosphate the C-6 myosin was rapidly purified by a procedure similar to that used for skeletal muscle myosin. The C-6 myosin resembles muscle myosin both physically and enzymatically. It contains heavy chains of 200,000 daltons and two classes of light chains of 17,000 and 19,000 daltons in approximately equal molar ratios. This myosin forms bipolar thick filaments in 0.1 M KCl and binds reversibly to skeletal muscle F-actin, the binding being inhibited by MgATP. Skeletal muscle F-actin stimulates the C-6 myosin adenosine triphosphatase 2- to 3-fold in the presence of KCl and Mg2+. The action activation of muscle myosin ATPase at low ionic strength is 10-fold greater than that of C-6 myosin. Ca2+ and EDTA stimulated the ATPase activities of both enzymes. When assayed in the presence of 0.6 M KCl and 1 mM EDTA the skeletal muscle myocin ATPase demonstrates substrate saturation while the C-6 myosin enzyme activity is stimulated by ATP concentrations above 2.5 mM.
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PMID:Purification and characterization of myosin from the clonal rat glial cell strain C-6. 12 31

The cyclic peptide phalloidin, one of the toxic components of Amanita phalloides prevented the drop of viscosity of F-actin solutions after the addition of 0.6 M KI and inhibited the ATP splitting of F-actin during sonic vibration. The data concerning ATP splitting are consistent with the assumption (a) that only 1 out of every 3 actin units of the filaments needs to be combined with phalloidin in order to suppress the contribution of these 3 actins to the ATPase activity of the filament and (b) that all actin units of the filaments can combine with phalloidin with a very high affinity. -halloidin did not only stabilize the actin-actin bonds in the F-actin structure but it also increased the rate of polymerization of G-actin to F-actin. The ability of F-actin to activate myosin ATPase was not affected by phalloidin. The tropomyosin-troponin complex did not prevent the stabilizing effect of phalloidin on the F-actin structure.
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PMID:Interaction of actin with phalloidin: polymerization and stabilization of F-actin. 12 84

The effects of D2O on the elementary steps in the contractile and transport ATPase [EC 3.6.1.3] reactions were studied, and the following results were obtained: 1. The rate of H-meromyosin ATPase in the steady state decreased in D2O to 60% of that in H2O. Deuterium oxide did not affect the size or rate of the initial burst of Pi liberation, i.e. the amount or rate of formation of the reactive myosin-phosphate-ADP complex, MADPP. Moreover, neither the rate of change in the fluorescence spectrum of H-meromyosin induced by ATP (the rate of formation of the second enzyme-ATP complex, M2ATP) nor the rate constant of decomposition of MADPP into M degrees + ADP + Pi was affected by D2O. However, the equilibrium constant of the step M2ATP in equilibrium MADPP decreased in D2O to about 1/2 the value in H2O. 2. In the case of the Na+-K+-dependent ATPase reactin, neither the rate constant of formation of the second enzyme-ATP complex, E2ATP, nor that of decomposition of a phosphorylated intermediate, EADP approximately P, was affected by D2O. However, the equilibrium constant of the step E2ATP in equilibrium EADP approximately P decreased in D2O to about 1/2.5-1/4 of the value in H2O. These results suggest a similarity between the modes of binding of phosphate in MADPP in the myosin ATPase reaction and in EADP approximatley P in the Na+-K+-dependent ATPase reaction.
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PMID:Effects of deuterium oxide on elementary steps in the ATPase reaction. Evidence for the similarity of key intermediates in contractile and transport ATPase. 13 92

The interaction between paramyosin and myosin has been studied by enzymological methods. Clam adductor paramyosin inhibits the actin-activated, Mg2+-requiring ATPase of both clam adductor and rabbit skeletal muscle myosins. Myosin and paramyosin must be rapidly coprecipitated for this inhibition. Incubation with F-actin in the absence of ATP does not alter this effect. This inhibition follows a hyperbolic function with respect to paramyosin concentration. Slow precipitation by dialysis of myosin and paramyosin together leads to copolymers with actin-activated ATPase equivalent to that of slowly formed myosin filaments. Both kinds of slowly formed filaments have enzymatic properties distinct from those of the rapidly precipitated proteins. Paramyosin is competitive with F-actin for their effects upon myosin. The apparent affinity of myosin for F-actin is markedly reduced by association with paramyosin, but the extrapolated maximal velocity of actomyosin is unaffected. The specificity of this inhibition is strongly suggested by marked quantitative differences between native and cleaved paramyosins. No inhibition of intrinsic myosin ATPase by paramyosin is seen. These studies suggest that at least two types of condition-dependent association between myosin and paramyosin are possible. One class of interactions is associated with enzymic inhibition in rapidly coprecipitated filaments, whereas slowly formed cofilaments exhibit catalytic activity similar to that of identically treat-d myosin and have a characteristic 14.5 nm axial repeat.
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PMID:Myosin-paramyosin cofilaments: enzymatic interactions with F-actin. 13 57

Important similarities are reported between human smooth muscle actomyosin and the human erythrocyte spectrin complex, primarily components 1, 2, and 5 (Fairbanks G., Steck, T.L., and Wallach, D.F.H. (1971), Biochemistry 10, 2606). The actin-like protein, component 5, is identical with human uterine actin in its ability to form 50-70-A filaments to stimulate myosin ATPase activity, and to bind rabbit heavy meromyoson specit heavy meromyosin specifically. Antibodies to human smooth muscle myosin(uterine) were prepared which were monospecific. A weak but specific cross-reaction of these antisera with components 1 and/or 2 (spectrin) was characterized and at least 25% of the antimyosin antibodies showed a low affinity reaction iwth spectrin. Antibodies generated against a soluble complex of spectrin components 1 and 2 reacted only with component 1 and did not cross-react with myosin. In addition to these structural similarities between smooth muscle actomyosin and the spectrin complex, we have found that spectrin is involved in ATP-dependent erythrocyte shape changes (Sheetz, M.P., Painter, R.G., AND Singer, S.J. (1976B), Cold Spring Harbor Symp. Cell Motility (in press) and, therefore, the spectrin complex is also a mechanochemical protein system.
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PMID:Relationships of the spectrin complex of human erythrocyte membranes to the actomyosins of muscle cells. 13 78

Bovine cardiac myosin ATPase activity was rapidly inactivated by the purine disulfide analog of ATP,6,6'-dithiobis(inosinyl imidodiphosphate). Kinetic investigations showed that this analog acted as a site-specific reagent at 0 degrees with a Ki of 130 muM and a half-life of 8.2 min at saturating inhibitor concentrations. Concentrations (50 to 500 muM) of ATP, adenyl-5'-yl imidodiphosphate (AMP-PNP), or ADP that saturated the active site caused an enhancement in the rate of inactivation, indicating the purine disulfide analog was not reacting at the active site. Under these conditions saturation kinetic data were still observed with Ki values remaining unchanged (120 muM) but with the half-life of inactivation decreasing to 6.0 min (ATP) and 4.6 min (AMP-PNP) at saturating inhibitor concentrations. At concentrations greater than 0.5 mM ATP, AMP-PNP, or ADP there was a decrease in the rate of inactivation, implying protection by these nucleotides. However, saturation kinetics of inactivation could no longer be demonstrated, implying a change in the mechanism of inactivation. A comparison of the inactivation of the Mg2+, Ca2+, and EDTA-ATPase activities of cardiac myosin after modification by the purine disulfide analog showed that the Mg2+- and Ca2+ATPase activities plateaued at approximately 60% and 40%, respectively, while the EDTA-ATPase activity continued to decrease to below 10%. This evidence supports the suggestion that the purine disulfide analog was not reacting at the active site. Equilibrium dialysis experiments were used to measure the binding of [8-3H]AMP-PNP to native cardiac myosin, the thiopurine nucleotide-modified myosin, and the derivative formed by displacing the thiopurine nucleotide by cyanide (thiocyanato-myosin). Native myosin bound a total of 2.1 mol of AMP-PNP with a binding constant of 6.0 X 10(6) M-1. There was a 15 to 40% decrease in the number of AMP-PNP binding sites in the enzyme derivatives, but the active sites appeared not to be blocked since the association constants remained essentially unchanged (KA=3.9 X 10(6) M-1 for thiopurine nucleotide-myosin and 12.0 X 10(6) M-1 for thiocyanato-myosin). The kinetic studies and the binding experiments indicate that the purine disulfide analog reacts at a specific site other than the active site but do not offer support to earlier suggestions from skeletal myosin studies that this site is a possible ATP control site.
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PMID:Reaction of cardiac myosin with a purine disulfide analog of adenosine triphosphate. I. Kinetics of inactivation and binding of adenylyl imidodiphosphate. 13 83

Kinetic measurement of the reaction of dynein ATPase (ATP phosphohydrolase, EC 3.6.1.3) extracted from the gills of Mytilus edulis shows that in the presence of Mg2+ there is a very rapid initial liberation of Pi from the dynein-ATP system, followed by a slower liberation in the steady state. In view of following results, we have confirmed that this phenomenon is not due to the accumulation of end products, a fall in substrate concentration, nor to the presence of labile impurities in ATP but is due to the catalytic activity of dynein ATPase. 1. The replacement of native dynein by heat denatured dynein or other kinds of Mg2+-ATPase could not produce such a burst phenomenon under the same condition. 2. Both the rate of initial burst and that of steady state were proportional to enzyme content over a wide range under our standard condition. 3. Initial burst was also observed under the constant ATP level by using a ATP generate system. 4. Preincubation of dynein with Pi prior to initiation of the reaction did not eliminate the initial burst. Some properties of the initial rapid liberation of dynein ATPase were also examined. These are shown below. 5. The free ADP liberation did not show any initial burst though the Pi liberation did in the initial phase and the rate of free ADP liberation was almost equal to that of Pi liberation of the steady state. 6. Mg2+ was more effective than Ca2+ for the appearance of the initial burst while the liberation of Pi in the steady state was activated more by Ca2+ than by Mg2+. The addition of K+ in the presence of Mg2+ resulted in a marked increase of Pi liberation in the steady state but not in the initial state. 7. The activation energy of the initial burst was 9.7 kcal, which is slightly smaller than that of myosin ATPase.
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PMID:Studies on the initial phase of dynein ATPase activity. 13 33


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