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)

We propose a mechanism for the cytoplasmic Ca++ oscillator which is thought to power shuttle streaming in strands of the slime-mold Physarum polycephalum. The mechanism uses a phosphorylation-dephosphorylation cycle of myosin light chain kinase. This kinase is bistable if the kinase phosphorylation chain, through adenylate cyclase and cAMP, is activated by calcium. Relaxation oscillations can then occur if calcium is exchanged between the cytoplasm and internal vacuoles known to exist in physarum. As contractile activity in physarum myosin is inhibited by calcium, this model can give calcium oscillations 180 degrees out of phase with actin filament tension as observed. Oscillations of ATP concentration are correctly predicted to be in phase with the tension, provided the actomyosin cycling rate is comparable with ATPase rates for phosphorylation of the myosin light chain and its kinase.
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PMID:Model of the Ca2+ oscillator for shuttle streaming in Physarum polycephalum. 153 35

The purpose of this study was to determine whether skeletal muscle mass, myofibrillar adenosinetriphosphatase activity, and the expression of myosin heavy (MHC) and light chain subunits are differentially affected in juvenile (4 wk) and young adult (12 wk) rats by a hypertrophic growth stimulus. Hypertrophy of the plantaris or soleus was studied 4 wk after ablation of either two [gastrocnemius (GTN) and soleus or plantaris] or one (GTN) synergistic muscle(s). There was no difference in the relative magnitude of hypertrophy because of age. Plantaris myofibrillar adenosinetriphosphatase activity was decreased 21 and 12% in juvenile and adult rats, respectively, as a result of ablation of both the GTN and soleus. Slow myosin light chain isoforms (1s and 2s) were expressed to a greater extent in hypertrophied plantaris muscles of both ages, but the increase in 1s was greater in juvenile rats. The relative expression of slow beta-MHC in hypertrophied plantaris muscles increased by 470 and 350%, whereas MHC IIb decreased by 70 and 33% in juvenile and adult rats, respectively. The relative expression of MHC IIa increased (56%) in the plantaris after ablation in juvenile rats only. These shifts in myosin subunit expression and the increases in mass were generally about one-half the magnitude when only the GTN was removed. There were no detectable myosin shifts in hypertrophied soleus muscles. Although the extent of muscle hypertrophy is similar, the shifts in myosin subunits were greater in juvenile than in young adult rats.
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PMID:Age effects on myosin subunit and biochemical alterations with skeletal muscle hypertrophy. 153 98

A peptide inhibitor, myosin kinase inhibitor (MKI), of myosin light chain kinase (MLCK) was tested for its effects on contractility and myosin light chain phosphorylation in Triton X-100 skinned guinea pig taenia coli. MKI is based on the amino acid sequence of the myosin light chain (residues 11-19 LC20) and is a competitive inhibitor [inhibitory constant (Ki) congruent to 10 microM] of purified MLCK with respect to myosin light chain (LC20). MKI inhibited unloaded shortening velocity (V(us)) and the calcium-sensitive ATPase activity of the skinned fibers but had no significant effect on steady-state isometric force or myosin light chain phosphorylation, as measured by IEF-polyacrylamide gel electrophoresis analysis. MKI had no significant effect on V(us) of thiophosphorylated fibers in the absence of calcium. MKI inhibited MLCK activity in protein extracts from taenia coli, as measured by radioactive phosphate incorporation into LC20. Surprisingly, MKI also inhibited the phosphatase activity of these same extracts. This peptide slowed the rate and extent of relaxation of calcium-contracted fibers and elicited a contraction in relaxed fibers. These results are consistent with the hypothesis that MKI may be a phosphatase inhibitor as well as an inhibitor of MLCK. Our data further suggest that the rate of phosphorylation-dephosphorylation turnover may be important in regulating V(us) in smooth muscle.
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PMID:Effects of myosin kinase inhibiting peptide on contractility and LC20 phosphorylation in skinned smooth muscle. 153 80

Phosphorylation/dephosphorylation of the 20-kDa light chain of smooth muscle myosin is a major regulator of actin-myosin interaction. Phosphatase inhibitors have thus been shown to enhance contraction in smooth muscle. The activity of type II phosphatase against phosphorylated myosin light chains is inhibited by polylysine. Thus we studied the effects of polylysine (10-13 kDa) on actin-myosin interaction in permeabilized guinea pig taenia coli fibers and in bovine aortic actomyosin. Addition of polylysine (10-20 microM) to Ca-ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid buffered solution ([Ca2+] less than 0.01 microM) elicited a contraction in fibers of 40 +/- 8% (n = 6) of maximally stimulated contractions ([Ca2+] congruent to 1.5 microM). Untreated fibers did not generate any significant force in parallel control experiments. Similarly, polylysine stimulated the ATPase activity both in fibers and actomyosin in a dose-dependent manner. This stimulation could be completely inhibited and abolished upon addition of heparin, a negatively charged heteropolysaccharide. In actomyosin previously phosphorylated with ATP gamma S, polylysine in a concentration range of 2-13 microM did not further stimulate enzyme activity. These increases in activity were not connected with significant changes in the phosphorylation of 20-kDa myosin light chain nor could any incorporation of 32P associated with polylysine stimulation be detected in both skinned fibers and actomyosin by autoradiography of SDS gels. Our data indicate that polylysine increases actin-myosin interaction in both smooth muscle model systems by directly influencing contractile proteins. As such, polylysine may be a useful probe for the mechanism of activation of smooth muscle.
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PMID:Polylysine activates smooth muscle actin-myosin interaction without LC20 phosphorylation. 153 81

Ca(2+)-calmodulin-dependent phosphorylation of the 20-kDa smooth muscle myosin light chain (MLC) results in high shortening velocities and rapid stress development. The stress maintained after a reduction in Ca2+ is associated with a decrease in MLC phosphorylation and velocity of shortening. This Ca(2+)-dependent stress without proportional MLC phosphorylation has been termed "latch" and has been postulated to reflect a population of dephosphorylated noncycling cross bridges or "latch bridges." Mg2+ is necessary for contraction of smooth muscle, and in high concentrations, Mg2+ elicits contractions that are MLC phosphorylation independent. The purpose of this study was to test the hypothesis that high concentrations of Mg2+ directly induce latch-bridge formation. This was accomplished by comparing the characteristics of Mg(2+)-induced contractions of Triton X-100-skinned swine carotid media with the known characteristics of the Ca(2+)-dependent latch state. In the absence of Ca2+, free Mg2+ (3-20 mM) caused an increase in the velocity of shortening and a concentration-dependent increase in stress, with no detectable increase in MLC phosphorylation. Mg(2+)-induced contractions could be supported by CTP, which is a substrate for the actin-activated myosin adenosinetriphosphatase but not the MLC kinase. Stress development in response to Mg2+ was abolished at long tissue lengths, which also inhibit the expression of latch bridges. The calmodulin antagonist, trifluoperazine (TFP), inhibited the MLC phosphorylation-independent contractions elicited by Mg2+. TFP also inhibited the latch state. The results of this study support the existence of a regulatory system in vascular smooth muscle that is independent of the MLC phosphorylation system and can be directly activated by pharmacological levels of Mg2+.
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PMID:Characterization of magnesium-induced contractions in detergent-skinned swine carotid media. 182 25

Human platelet myosin forms 10S and 6S conformations, and its Ca(2+)- and Mg(2+)-ATPase activities are parallel with the transition between 10S and 6S conformation, as judged by the gel filtration, intrinsic fluorescence, and viscosity methods. The 20,000-dalton myosin light chain (LC20) is phosphorylated by both myosin light chain kinase (MLC kinase) and Ca2+, phospholipid-dependent protein kinase (protein kinase C [PKC]). The phosphorylation (1 mol of phosphate/mol of LC20) by MLC kinase shifts the equilibrium toward the 6S conformation, but that by PKC does not. The prephosphorylation of myosin by PKC prevents the effect of phosphorylation by MLC kinase on actin-activated Mg(2+)-ATPase activity, but not the effect on conformational change. Inhibition of actin-activated ATPase activity by PKC is due to a decreased affinity of myosin for actin, and no change in Vmax is observed. These results suggest that sequential phosphorylation of myosin by both kinases plays an important role in the ATPase activities of human platelet myosin.
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PMID:Effect of phosphorylation of myosin light chain by myosin light chain kinase and protein kinase C on conformational change and ATPase activities of human platelet myosin. 183 91

Permeabilized endothelial cell monolayers retracted on exposure to ATP and Ca2+. ADP, inosine triphosphate (ITP), GTP, adenosine 5'-(gamma-thio)triphosphate (ATP-gamma S), and 5'-adenylylimidodiphosphate failed to support retraction. However, ATP gamma S, a substrate for myosin light-chain kinase (MLCK) but not myosin adenosinetriphosphatase (ATPase), combined with ITP, a substrate for myosin ATPase but not MLCK, supported retraction. Two MLCK pseudosubstrate peptides, M5 and SM-1, inhibited endothelial cell retraction equally and more effectively than myosin kinase-inhibitory peptide with a sequence based on the phosphorylated site of myosin light chain. M5 was shown to inhibit thiophosphorylation of endothelial cell myosin light chains. Endothelial cells incubated with exogenous unregulated kinase in the presence of ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetra-acetic acid retracted on addition of ATP. This retraction was accompanied by thiophosphorylation of the 19 kDa myosin light chains in the presence of ATP gamma 35S. The N-ethylmaleimide-modified subfragment 1 of myosin heads, a specific inhibitor of actin-myosin interaction, prevented retraction. These data add support to the proposal of a central role for MLCK activation of myosin in endothelial retraction.
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PMID:Regulation of permeabilized endothelial cell retraction by myosin phosphorylation. 185 58

Vasopressin (VP) and other hormones that elevate intracellular Ca2+ concentration also increase tight junctional permeability in the liver cell. Data derived from study of other tissues suggest that microfilaments are instrumental in regulating tight junctional permeability. By analogy to microfilament contraction in smooth muscles, it is likely that the transduction pathway for these hormones involves Ca(2+)-stimulated complex formation between calmodulin and myosin light chain (MLC) kinase with activation of this latter enzyme. MLC kinase then phosphorylates MLC, which, in the presence of actin, exerts adenosinetriphosphatase activity and produces microfilament contraction. This transduction pathway in the hepatocyte remains speculative. To demonstrate the likelihood of this pathway, we stimulated isolated hepatocytes with 10(-8) M VP and assayed MLC phosphorylation. We did this by immunoprecipitation of myosin from homogenates of liver cells prelabeled with [32P]-orthophosphate. We used a polyclonal antibody raised in rabbits against rat liver cell myosin. Our data demonstrate that VP is a potent stimulator of MLC phosphorylation. Maximal rises in intracellular Ca2+ and maximal MLC phosphorylation occur within 40 s of VP administration. The dose-response curve for MLC phosphorylation by VP is similar to that for tight junctional permeabilization in perfused liver with maximal effect at about 10(-8) M VP. The calcium ionophore A23187 also stimulated MLC phosphorylation. MLC phosphorylation, therefore, is at least coincident with, and probably responsible for, tight junctional permeabilization caused by elevation of intracellular Ca2+ in the liver cell.
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PMID:Vasopressin and A23187 stimulate phosphorylation of myosin light chain-1 in isolated rat hepatocytes. 187

Myosin light chain phosphorylation in permeable skeletal muscle fibers increases isometric force and the rate of force production at submaximal levels of calcium activation; myosin light chain phosphorylation may underlie the increased rate and extent of force production associated with isometric twitch potentiation in intact fibers. To understand the mechanism by which myosin light chain phosphorylation manifests these effects, we have measured isometric force, isometric stiffness, rate of isometric force redevelopment after isotonic shortening, and isometric ATPase activity in permeabilized rabbit psoas muscle fibers. These measurements were made in the presence and absence of myosin light chain phosphorylation over a range of calcium concentrations that caused various levels of activation. The results were analyzed with a two-state cross-bridge cycle model as suggested by Brenner [Brenner, B. (1988) Proc. Natl. Acad. Sci. USA 85, 3265-3269]. The results indicate that myosin light chain phosphorylation exerts its effect on force generation and the isometric rate of force redevelopment in striated muscle through a single mechanism, namely, by increasing the rate constant describing the transition from non-force-generating cross-bridges to force-generating states (fapp). gapp, the reverse rate constant, is unaffected by phosphorylation as are the number of cycling cross-bridges. Since both calcium and myosin light chain phosphorylation increase fapp, the possibility is considered that modulation of fapp may represent a general mechanism for regulating force in actin-myosin systems.
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PMID:Alteration of cross-bridge kinetics by myosin light chain phosphorylation in rabbit skeletal muscle: implications for regulation of actin-myosin interaction. 213 51

Isometric force developed by skinned gizzard muscle fiber bundles and levels of phosphorylation and thiophosphorylation of the 20,000-dalton myosin light chain were determined. These data showed a highly non-linear relationship between isometric force and myosin light-chain phosphorylation. Maximum force was developed at approximately 0.2 mol of phosphate/mol of light chain as reported previously (Hoar, P. E., Kerrick, W. G. L., and Cassidy, P. S. (1979) Science 204, 503-506). In contrast, the relationship between isometric force and myosin light-chain thiophosphorylation was linear, with maximum force occurring at 1.0 mol of thiophosphate/mol of myosin light chain. These observations are consistent with the latch-bridge hypothesis for conditions of varying myosin light-chain phosphatase/myosin light-chain kinase activity ratios as discussed by Hai and Murphy [1988) Am. J. Physiol. 254, C99-C106). To further test the latch-bridge hypothesis, ATPase activity was also measured during isometric force development in these fiber bundles. The relationship between isometric force and ATPase activity was linear whether the myosin light chains were phosphorylated or thiophosphorylated. Thus the number of cycling myosin cross-bridges, as measured by ATPase activity, was directly proportional to the force the muscle developed, not to the level of myosin light-chain phosphorylation. This finding that high levels of tension generated at low levels of light-chain phosphorylation are associated with high levels of ATPase activity is inconsistent with the latch-bridge model (Hai and Murphy, 1988).
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PMID:The relationship between ATPase activity, isometric force, and myosin light-chain phosphorylation and thiophosphorylation in skinned smooth muscle fiber bundles from chicken gizzard. 214 Mar 60


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