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)

Effects of K-252a, (8R*, 9S*, 11S*)-(-)-9-hydroxy-9-methoxycarbonyl-8-methyl-2,3,9,10-tetrahydro-8, 11-epoxy-1H,8H,11H-2,7b,11a-triazadibenzo[a,g]cycloocta [cde]trinden-1-one, purified from the culture broth of Nocardiopsis sp., on the activity of myosin light chain kinase were investigated. 1) K-252a (1 x 10(-5) M) affected three characteristic properties of chicken gizzard myosin-B, natural actomyosin, to a similar degree: the Ca2+-dependent activity of ATPase, superprecipitation, and the phosphorylation of the myosin light chain. 2) K-252a inhibited the activities of the purified myosin light chain kinase and a Ca2+-independent form of the enzyme which was constructed by cross-linking of myosin light chain kinase and calmodulin using glutaraldehyde. The degrees of inhibition by 3 x 10(-6) M K-252a were 69 and 48% of the control activities with the purified enzyme and the cross-linked complex, respectively. Chlorpromazine (3 x 10(-4) M), a calmodulin antagonist, inhibited the native enzyme, but not the cross-linked one. These results suggested that K-252a inhibited myosin light chain kinase by direct interaction with the enzyme, whereas chlorpromazine suppressed the enzyme activation by interacting with calmodulin. 3) The inhibition by K-252a of the cross-linked kinase was affected by the concentration of ATP, a phosphate donor. The concentration causing 50% inhibition was two orders magnitude lower in the presence of 100 microM ATP than in the presence of 2 mM ATP. 4) Kinetic analyses using [gama-32P]ATP indicated that the inhibitory mode of K-252a was competitive with respect to ATP (Ki = 20 nM). These results suggest that K-252a interacts at the ATP-binding domain of myosin light chain kinase. The direct action of the compound on the enzyme would explain the multivarious inhibition of myosin ATPase, of superprecipitation, and of the contractile response of smooth muscle.
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PMID:K-252a, a novel microbial product, inhibits smooth muscle myosin light chain kinase. 296 51

A calmodulin-independent kinase isolated from chicken intestinal brush border phosphorylates brush border myosin mainly at an apparently single threonine on its 20 kDa light chains. Phosphorylation to 1.9 mol phosphate/mol myosin activated the myosin actin-activated ATPase about 12-fold, to about 100 nmol/min per mg. Brush border myosin ATPase can thus be activated by phosphorylation either at threonine, by calmodulin-independent kinase, or at serine, by calmodulin-dependent myosin light chain kinase, as previously shown [(1987) FEBS Lett. 223, 262-266].
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PMID:Phosphorylation of brush border myosin at threonine on its 20 kDa light chains by a calmodulin-independent kinase activates its ATPase. 296 28

Previous work from our laboratory indicated that pancreatic islets contain myosin light chain kinase, a calcium- and calmodulin-activated enzyme. This enzyme catalyzes phosphorylation of myosin which, in tissues containing smooth muscle, is believed to permit the ATPase of myosin to be activated by actin. The current report shows that incubating islet cytosol with ATP under conditions that should permit phosphorylation of myosin markedly enhances islet myosin ATPase activity in the presence of actin. It has been suggested that contractile proteins power insulin granule movements in the beta cell. Phosphorylation of myosin may be one of the means of coupling stimuli to insulin secretion.
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PMID:Activation of pancreatic islet myosin ATPase by ATP and actin. 316 Mar 44

Myosin light chain kinase was partially purified from bovine adrenal medulla. A polypeptide of Mr 165,000 dalton was identified as kinase by using anti-gizzard myosin light chain kinase IgG on immunoreplica. Phosphorylation of medullary myosin was Ca2+- and calmodulin-dependent. The phosphorylated myosin was showed to enhance the actin-activated Mg2+-ATPase activity. In contrast, the myosin ATPase activity was dramatically decreased by dephosphorylation of myosin.
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PMID:Phosphorylation of myosin light chain and the actin-activated ATPase activity of adrenal medullary myosin. 316 Jun 91

Many non-muscle cells including chromaffin cells contain actin and myosin. The 20,000 dalton light chain subunits of myosin can be phosphorylated by a Ca2+/calmodulin-dependent enzyme, myosin light chain kinase. In tissues other than striated muscle, light chain phosphorylation is required for actin-induced myosin ATPase activity. The possibility that actin and myosin are involved in catecholamine secretion was investigated by determining whether increased phosphorylation in the presence of [gamma-32P]ATP of myosin light chain by myosin light chain kinase enhances secretion from digitonin-treated chromaffin cells. In the absence of exogenous myosin light chain kinase, 1 microM Ca2+ caused a 30-40% enhancement of the phosphorylation of a 20 kDa protein. This protein was identified on 2-dimensional gels as myosin light chain by its comigration with purified myosin light chain. Purified myosin light chain kinase (400 micrograms/ml) in the presence of calmodulin (10 microM) caused little or no enhancement of myosin light chain phosphorylation in the absence of Ca2+ in digitonin-treated cells. In the presence of 1 microM Ca2+, myosin light chain kinase (400 micrograms/ml) caused an approximately two-fold increase in myosin light chain phosphorylation in digitonin-treated cells in 5 min. The phosphorylation required permeabilization of the cells by digitonin and occurred within the cells rather than in the medium. Myosin light chain kinase-induced phosphorylation of myosin light chain was maximal at 1 microM Ca2+. Under identical conditions to those of the phosphorylation experiments, secretion was unaltered by myosin light chain kinase. The experiments indicate that the phosphorylation of myosin light chain by myosin light chain kinase is not a limiting factor in secretion in digitonin-treated chromaffin cells and suggest that the activation of myosin is not directly involved in secretion from the cells. The experiments also demonstrate the feasibility of investigation of effects of exogenously added proteins on secretion in digitonin-treated cells.
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PMID:Effects of purified myosin light chain kinase on myosin light chain phosphorylation and catecholamine secretion in digitonin-permeabilized chromaffin cells. 367 47

We have been using lysed cell models of teleost retinal cones to examine the mechanism of contraction in nonmuscle cells. We have previously reported that dark-adapted retinas can be lysed with the detergent Brij-58 to obtain cone motile models that undergo Ca++- and adenosine triphosphate (ATP)-dependent reactivated contraction. In this report we further dissect the roles of ATP and Ca++ in activation of contraction and force production by (a) characterizing the Ca++ and nucleotide requirements in more detail, (b) by analyzing the effects of inosine triphosphate (ITP) and the ATP analog ATP gamma S and (c) by testing effects of cyclic adenosine monophosphate (cAMP) on reactivated cone contraction. Exposing lysed cone models to differing free Ca++ concentrations produced reactivated contraction at rates proportional to the free Ca++ concentration between 3.16 X 10(-8) and 10(-6) M. A role for calmodulin (CaM) in this Ca++ regulation was suggested by the inhibition of reactivated contraction by the calmodulin inhibitors trifluoperazine and calmidazolium ( R24571 ). The results of analysis of nucleotide requirements in lysed cone models were consistent with those of smooth muscle studies suggesting a role for myosin phosphorylation in Ca++ regulation of contraction. ATP gamma S and ITP are particularly interesting in that ATP gamma S, on the one hand, can be used by kinases to phosphorylate proteins (e.g., myosin light chains) but resists cleavage by phosphatases or adenosine triphosphatases (ATPases), e.g., myosin ATPase. ITP, on the other hand, can be used by myosin ATPase but does not support Ca++/calmodulin mediated phosphorylation of myosin light chains by myosin light chain kinase. Thus, these nucleotides provide an opportunity to distinguish between the kinase and myosin ATPase requirements for ATP. When individual nucleotides were tested with cone motile models, the nucleotide requirement was highly specific for ATP; not only ITP and ATP gamma S, but also guanosine triphosphate, cytosine triphosphate, adenylyl-imidodiphosphate (AMPPNP) failed to support reactivated contraction when substituted for ATP throughout the incubation. However, if lysed cones were initially incubated with ATP gamma S and then subsequently incubated with ITP, the cones contracted to an extent that was comparable to that observed with ATP. As observed in skinned smooth muscle, adding cAMP to contraction medium strongly inhibited contraction in lysed cone models.
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PMID:Regulation of reactivated contraction in teleost retinal cone models by calcium and cyclic adenosine monophosphate. 632 28

Limited digestion of calmodulin (CaM)-dependent myosin light chain kinase from turkey gizzard with alpha-chymotrypsin in the presence of bound CaM generated an 80,000-dalton kinase fragment that was fully active in the absence of Ca2+. This kinase catalyzed specific Ca2+-independent phosphorylation of the 20,000-dalton light chain of myosin using isolated light chains, intact myosin, and actomyosin. Phosphorylation of myosin in the absence of Ca2+ allowed us to dissociate myosin phosphorylation from other potential Ca2+-dependent regulatory mechanisms, thus permitting an evaluation of the postulated central role of myosin phosphorylation in the regulation of smooth muscle contraction. Ca2+-independent myosin phosphorylation was found to cause loss of Ca2+ sensitivity of 1) actin-activated myosin ATPase activity in a crude actomyosin preparation, and 2) tension development in skinned smooth muscle fibers in the absence of Ca2+. Myosin phosphorylation is, therefore, the key event in actin activation of ATPase activity and initiation of contraction in skinned chicken gizzard fibers.
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PMID:Gizzard Ca2+-independent myosin light chain kinase: evidence in favor of the phosphorylation theory. 684 77

Protein phosphorylation may play a critical role in stimulus-coupled secretion of platelets. Some platelet proteins become phosphorylated on exposure to agents such as thrombin and collagen, and the smallest of these phosphoproteins (molecular weight 20,000), has been identified as a light chain of myosin. Phosphorylation of myosin light chain increases the activity of actin-activated myosin ATPase and the resultant contraction of the actomyosin presumably mediates the release reaction. Platelet myosin light chain kinase has been identified as a calcium-dependent enzyme requiring calmodulin for its activity. Calmodulin is a Ca2+-binding protein with a molecular weight of approximately 18,000 which seems to be involved in a wide variety of cellular processes. Although a growing body of evidence suggests that non-muscle actomyosin, such as that isolated from platelets, is regulated by Ca2+-calmodulin-dependent light chain phosphorylation, the precise relationship between the phosphorylation and the function of platelets is not clearly established. We now present pharmacological evidence that a calmodulin-mediated system, such as Ca2+-dependent myosin light chain phosphorylation, also plays an important role in the phenomenon of the release reaction. N-(6-aminohexyl)-5-chloro-1-napthalene-sulphonamide (W-7) (refs 13-15) is shown to bind selectively to calmodulin in vitro and inhibit its biological activity.
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PMID:Ca2+-calmodulin-dependent phosphorylation and platelet secretion. 743 2

Smooth muscle hypertrophy is often found in tissue subjected to abnormal physical stress. To determine if physical stress (strain) per se could increase the contractile potential of airway smooth muscle (ASM), we compared cultured ASM cells subjected to strain to control cells (no strain) for rates of 1) myosin light chain kinase (MLCK)-mediated myosin light chain (LC20) phosphorylation, 2) actin-activated myosin ATPase, and 3) myosin light chain phosphatase-mediated myosin dephosphorylation. Lysates from strained cells showed increases in both LC20 phosphorylation activity and actomyosin ATPase activity but decreased rates of phosphatase-dependent myosin dephosphorylation. The increased LC20 phosphorylation activity and ATPase activity of the strained cells were accompanied by increases in cellular content of MLCK and myosin, respectively, compared with control. Because the cultured ASM cells exposed to strain expressed higher MLCK activity and actomyosin ATPase activity but lower myosin light chain phosphatase activity, these data suggest that physical stress in part determines ASM potential for contractile state.
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PMID:Mechanical strain increases contractile enzyme activity in cultured airway smooth muscle cells. 761 41

Polylysine (10-13 kDa) stimulates contraction in smooth muscle skinned fibers and activates actomyosin adenosinetriphosphatase (ATPase) activity in the absence of myosin light chain phosphorylation [P. T. Szymanski and R. J. Paul. Adv. Exp. Med. 304: 363-368, 1991; P. T. Szymanski, J. D. Strauss, G. Doerman, J. DiSalvo, and R. J. Paul. Am J. Physiol. 262 (Cell Physiol. 31): C1445-C1455, 1992]. To provide further information on the mechanism of polylysine action on contractility in smooth muscle, we investigated its effect on ATPase activity and conformation of purified gizzard myosin. We report here that polylysine directly stimulates myosin ATPase activity in a concentration-dependent manner. This stimulation could be completely abolished with the addition of heparin, a negatively charged heteropolysaccharide. Polylysine (10 microM) increases myosin ATPase activity to a level similar to that of myosin phosphorylation. Addition of 10 microM polylysine to phosphorylated myosin [with myosin light chain kinase and adenosine 5'-O-(3-thiotriphosphate) (ATP gamma S), to approximately 1.9 mol P/mol myosin], however, did not further stimulate ATPase activity. At 0.2 M KCl (the salt concentration at which myosin exists primary in the 10S form), the addition of polylysine increases myosin ATPase activity to a level comparable to that of untreated myosin in 0.3 M KCl. These changes parallel the increase in solution viscosity elicited by polylysine. These results suggest that polylysine induces a transition in myosin conformation from the 10S to the 6S form, and this was confirmed by electron microscopy.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Polylysine activates smooth muscle myosin ATPase activity via induction of a 10S to 6S transition. 836 68


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