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)

Aorta smooth muscle myosin contains two kinds of 17-kDa essential light chain, LC17nm (nonmuscle-type) and LC17gi (gizzard-type) [Hasegawa, Y., Ueda, Y., Watanabe, M., & Morita, F. (1992) J. Biochem. 111, 798-803]. The LC17 isoforms were released from porcine aorta myosin by incubation at 46 degrees C. The rate of release was 1.5 to 2 times higher with LC17gi than with LC17nm. Aorta myosins containing the two LC17 isoforms in various ratios could be reconstituted. The actin-activated ATPase activity was measured as a function of LC17nm content. The Vm value was lower with myosin which contained more LC17nm. The apparent dissociation constant for F-actin, Km, was 20-fold less with myosin which contained 81% LC17nm than myosin which contained 23% LC17nm. A similar difference in the dissociation constants of myosin for F-actin was observed in the presence of adenylyl imidodiphosphate. The role of LC17nm appears to be to make aorta myosin suitable for maintaining the muscle tension with a low expenditure of energy. The isoform-dependent difference in the F-actin-binding affinities of myosin seems partly due to the difference in the affinities of LC17 isoforms themselves for F-actin. We found that the isolated LC17nm itself could bind with F-actin with a dissociation constant of 64 microM, but LC17gi could not.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Role of 17-kDa essential light chain isoforms of aorta smooth muscle myosin. 138 50

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

KT5926, (8R*,9S*,11S*)-(-)-9-hydroxy-9-methoxycarbonyl-8-methyl-14-n-propoxy-2,3 ,9, 10-tetrahydro-8,11-epoxy, 1H,8H, 11H-2,7b,11a-triazadibenzo[a,g]cycloocta[cde] trinden-1-one, was found to be a potent and selective inhibitor of myosin light chain kinase. The compound inhibited both Ca2+/calmodulin-dependent and -independent smooth muscle myosin light chain kinases to a similar extent. The inhibition was not affected by the concentration of calmodulin. Kinetic analyses showed that the mode of inhibition was of the competitive type with respect to ATP (Ki, 18 nM) and of the noncompetitive type with respect to myosin light chain (Ki, 12 nM). These results indicated that KT5926 directly interacted with the enzyme at the catalytic site. KT5926 also inhibited other protein kinases, but with relatively high Ki values; the values for protein kinase C, cAMP-dependent protein kinase, and cGMP-dependent protein kinase were 723, 1200, and 158 nM, respectively. Ca2(+)-ATPase, Na+/K(+)-ATPase, hexokinase, and 5'-nucleotidase were not inhibited by KT5926 at less than 10 microM. The effect of KT5926 on serotonin secretion and protein phosphorylation induced by platelet-activating factor or phorbol ester was examined in rabbit platelets. KT5926 inhibited the phosphorylation of a 20-kDa protein but had no effect on the phosphorylation of a 40-kDa protein, thereby indicating that the compound exerts its selective inhibition of myosin light chain kinase in intact cells. The compound inhibited serotonin secretion induced by platelet-activating factor, but its potency was significantly less than that 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, which inhibited the phosphorylation of both the 20-kDa protein and the 40-kDa protein. Phorbol ester-induced secretion was not suppressed by KT5926. These results provide the evidence that both the 20-kDa protein phosphorylation by myosin light chain kinase and the 40-kDa protein phosphorylation by protein kinase C substantially contribute to the secretion response in platelets.
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PMID:KT5926, a potent and selective inhibitor of myosin light chain kinase. 232 35

The actin-activated ATPase activity of myosin II from Acanthamoeba castellanii is inhibited by phosphorylation of 3 serine residues near the carboxyl end of the heavy chain of the molecule. We have purified a protein phosphatase from Acanthamoeba using myosin II as a substrate. This phosphatase has a molecular weight of 39,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and an isoelectric point in urea of 5.2. The enzyme also is active against other phosphoserine protein substrates such as turkey gizzard smooth muscle myosin light chain, but not against a synthetic phosphotyrosine protein substrate. It does not hydrolyze ATP or p-nitrophenol phosphate. No effector has been found to increase substantially the activity of the enzyme as isolated, but it is inhibited by ATP, pyrophosphate, and NaF. This inhibition is reduced in the presence of MnCl2. The Mg2+-dependent actin-activated ATPase of myosin II is activated by dephosphorylation of phosphorylated myosin II by the phosphatase. Its broad substrate specificity, molecular weight, and response to protein phosphatase inhibitors suggest that the Acanthamoeba protein phosphatase is a type 2A phosphatase (Cohen, P. (1982) Nature (Lond.) 206, 613-620).
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PMID:Purification of a protein phosphatase from Acanthamoeba that dephosphorylates and activates myosin II. 631 29

Acanthamoeba myosin I heavy chain kinase activates the actin-activated Mg2+ -ATPase activity of the Acanthamoeba myosin I isoenzymes, myosins IA and IB, by phosphorylating a single site within the myosin heavy chain. In this paper, we report that myosin I heavy chain kinase also phosphorylates isolated turkey gizzard smooth muscle myosin light chains, gizzard smooth muscle heavy meromyosin, and intact gizzard smooth muscle myosin, all in the absence of Ca2+ and with specific activities close to those measured for purified Ca2+/calmodulin-dependent gizzard smooth muscle myosin light chain kinase. Myosin I heavy chain kinase incorporates a maximum of 2 mol of phosphate/mol of heavy meromyosin, both by itself and together with smooth muscle myosin light chain kinase (the light chain kinase alone incorporates 1.6 mol of phosphate/mol of heavy meromyosin). Both kinases phosphorylate intact smooth muscle myosin to a maximum of 2 mol of phosphate/mol of myosin. Myosin I heavy chain kinase fully activates the actin-activated Mg2+ -ATPase of both myosin and heavy meromyosin. Two-dimensional tryptic peptide maps of isolated light chains phosphorylated by myosin I kinase show the same phosphopeptide as for light chains phosphorylated by the light chain kinase. These results support the conclusion that myosin I heavy chain kinase phosphorylates gizzard smooth muscle myosin at the same site within the 20,000-Da light chain as does smooth muscle myosin light chain kinase. The results suggest that the amino acid sequence around the phosphorylation site within the heavy chain of Acanthamoeba myosin I isoenzymes may be similar to the primary sequence around the phosphorylation site within the smooth muscle myosin light chain.
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PMID:Phosphorylation and activation of smooth muscle myosin by Acanthamoeba myosin I heavy chain kinase. 632 1

Two anti-17,000 Da myosin light chain (LC17) monoclonal antibodies (MM2 and MM10), which increase the actin-activated Mg(2+)-ATPase activity of dephosphorylated smooth muscle myosin, inhibited the exchange of the 20,000 Da regulatory light chain of myosin (LC20). MM2, which shows higher potency of activation of ATPase activity, inhibited the exchange more extensively than MM10, suggesting that there is a correlation between the activation of ATPase activity and the inhibition of the LC20 exchange. The inhibition of the exchange was observed for intact myosin and heavy meromyosin but not subfragment 1, suggesting that the heavy chain at the head-rod junction is involved in the inhibition of LC20 exchange by anti-LC17 antibodies. Alternatively, the interaction between the two heads of the myosin molecule may influence the inhibition of LC20 exchange. These results suggest that LC20 interacts with both LC17 and the heavy chain, and the interaction between LC20 and LC17 is involved in the activation of actin-activated ATPase activity of smooth muscle myosin.
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PMID:Inhibition of 20-kDa myosin light chain exchange by monoclonal antibodies against 17-kDa myosin light chain. 772 54

Aorta smooth myosin contains two types of light chain, LC20 and LC17, which fold together with the N-terminal region of each heavy chain to form the globular head region of myosin. We demonstrate an altered conformation of LC20 after its separation from heavy chain by high concentrations of urea, on the basis of the following evidence: 1) A polyclonal antibody against LC20 was not able to recognize this conformationally altered form; 2) Myosin reconstituted from heavy chains and urea-dissociated light chains exhibited extremely low ATPase activity. Circular dichroism unfolding profiles showed that light chains dissociated from heavy chains by SDS appeared to be more stable than those generated by urea dissociation.
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PMID:Conformationally altered aortic myosin light chains. 784 64

The 20-kDa regulatory (LC20) and 17-kDa essential (LC17) light chain subunits could be removed from porcine aorta smooth muscle myosin by the use of trifluoperazine and ammonium chloride. The isolated heavy chain rebound both light chains, resulting in the restoration of native properties. Experiments on reconstitution of the isolated heavy chain with LC17 and/or LC20 showed that both light chains were required for folding into the 10 S conformation and thus for the phosphorylation-dependent filament formation of smooth muscle myosin. However, LC17 was not essential for the phosphorylation-dependent regulation of actin-activated ATPase activity and superprecipitation but was required for full regulation. LC17 and phosphorylated LC20 were found to act as activators, and dephosphorylated LC20 was found to act as a repressor of the motor activities of smooth muscle myosin.
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PMID:Roles of light chains in the activity and conformation of smooth muscle myosin. 862 39

Porcine aorta smooth muscle myosin contains two essential light chain (LC17) isoforms and the light chain was replaced with one of the LC17 isoforms, rabbit skeletal muscle myosin alkali light chain 2 (A2), or scallop striated muscle myosin essential light chain (SHLC). The myosin containing either an LC17 isoform or A2 showed phosphorylation-dependent properties in the monomer conformation, filament formation, ATPase activities, and superprecipitation, behaving in essentially the same way as native myosin. On the other hand, the replacement of LC17 with SHLC destabilized the 10S conformation and the myosin was predominantly filamentous under physiological conditions, irrespective of the phosphorylation state. This myosin containing dephosphorylated regulatory light chain showed higher actin-activated ATPase activity than native dephosphorylated myosin and was further activated by Ca2+, resulting in a decrease of phosphorylation-dependent regulation. Superprecipitation for the myosin was observed only when the regulatory light chain was phosphorylated. Superprecipitation for myosin containing SHLC was significantly slow in the absence of Ca2+ in comparison with that for myosin containing LC17, and was further activated by the presence of Ca2+. On the basis of the differences in amino acid sequences of these essential light chains, it appears that the N-terminal domain of LC17 may be implicated in these phosphorylation-dependent properties of smooth muscle myosin.
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PMID:Conformation and activity of smooth muscle myosin probed by various essential light chains. 905 92

The exchange of 17-kDa essential light chain (LC17) in smooth muscle myosin was carried out by incubating myosin with a 10-fold molar excess of exogenously added LC17 over the corresponding endogenous light chain in the presence of trifluoperazine and 4.5 m ammonium chloride. Porcine aorta myosin contains two kinds of LC17 isoform, LC17nm and LC17gi, while chicken gizzard myosin contains only one kind of LC17 isoform. As LC17gi can be separated from LC17nm and gizzard LC17 by urea-gel electrophoresis, LC17nm in aorta myosin and LC17 in gizzard myosin were exchanged with LC17gi and LC17gi in aorta myosin was exchanged with LC17nm, and the degree of exchange was estimated by urea-gel electrophoresis. Under the optimal conditions (6 and 10 degrees C for aorta and gizzard myosin, respectively), nearly 90% of exchange, which is close to the theoretical value, was achieved for the former combinations, and a slightly lower exchange was obtained for the latter. The LC17-exchanged myosins contained stoichiometric amounts of the heavy and light chains and retained the original nature in the phosphorylation-dependent actin-activated ATPase activity, 6S-10S conformational transition, and filament assembly.
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PMID:Essential light chain exchange in smooth muscle myosin. 935 45


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