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)

We have characterized the structure and expression of rodent mRNAs encoding the fast and slow skeletal muscle isoforms of the contractile regulatory protein, troponin I (TnIfast and TnIslow). TnIfast and TnIslow cDNA clones were isolated from mouse and rat muscle cDNA clone libraries and were used as isoform-specific probes in Northern blot and in situ hybridization studies. These studies showed that the TnIfast and TnIslow mRNAs are expressed in skeletal muscle, but not cardiac muscle or other tissues, and that they are differentially expressed in individual muscle fibers. Fiber typing on the basis of in situ hybridization analysis of TnI isoform mRNA content showed an excellent correlation with fiber type as assessed by myosin ATPase histochemistry. These results directly demonstrate that the differential expression of skeletal muscle TnI isoforms in the various classes of vertebrate striated muscle cells is based on gene regulatory mechanisms which control the abundances of specific TnI mRNAs in individual muscle cells. Both TnIfast and TnIslow mRNAs are expressed, at comparable levels, in differentiated cultures of rat L6 and mouse C2 muscle cell lines. Thus, although neuronal input has been shown to be an important factor in determining fast versus slow isoform-specific expression in skeletal muscle, both TnIfast and TnIslow genes can be expressed in muscle cells in the absence of nerve. Comparison of the deduced rodent TnI amino acid sequences with previously determined rabbit protein sequences showed that residues with potential fast/slow isoform-specific function are present in several discrete clusters, two of which are located near previously identified actin and troponin C binding sites.
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PMID:cDNA clone and expression analysis of rodent fast and slow skeletal muscle troponin I mRNAs. 276 67

Myosin and actin competition tests indicated the presence of both thin-filament and myosin-linked Ca2+-regulatory systems in pig aorta and turkey gizzard smooth-muscle actomyosin. A thin-filament preparation was obtained from pig aortas. The thin filaments had no significant ATPase activity [1.1 +/- 2.6 nmol/mg per min (mean +/- S.D.)], but they activated skeletal-muscle myosin ATPase up to 25-fold [500 nmol/mg of myosin per min (mean +/- S.D.)] in the presence of 10(-4) M free Ca2+. At 10(-8) M-Ca2+ the thin filaments activated myosin ATPase activity only one-third as much. Thin-filament activation of myosin ATPase activity increased markedly in the range 10(-6)-10(-5) M-Ca2+ and was half maximal at 2.7 x 10(-6) M (pCa2+ 5.6). The skeletal myosin-aorta-thin-filament mixture gave a biphasic ATPase-rate-versus-ATP-concentration curve at 10(-8) M-Ca2+ similar to the curve obtained with skeletal-muscle thin filaments. Thin filaments bound up to 9.5 mumol of Ca2+/g in the presence of MgATP2-. In the range 0.06-27 microM-Ca2+ binding was hyperbolic with an estimated binding constant of (0.56 +/- 0.07) x 10(6) M-1 (mean +/- S.D.) and maximum binding of 8.0 +/- 0.8 mumol/g (mean +/- S.D.). Significantly less Ca2+ bound in the absence of ATP. The thin filaments contained actin, tropomyosin and several other unidentified proteins. 6 M-Urea/polyacrylamide-gel electrophoresis at pH 8.3 showed proteins that behaved like troponin I and troponin C. This was confirmed by forming interspecific complexes between radioactive skeletal-muscle troponin I and troponin C and the aorta thin-filament proteins. The thin filaments contained at least 1.4 mumol of a troponin C-like protein/g and at least 1.1 mumol of a troponin I-like protein/g.
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PMID:Calcium ion-regulated thin filaments from vascular smooth muscle. 644 98

Cardiac hypertrophy induced by thyrotoxic stress leads to an increase in the rate of force development, velocity of shortening, tension-dependent heat generation, and myosin ATPase activity. We did studies to see whether alterations in covalent phosphorylation of myofibrillar proteins correlate with these changes. The protein preparations were isolated from control and thyrotoxic hearts of male albino rabbits freeze-clamped in situ. We measured myofibrillar ATPase, and the covalent phosphate content of ventricular myosin 19,000 (mol wt) light chain (P-light chain) and troponin I (TnI). The myofibrillar ATPase activity was increased 2-fold in the thyrotoxic preparations with no change in the level of myofibrillar phosphorylation. The covalent phosphate content of TnI was 1.21 +/- 0.09 mol P/mol TnI in control hearts and 1.14 +/- 0.04 mol P/mol TnI in thyrotoxic hearts. The covalent phosphate content of the light chain fraction was 0.41 +/- 0.06 mol P/mol P-light chain in control hearts and 0.37 +/- 0.04 mol P/mol P-light chain in thyrotoxic hearts. The dependence of the normalized myofibrillar ATPase on free calcium concentration was the same in control and thyrotoxic preparations. Thus the mechanical, thermal, and biochemical changes found in hearts from thyrotoxic animals probably occur with no change in phosphorylation of TnI or myosin light chains.
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PMID:Phosphorylation and adenosine triphosphatase activity of myofibrils from thyrotoxic rabbit hearts. 645 Jun 50

In the newborn rat all cells of soleus, extensor digitorum longus (EDL), and tibialis anterior (TA) muscles stained for fast troponin I. A proportion of the cells, that was much higher in the soleus, also stained for slow troponin I. Fast and slow troponin I were segregated in different cell types in all three muscles 10 to 12 days after birth. No subsequent changes in the distribution of the two forms of troponin I occurred with further growth of EDL and TA muscles. The number of type I cells in soleus steadily increased with increasing age to 24 weeks. Three weeks after denervation at birth, almost all cells in soleus muscle stained for fast troponin I but less than 5% stained significantly dark for slow troponin I. All cells stained for myosin ATPase after alkaline preincubation, but very few after acid preincubation. Three weeks after denervation of EDL and to a lesser extent with TA muscle, fast and slow troponin I were still segregated in different cells. After alkaline preincubation all cells stained equally dark for myosin ATPase but only those positive for slow troponin I stained for myosin ATPase after acid preincubation.
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PMID:Effect of denervation at birth on the development of skeletal muscle cell types in the rat. 662 4

The Ca2+-binding component of troponin (TnC) and its proteolytic fragments containing Ca2+-binding sites I-III (TH1) or sites III and IV (TR2C) have been labeled with the fluorescent probes dansylaziridine (DANZ) at methionine 25 or 5-(iodoacetamidoethyl)amino-naphthalene-1-sulfonic acid (AEDANS) at cysteine-98. These probes report binding of Ca2+ to the low and high affinity sites, respectively. Fluorescence changes as a function of [Ca2+] were measured for the free peptides, their complexes with troponin I + troponin T, and these complexes bound to actin-tropomyosin in the presence of Mg2+ and ATP with and without myosin. An apparent Hill coefficient of 1.0-1.1 has been obtained for the Ca2+-induced fluorescence changes in TnC, its fragments, and their ternary complexes regardless of the label used. When a ternary complex containing appropriately labeled TnC or its fragment is bound to the actin-tropomyosin complex, the Hill coefficient for the titration of the low affinity sites increases to 1.5-1.6 and further increases to greater than 2 in the presence of myosin. To interpret the apparent Hill coefficients, we used a model containing two binding sites and a single reporter of the conformational change. Hill coefficients between 1.0 and 1.2 can be obtained for the fluorescence change without true cooperativity in metal binding, depending on the mechanism of the fluorescence change; i.e. the contribution of the singly or doubly occupied species to the fluorescence change. A Hill coefficient between 1.2 and 2, however, always indicates cooperativity in binding independently of the mechanism. Thus, our finding that fluorescence titrations of Ca2+ binding to TnCDANZ bound to actin-tropomyosin exhibit a Hill coefficient of 1.5 in the absence of myosin and 2.4 in its presence indicates the existence of true positive cooperativity in metal binding to sites I and II. No cooperativity was observed for AEDANS-labeled complexes that reflect Ca2+-binding to the high affinity sites. Plots of the Ca2+ dependence of myosin ATPase activity activated by actin-tropomyosin in the presence of any of the troponin complexes used had apparent Hill coefficients of approximately 4. The higher value suggests cooperative interactions in the activation of ATPase beyond those involved in Ca2+-binding to the Ca2+-specific sites.
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PMID:Cooperative binding to the Ca2+-specific sites of troponin C in regulated actin and actomyosin. 664 69

Adaptive cardiac hypertrophy in the rat has been characterized as pathological or physiological reflecting the nature of the inciting stimulus. These two adaptations are distinguished by alterations in contractility and in the myosin ATPase composition of the affected muscle. We investigated the relative amounts of the mRNAs encoding cardiac sarcoplasmic reticular calcium ATPase (SERCA2), cardiac and skeletal troponin I (TnI), atrial natriuretic factor (ANF), and myosin light chain 1 (MLC1) in the hearts of rats that had been subjected to either conditioning by swimming (Sw), to renovascular hypertension (H) or to the combined stimulus (H-Sw) for 6 weeks. Compared to control animals, the mRNA levels for SERCA2 and cardiac TnI were slightly increased with Sw and moderately depressed with H. H-Sw animals showed a trend towards normalized mRNA levels for both genes. ANF mRNA levels were slightly elevated with Sw and markedly elevated with both H and H-Sw. MLC1 mRNA levels did not change with either or both stimuli. These data confirm that these two types of adaptive hypertrophy can be distinguished at the level of gene expression and suggest that the mechanical alterations seen in adaptive hypertrophy reflect a spectrum of pre-translational alterations which are not limited to changes in myosin heavy chain gene expression.
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PMID:Alterations in gene expression in the rat heart after chronic pathological and physiological loads. 819 70

Diabetes is one of the most prevalent chronic conditions that has a high association with death from cardiovascular disease(s). An impaired cardiac function independent of vascular disease suggests the existence of a primary myocardial defect in diabetes mellitus. We and others have documented that myocardial performance is impaired in the hearts of chronically diabetic rats and rabbits. Abnormalities in the contractile proteins and regulatory proteins could be responsible for the mechanical defects in streptozotocin (STZ)-diabetic hearts. The major focus of research on contractile proteins in the diabetic state has been on myosin ATPase and its isoenzymes. However, in the contractile protein system, this could be only one of the mechanisms that might be a controlling factor in myofilament contraction in diabetes. To define the role of cardiac contractile as well as regulatory proteins (troponin-tropomyosin) as a whole in the regulation of actomyosin system in diabetic cardiomyopathy, individual proteins of the cardiac system were reconstituted under controlled conditions. Enzymatic data confirmed a diminished calcium sensitivity in the regulation of the cardiac actomyosin system when regulatory protein(s) complex was recombined from diabetic hearts. This diminished calcium sensitivity along with shifts in cardiac myosin heavy chain (V1-->V3) could contribute to the impaired cardiac function in the hearts of chronic diabetic rats. It has also been reported that sarcomeric proteins such as myosin light chain-2 (MLC-2) and troponin I (TnI) could be involved in regulating muscle contraction and in calcium sensitivity. Since phosphorylation of cardiac TnI is associated with altered maximum enzymatic activity and calcium force relationship in isolated muscle preparations. TnI phosphorylation could contribute to depressed myocardial contractility in experimental diabetes. While we have yet to understand the exact function of each component in cardiac muscle and their behavior in concert where all of them act in tandem, we have focussed on the role of contractile proteins and their regulation in diabetes in this review. We have also included a brief discussions on other relevant intracellular components. In summary, there is substantial evidence to suggest that there are independent processes associated with diabetes which effect cardiac performance in experimental animals and in man. The focus of this review has been the explication of a biochemical defect which underlies cardiac contractile dysfunction in experimental models of diabetes.
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PMID:Regulation of contractile proteins in diabetic heart. 921 70

Ca(2+) sensitizers may be advantageous for treatment in human heart failure by increasing cardiac force without increasing the Ca(2+) transient or energy consumption. To study the mode of action of the Ca(2+) sensitizers EMD 57033 (EMD) and CGP 48506 (CGP), their influence on butanedione monoxime (BDM)-mediated depression of cross-bridge cycling was analyzed in human myocardium (explanted hearts, dilated cardiomyopathy, n = 19). In Triton X (1%)-skinned fiber preparations of left ventricular myocardium from patients suffering from dilated cardiomyopathy, troponin I was extracted by vanadate (10 mM) treatment, resulting in a Ca(2+)-independent contraction. In troponin I-depleted fibers BDM (5-50 mM) was applied in the absence and presence of EMD (10 microM) or CGP (10 microM). To analyze the influence on cross-bridge kinetics, tension cost (ratio of ATPase activity and tension development) was studied. BDM exerted a dose-dependent force inhibition in troponin I-depleted fibers (IC(50) = 7.22 mM), which was antagonized by EMD (IC(50) of BDM + EMD = 19.97 mM) and CGP (IC(50) of BDM + CGP = 15.30 mM). EMD increased Ca(2+) sensitivity of force and maximal force in Triton X-skinned fibers. The Ca(2+)-sensitizing effect of CGP was accompanied by an increased Ca(2+) sensitivity of myosin-ATPase activity, an increased slope of the Ca(2+) force and Ca(2+) ATPase curve, as well as a reduced maximal myosin ATPase activity. CGP and EMD reduced tension cost. In conclusion, EMD and CGP antagonize the BDM-mediated relaxation in troponin I-depleted cardiac muscle fibers. The Ca(2+)-sensitizing effect of CGP seems to be dependent on an improvement of the myofilament cooperativity, whereas EMD seems to operate by increasing the force per cross-bridge.
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PMID:Different effect of the Ca(2+) sensitizers EMD 57033 and CGP 48506 on cross-bridge cycling in human myocardium. 1108 66

Cardiac troponin C is the Ca2+-dependent switch for heart muscle contraction. Troponin C is associated with various other proteins including troponin I and troponin T. The interaction between the subunits within the troponin complex is of critical importance in understanding contractility. Following a Ca2+ signal to begin contraction, the inhibitory region of troponin I comprising residues Thr128-Arg147 relocates from its binding surface on actin to troponin C, triggering movement of troponin-tropomyosin within the thin filament and thereby freeing actin-binding site(s) for interactions with the myosin ATPase of the thick filament to generate the power stroke. The structure of calcium-saturated cardiac troponin C (C-domain) in complex with the inhibitory region of troponin I was determined using multinuclear and multidimensional nuclear magnetic resonance spectroscopy. The structure of this complex reveals that the inhibitory region adopts a helical conformation spanning residues Leu134-Lys139, with a novel orientation between the E- and H-helices of troponin C, which is largely stabilized by electrostatic interactions. By using isotope labeling, we have studied the dynamics of the protein and peptide in the binary complex. The structure of this inhibited complex provides a framework for understanding into interactions within the troponin complex upon heart contraction.
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PMID:Structure and dynamics of the C-domain of human cardiac troponin C in complex with the inhibitory region of human cardiac troponin I. 1273 41

The complex of tropomyosin and troponin binds to actin and inhibits activation of myosin ATPase activity and force production of striated muscles at low free Ca(2+) concentrations. Ca(2+) stimulates ATP activity, and at subsaturating actin concentrations, the binding of NEM-modified S1 to actin-tropomyosin-troponin increases the rate of ATP hydrolysis even further. We show here that the Delta14 mutation of troponin T, associated with familial hypertrophic cardiomyopathy, results in an increase in ATPase rate like that seen with wild-type troponin in the presence of NEM-S1. The enhanced ATPase activity was not due to a decreased incorporation of mutant troponin T with troponin I and troponin C to form an active troponin complex. The activating effect was more prominent with a hybrid troponin (skeletal TnI, TnC, and cardiac TnT) than with all cardiac troponin. Thus it appears that changes in the troponin-troponin contacts that result from mutations or from forming hybrids stabilize a more active state of regulated actin. An analysis of the effect of the Delta14 mutation on the equilibrium binding of S1-ADP to actin was consistent with stabilization of an active state of actin. This change in activation may be important in the development of cardiac disease.
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PMID:The Delta 14 mutation of human cardiac troponin T enhances ATPase activity and alters the cooperative binding of S1-ADP to regulated actin. 1556 20


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