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)

The purification of axonal membranes of crustaceans was followed by measuring enrichment in [3H]tetrodotoxin binding capacity and in Na+, K+-ATPase activity. A characteristic of these membranes is their high content of lipids and their low content of protein as compared to other types of plasmatic membranes. The axonal membrane contains myosin-like, actin-like, tropomyosin-like, and tubulin-like proteins. It also contains Na+, K+-ATPase and acetylcholinesterase. The molecular weights of these two enzymes after solubilization are 280,000 and 270,000, respectively. The molecular weights of the catalytic subunits are 96,000 for ATPase and 71,000 for acetylcholinesterase. We confirmed the presence of a nicotine binding component in the axonal membrane of the lobster but we have been unable to find [3H]nicotine binding to crab axonal membranes. The binding to axonal membranes og of the sodium channel, has been studied in detail. The dissociation constant for the binding of [3H]tetrodotoxin to the axonal membrane receptor is 2.9 nM at pH 7.4. The concentration of the tetrodotoxin receptor in crustacean membranes is about 10 pmol/mg of membrane protein, 7 times less than the acetylcholinesterase, 30 times less than the Na+, K+-ATPase, and 30 times less than the nicotine binding component in the lobster membrane. A reasonable estimate indicates that approximately only one peptide chain in 1000 constitutes the tetrodotoxin binding part of the sodium channel in the axonal membrane. Veratridine, which acts selectively on the resting sodium permeability, binds to the phospholipid part of the axonal membrane. [3H]Veratridine binding to membranes parallels the electrophysiological effect. Veratridine and tetrodotoxin have different receptor sites. Although tetrodotoxin can repolarize the excitable membrane of a giant axon depolarized by veratridine, veratridine does not affect the binding of [3H]tetrodotoxin to purified axonal membranes. Similarly, tetrodotoxin does not affect the binding of [3H]veratridine to axonal membranes. Scorpion neurotoxin I, a presynaptic toxin which affects both the Na+ and the K+ channels, does not interfere with the binding of [3H]tetrodotoxin or [3H]veratridine to axonal membranes. Tetrodotoxin, veratridine, and scorpion neurotoxin I, which have in common the perturbation of the normal functioning of the sodium channel, act upon three different types of receptor sites.
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PMID:Constitution and properties of axonal membranes of crustacean nerves. 0 58

Fluorescein-labeled heavy meromyosin subfragment-1 (F-S-1) has been purified by ion exchange chromatography and characterized in terms of its ability to bind specifically to actin. F-S-1 activates the Mg++-adenosine triphosphatase activity of rabbit skeletal muscle actin and decorates actin as shown by negative stains and thin sections of rabbit actin and rat embryo cell microfilament bundles, respectively. Binding of F-S-1 to cellular structures is prevented by pyrophosphate and by competition with excess unlabeled S-1. The F-S-1 is used in light microscope studies to determine the distribution of actin-containing structures in wnterphase and mitotic rat embryo and rat kangaroo cells. Interphase cells display the familiar pattern of fluorescent stress fibers. Chromosome-to-pole fibers are fluorescent in mitotic cells. The glycerol extraction procedures employed provide an opportunity to examine cells prepared in an identical manner by light and electron microscopy. The latter technique reveals that actin-like microfilaments are identifiable in spindles of glycerinated cells before and after addition of S-1 or HMM. In some cases, microfilaments appear to be closely associated with spindle microtubles. Comparison of the light and electron microscope results aids in the evaluation of the fluorescent myosin fragment technique and provides further evidence for possible structural and functional roles of actin in the mitotic apparatus.
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PMID:Myosin subfragment binding for the localization of actin-like microfilaments in cultured cells. A light and electron microscope study. 7 3

Solution of thrombosthenin, the contractile protein complex isolated from pig platelets, have been studied by analytical ultracentrifugation and zone sedimentation in sucrose density gradients. Freshly prepared thrombosthenin in 0.6 M KCl shows a prominent peak in the ultracentrifuge with S degrees 20w about 5.5 and higher molecular weight aggregates (greater than 100S) sedimenting quickly to the bottom of the cell. Short term storage of high ionic strength solutions of thrombosthenin induces actomyosin-like gel formation and these gels dissociate with ATP and Mg2+ ions into two components of S degrees 20w 8.0 and S degrees 20w50. The supernatant, after actomyosin gel removal, contains only the S degrees 20w5.5 protein. From results of Ca2+ ATPase activity measurements and SDS polyacrylamide gel electrophoretic mobilities of dissociated thrombosthenin separated into fractions in sucrose density gradients, it is concluded that the S degrees20w5.5 protein species is the myosin-like protein of thrombosthenin. The S degrees 20w8.0 protein is not fibrinogen but also has myosin-like properties and is believed to be myosin dimer. Species of higher S values seen in the presence of ATP and Mg2+ in the analytical ultracentrifuge and located in the higher density zones of the sucrose gradients all gave in SDS polyacrylamide gel electrophoresis a single band of molecular weight 46-47,000 daltons. These subunit proteins appear to be derived from a range of polymeric variants of the F-actin-like protein of the contractile complex. All these higher density F-actin-like proteins readily form superprecipitates and display syneresis when combined with rabbit skeletal muscle myosin or platelet myosin. They are also all capable of conferring upon these two myosins a Mg2+ activated ATPase activity. It is suggested that in thrombosthenin solutions a myosin monomer-dimer equilibrium state exists which can be directionally influenced by a number of factors. The coexistence in the solution of F-actin and Mg2+ ATP, for example, increases the propensity of the myosin-like protein to form the higher molecular weight aggregate. Such aggregation may be the initiating mechanism for the intracellular organization of the thick filaments of the actomyosin complex, preparatory to a contractile event.
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PMID:Platelet contractile proteins: separation and characterization of the actin and myosin-like components. 12 96

A myosin-like protein was identified in isolated rabbit liver cells. It was extracted with high-ionic-strength buffer containing ATP, and purified by gel filtration in the presence of iodide. The myosin polypeptide was indistinguishable in size from the heavy chain of muscle myosin as determined by electrophoresis on polyacrylamide gels and gel filtration in the presence of sodium dodecyl sulfate. The hepatic myosin had an amino acid composition similar to that of muscle myosin, but lacked 3-methylhistidine. The Mg2+ -ATPase of the myosin was not activated by muscle actin. At low ionic strength, in the presence of Mg2+, the protein aggregated to form bipolar filaments 0.3 mum in length. A protein which resembled muscle actin in size and amino acid composition was extracted along with the myosin. Based on scans of stained sodium dodecyl sulfate polyacrylamide gels, the myosin content was estimated as 0.3% to 0.4% of the cell protein. The actin-like component was present in approximately ten-fold excess by weight. This ratio suggests that the organization and function of myosin in the hepatocyte is very different from that in the muscle cell.
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PMID:The identification of myosin in rabbit hepatocytes. 13 48

Differential ultracentrifugation of an extract of the plasmodium of Physarum polycephalum yields a high-speed fraction which exhibits calcium-sensitive adenosine triphosphate activity at low ionic strength. The rate of inorganic phosphate production increased from 2- to 25-fold in different preparations when the calcium concentration was increased from about 10(-8) to 10(-5) M. Complement fixation using specific antibody to Physarum myosin showed the fraction to contain 3% myosin. By electron microscopy, actin-like microfilaments 50--150 nm long were present. Addition of pure rabbit F-actin or myosin to this fraction activated the ATPase measured in EGTA and so partially reversed the calcium sensitivity. If muscle myosin was added to the supernatant from which the fraction was centrifuged, a "hybrid complex" was obtained which included actin and additional protein from the plasmodium, and this hybrid was also calcium sensitive. Over 85% of the calcium-sensitive, magnesium-activated ATPase could be precipitated by sequential "hybrid" formation. The calcium sensitivity of the hybrid was maximal when formed at the lowest ratios of added myosin to Physarum proteins. It is concluded that the results do not allow a simple interpretation along the lines of either actin-linked or myosin-linked sensitivity. Evidence consistent with both a form of actin-linked and myosin-linked sensitivity is present in our results.
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PMID:A calcium-sensitive preparation from Physarum polycephalum. 13 44

Lithium treatment is known to cause tubule dilation in distal nephron segments both in rat and in man. However, due to the heterogeneous cell composition of the distal nephron and the cellular changes following lithium treatment, it has been difficult to identify the structurally changed segments. In this study we have therefore applied computer-assisted reconstruction of cortical distal nephron segments. Tubule dilation was demonstrated in connecting and initial collecting tubules and in the first part of cortical collecting ducts (CCD) whereas it was absent from distal straight and distal convoluted tubules. Principal cells (P cells) in the CCD showed swelling of the cytoplasm, accumulation of actin-like microfilaments, and abnormal arrangements of basolateral membranes. Connecting tubule cells (CNT cells) showed similar but less pronounced changes. Intercalated cells (I cells) showed an accumulation of vesicles in the apical cytoplasm and a reduced luminal surface area. Lesions in P and CNT cells may, at least in part, explain the diabetes insipidus and sodium loss found during lithium treatment. Proton secretion in I cells is probably mediated by an ATPase present in the luminal membrane. The reduction in area of this membrane may explain why lithium-treated animals have a lowered ability to excrete an acid load.
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PMID:Lithium-induced structural changes in the cortical distal nephron localized by computer-assisted three-dimensional reconstruction. 324 74

In our opinion, all of the phenomena that are inhibited by cytochalasin can be thought of as resulting from contractile activity of cellular organelles. Smooth muscle contraction, clot retraction, beat of heart cells, and shortening of the tadpole tail are all cases in which no argument of substance for alternative causes can be offered. The morphogenetic processes in epithelia, contractile ring function during cytokinesis, migration of cells on a substratum, and streaming in plant cells can be explained most simply on the basis of contractility being the causal event in each process. The many similarities between the latter cases and the former ones in which contraction is certain argue for that conclusion. For instance, platelets probably contract, possess a microfilament network, and behave like undulating membrane organelles. Migrating cells possess undulating membranes and contain a similar network. It is very likely, therefore, that their network is also contractile. In all of the cases that have been examined so far, microfilaments of some type are observed in the cells; furthermore, those filaments are at points where contractility could cause the respective phenomenon. The correlations from the cytochalasin experiments greatly strengthen the case; microfilaments are present in control and "recovered" cells and respective biological phenomena take place in such cells; microfilaments are absent or altered in treated cells and the phenomena do not occur. The evidence seems overwhelming that microfilaments are the contractile machinery of nonmuscle cells. The argument is further strengthened if we reconsider the list of processes insensitive to cytochalasin (Table 2). Microtubules and their sidearms, plasma membrane, or synthetic machinery of cells are presumed to be responsible for such processes, and colchicine, membrane-active drugs, or inhibitors of protein synthesis are effective at inhibiting the respective phenomena. These chemical agents would not necessarily be expected to affect contractile apparatuses over short periods of time, they either do not or only secondarily interfere with the processes sensitive to cytochalasin (Table 1). It is particularly noteworthy in this context that microtubules are classed as being insensitive to cytochalasin and so are not considered as members of the "contractile microfilament" family. The overall conclusion is that a broad spectrum of cellular and developmental processes are caused by contractile apparatuses that have at least the common feature of being sensitive to cytochalasin. Schroeder's important insight (3) has, then, led to the use of cytochalasin as a diagnostic tool for such contracile activity: the prediction is that sensitivity to the drug implies presence of some type of contractile microfilament system. Only further work will define the limits of confidence to be placed upon such diagnoses. The basis of contraction in microfilament systems is still hypothetical. Contraction of glycerol-extracted cells in response to adenosine triphosphate (53), extraction of actin-like or actomyosin-like proteins from cells other than muscle cells (54), and identification of activity resembling that of the actomyosin-adenosine triphosphatase system in a variety of nonmuscle tissues (40, 54) are consistent with the idea that portions of the complex, striated muscle contractile system may be present in more primitive contractile machinery. In the case of the egg cortex, calcium-activated contractions can be inhibited by cytochalasin. If, as seems likely, microfilaments are the agents activated by calcium, then it will be clear that they have the same calcium requirement as muscle. Biochemical analyses of primitive contractile systems are difficult to interpret. Ishikawa's important observation (31), that heavy meromyosin complexes with fine filaments oriented parallel to the surface of chondrocytes and perpendicular to the surface of intestinal epithelial cells, implies that both types of filaments are "actin-like" in this one respect. Yet, it is very likely that these actin-like filaments correspond respectively to the cytochalasin-insensitive sheath of glial and heart fibroblasts and the core filaments of oviduct microvilli. No evidence from our studies links contractility directly to these meromyosin-binding filaments. Apart from this problem, activity resembling that of the myosin-adenosine triphosphatase has been associated with the microtubule systems of sperm tails and cilia (55), but those organelles are insensitive to cytochalasin in structure and function. Clearly, a means must be found to distinguish between enzymatic activities associated with microfilament networks, microfilament bundles, microtubules, and the sheath filaments of migratory cells. Until such distinctions are possible, little of substance can be said about the molecular bases of primitive contractile systems. Three variables are important for the control of cellular processes dependent upon microfilaments: (i) which cells of a population shall manufacture and assemble the filaments; (ii) where filaments shall be assembled in cells; and (iii) when contractility shall occur. With respect to distribution among cells, the networks involved in cell locomotion are presumed to be present in all cells that have the potential to move in cell culture. In this respect, the networks can be regarded as a common cellular organelle in the sense that cytoplasmic microtubules are so regarded. In some developing systems, all cells of an epithelium possess microfilament bundles (7, 13), whereas, in others, only discrete subpopulations possess the bundles (5, 6). In these cases the filaments can be regarded as being differentiation products associated only with certain cell types. These considerations may be related to the fact that microfilament networks are associated with behavior of individual cells (such as migration, wound healing, and cytokinesis), whereas the bundles are present in cells that participate in coordinated changes in shape of cell populations. With respect to placement in cells, two alternatives are apparent, namely, localized or ubiquitous association with the plasma membrane. Microfilament bundles of epithelial cells are only found extending across the luminal and basal ends of cells. In this respect they contrast with desmosomal tonofilaments and with microtubules, each of which can curve in a variety of directions through the cell. The strict localization of microfilament bundles probably rests upon their association with special junctional complex insertion regions that are only located near the ends of cells. In the case of mitotically active cells, the orientation of the spindle apparatus may determine the site at which the contractile ring of microfilaments will form (4, 56); this raises the question of what sorts of cytoplasmic factors can influence the process of association between filament systems and plasma membranes. In contrast to such cases of localized distribution, contractile networks responsible for cell locomotion are probably found beneath all of the plasma membrane, just as the network of thrombosthenin may extend to all portions of the periphery of a blood platelet. This ubiquitous distribution probably accounts for the ability of a fibroblast or glial cell to establish an undulating membrane at any point on its edge, or of an axon to form lateral microspikes along its length. The third crucial aspect of control of these contractile apparatuses involves the choice of when contraction shall occur (and as a corollary the degree or strength of contraction that will occur). In the simplest situation, contraction would follow automatically upon assembly of the microfilament bundles or networks. In cleavage furrows of marine embryos (4), for instance, microfilaments are seen beneath the central cleavage furrow and at its ends, but not beyond, under the portion of plasma membrane that will subsequently become part of the furrow. This implies that the furrow forms very soon after the contractile filaments are assembled in the egg cortex. In other cases, microfilaments are apparently assembled but not in a state of (maximal?) contraction. Thus, networks are seen along the sides of migratory cells, although such regions are not then active as undulating membrane organelles. Similarly, microfilament bundles occur in all epithelial cells of the salivary gland (13), or pancreatic anlage (7), although only the ones at discrete points are thought to generate morphogenetic tissue movements. Likewise, bundles begin to appear as early as 12 hours after estrogen administration to oviduct, although visible tubular gland formation does not start until 24 to 30 hours. Finally, streaming in plant cells can wax and wane, depending upon external factors such as auxin (57). All of these cases imply a control mechanism other than mere assembly of the microfilament systems and even raise the possibility that within one cell some filaments may be contracting while others are not. In discussing this problem, it must be emphasized that different degrees of contraction or relaxation cannot as yet be recognized with the electron microscope. In fact, every one of the cases cited above could be explained by contraction following immediately upon some subtle sort of "assembly." Inclusive in the latter term are relations between individual filaments, relations of the filaments and their insertion points on plasma membrane, and quantitative alterations in filament systems. Furthermore, the critical role of calcium and high-energy compounds in muscle contraction suggest that equivalent factors may be part of primitive, cytochalasinsensitive systems. The finding that calcium-induced contraction in the cortex of eggs is sensitive to cytochalasin strengthens that supposition and emphasizes the importance of compartmentalization of cofactors as a means of controlling microfilaments in cells.
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PMID:Microfilaments in cellular and developmental processes. 553 22

Using affinity chromatography on DNAase I-Sepharose, an actin-like protein was isolated from rat liver mitochondria and purified 60-fold. SDS electrophoresis in polyacrylamide gel revealed that the protein migrated with muscle actin and thus had the molecular weight of 42 000 Da. Evidence for the actin-like nature of the mitochondrial protein could be obtained from the fact that the protein inhibited the activity of pancreatic DNAase I which, similar to the smooth muscle protein, was less conspicuous than that of its muscle counterpart. Unlike striated muscle actin but similar to the smooth muscle protein, the mitochondrial actin weakly stimulated the Mg-ATPase activity of rabbit skeletal muscle myosin. After manyfold washing of the mitochondria with isotonic isolation media, the content of the actin-like protein remained unchanged, which indirectly points to the presence of insignificant cytoplasmic actin contaminations. During isoelectrofocusing, the mitochondrial actin-like protein yielded two forms, i. e., beta- and gamma-isoactins, whose ratio was 8:1. The pI values for the beta- and gamma-isoforms were 5.52 and 5.59, respectively. The identical position of the absorption spectra (260 nm) and fluorescence excitation spectra (around 280 nm) maxima of the actin-like protein and smooth and skeletal muscle actins testify to their homology.
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PMID:[Isolation and characteristics of actin-like proteins in liver mitochondria]. 624 Sep 91

In order to search for a common structural motif in the phosphate-binding sites of protein-mononucleotide complexes, we investigated the structural variety of phosphate-binding schemes by an all-against-all comparison of 491 binding sites found in the Protein Data Bank. We found four frequently occurring structural motifs composed of protein atoms interacting with phosphate groups, each of which appears in different protein superfamilies with different folds. The most frequently occurring motif, which we call the structural P-loop, is shared by 13 superfamilies and is characterized by a four-residue fragment, GXXX, interacting with a phosphate group through the backbone atoms. Various sequence motifs, including Walker's A motif or the P-loop, turn out to be a structural P-loop found in a few specific superfamilies. The other three motifs are found in pairs of superfamilies: protein kinase and glutathione synthetase ATPase domain like, actin-like ATPase domain and nucleotidyltransferase, and FMN-linked oxidoreductase and PRTase.
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PMID:Structural motif of phosphate-binding site common to various protein superfamilies: all-against-all structural comparison of protein-mononucleotide complexes. 1006 5

Plasmids encode partitioning genes (par) that are required for faithful plasmid segregation at cell division. Initially, par loci were identified on plasmids, but more recently they were also found on bacterial chromosomes. We present here a phylogenetic analysis of par loci from plasmids and chromosomes from prokaryotic organisms. All known plasmid-encoded par loci specify three components: a cis-acting centromere-like site and two trans-acting proteins that form a nucleoprotein complex at the centromere (i.e. the partition complex). The proteins are encoded by two genes in an operon that is autoregulated by the par-encoded proteins. In all cases, the upstream gene encodes an ATPase that is essential for partitioning. Recent cytological analyses indicate that the ATPases function as adaptors between a host-encoded component and the partition complex and thereby tether plasmids and chromosomal origin regions to specific subcellular sites (i.e. the poles or quarter-cell positions). Two types of partitioning ATPases are known: the Walker-type ATPases encoded by the par/sop gene family (type I partitioning loci) and the actin-like ATPase encoded by the par locus of plasmid R1 (type II partitioning locus). A phylogenetic analysis of the large family of Walker type of partitioning ATPases yielded a surprising pattern: most of the plasmid-encoded ATPases clustered into distinct subgroups. Surprisingly, however, the par loci encoding these distinct subgroups have different genetic organizations and thus divide the type I loci into types Ia and Ib. A second surprise was that almost all chromosome-encoded ATPases, including members from both Gram-negative and Gram-positive Bacteria and Archaea, clustered into one distinct subgroup. The phylogenetic tree is consistent with lateral gene transfer between Bacteria and Archaea. Using database mining with the ParM ATPase of plasmid R1, we identified a new par gene family from enteric bacteria. These type II loci, which encode ATPases of the actin type, have a genetic organization similar to that of type Ib loci.
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PMID:Plasmid and chromosome partitioning: surprises from phylogeny. 1093 39


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