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)

Muscular dysgenesis is a lethal mutation in mice that results in a complete absence of skeletal muscle contraction due to the failure of depolarization of the transverse tubular membrane to trigger calcium release from the sarcoplasmic reticulum. In order to determine whether the defect in muscular dysgenesis leads to a specific loss of one of the components of excitation-contraction coupling or to a generalized loss of all components of excitation-contraction coupling, we have analyzed skeletal muscle from control and dysgenic mice for the sarcoplasmic reticulum and transverse tubular proteins which are believe to function in excitation-contraction coupling. We report that the proteins involved in sarcoplasmic reticulum calcium transport, storage, and release [Ca2+ + Mg2+)-ATPase, calsequestrin, and calcium release channel) are present in dysgenic muscle. Also present in dysgenic muscle is the 175/150-kDa glycoprotein subunit (alpha 2) of the dihydropyridine receptor. However, the 170-kDa dihydropyridine binding subunit (alpha 1) of the dihydropyridine receptor is absent in dysgenic muscle. These results suggest that the specific absence of the alpha 1 subunit of the dihydropyridine receptor is responsible for the defects in muscular dysgenesis and that the alpha 1 subunit of the dihydropyridine receptor is essential for excitation-contraction coupling in skeletal muscle.
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PMID:Specific absence of the alpha 1 subunit of the dihydropyridine receptor in mice with muscular dysgenesis. 253 62

Limited tryptic digestion of fluorescein isothiocyanate (FITC)-labeled (H+-K+)-ATPase from rat resting light gastric membranes produced a soluble 27-kDa polypeptide which retained the fluorescence of the parent enzyme. Its production was markedly enhanced in the presence of an amphiphilic detergent, Zwittergent 3-14, which potently inhibits the ATPase activity. This increase is probably due to protection of certain tryptic cleavage sites through conformational changes of the membrane enzyme by the detergent. The NH2-terminal sequence of the 27-kDa polypeptide corresponded exactly to that beginning at Asn-369 of the cDNA-deduced primary structure of the rat ATPase. The presence of the phosphorylation site, Asp-385, and FITC-labeled Lys-517, which is known to be a part of the ATP-binding site, indicates that the 27-kDa polypeptide contains a major cytoplasmic portion of (H+-K+)-ATPase. Interestingly, the polypeptide was stained with periodate-Schiff's base, indicating its glycoprotein nature. The carbohydrate group attached to the polypeptide seems to include at least an N-linked high-mannose moiety, since the polypeptide showed Con A binding activity as detected with a Con A-biotin/avidin-peroxidase assay on nitrocellulose transblots. Also, its Con A binding activity was inhibited by excess methyl alpha-D-mannopyranoside and disappeared upon treatment of the polypeptide with endoglycosidase H and N-glycanase. Further tryptic action converted the 27-kDa polypeptide to 2 smaller FITC-labeled polypeptides of 25 and 15 kDa, which lost 18 and 96 amino acid residues, respectively, from the NH2 terminus of the parent polypeptide.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Evidence for the presence of a carbohydrate moiety in fluorescein isothiocyanate labeled fragments of rat gastric (H+-K+)-ATPase. 254 51

A convenient method for highly efficient and directional immobilization of intact sodium- and potassium-activated ATPase (Na,K-ATPase) using wheat germ agglutinin linked on microtiter plates was developed. Wheat germ agglutinin, which bound tightly to the beta-subunit of Na,K-ATPase and had no effect on the Na,K-ATPase activity, the potassium-activated p-nitrophenylphosphatase activity, or the inhibitory action of ouabain, was covalently linked to microtiter plates and used as an immobilizer of the enzyme. The amount of Na,K-ATPase coupled to microtiter plates in this immobilizing system was more than 10-fold greater than that used in the direct immobilizing system (O. Urayama, M. Nakao, H. Nagamune, and H. Sugiyama, (1984) Anal. Biochem. 141, 194-198). Also in this system, the cytoplasmic domain of Na,K-ATPase was exposed to the liquid phase. This technique was useful for investigating the reactivities of monoclonal antibody specific for the cytoplasmic domain of the enzyme. Moreover, because this technique was used successfully in the immobilization of periodic acid--Schiff positive staining glycoprotein 1 prepared from human erythrocytes and human alpha 2-macroglobulin, the technique should also be useful for other membrane or secreted proteins that possess N-linked sugar chains containing bisecting N-acetylglucosamine or a high amount of sialic acid.
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PMID:Directional immobilization of sodium- and potassium-activated ATPase to expose its cytoplasmic part to the liquid phase on microtiter plates by wheat germ agglutinin. 255 55

This paper describes further characterization of the 170-180-kDa glycoprotein (P-glycoprotein) recognized by the monoclonal antibody MRK 16 in the human adrenal. By electron microscopy, P-glycoprotein was observed in the adrenal cell membranes. However, MRK 16-defined P-glycoprotein was not found in cow, pig, horse, monkey or rabbit adrenal, indicating that MRK 16 recognizes the non-homologous part of P-glycoprotein of various species. Eleven out of 16 adrenal tumors including 4 cases of primary aldosteronism and 7 cases of Cushing syndrome were intensely stained with MRK 16, whereas pheochromocytoma, non-functioning adrenocortical adenoma with no associated increase of serum adrenal-derived hormones and myolipoma of the adrenal were not. Finally, P-glycoprotein-MRK 16-protein A-Sepharose complex derived from human adrenal possessed marked ATPase activity. Taken together, these data suggest that P-glycoprotein may play a physiological role in the human adrenal.
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PMID:Further characterization of the human adrenal-derived P-glycoprotein recognized by monoclonal antibody MRK 16 reacting with only human P-glycoprotein. 257 26

In this paper, progress towards the goal of understanding communication between the nucleus and cytoplasm using an in vitro system is reviewed. To probe the mechanism of nuclear targeting, we developed an in vitro transport system and have begun to dissect the highly selective process of nuclear transport. The basic parameters of transport were defined using an easily isolated nuclear protein, nucleoplasmin. To study the interaction of nuclear targeting signals with the pore, an artificial nuclear transport substrate was constructed, which consists of human serum albumin coupled to the signal sequence of the SV40 T-antigen. A similar peptide-protein conjugate was made using a mutant signal sequence. These conjugates were fluorescently labeled and/or tagged with gold and tested for transport in the in vitro system. High levels of nuclear transport of the wild-type signal sequence-containing protein were observed, while no transport of the mutant signal sequence-containing protein was seen. Thus, the in vitro system correctly recognizes the single amino acid change between the wild-type and mutant signal sequences. We found that the observed nuclear transport was completely dependent on the presence of ATP. Using the in vitro system we identified a specific inhibitor of nuclear transport, the lectin wheat germ agglutinin (WGA), which we find binds directly to the nuclear pore. Probing blots of nuclear proteins with 125I-WGA identified a family of nuclear pore glycoproteins, including one major glycoprotein of 62K (K = 10(3)Mr) molecular weight. With the inhibitor and the in vitro assay, it has been possible to experimentally separate nuclear transport into two steps: (1) a step in which the signal sequence-bearing protein binds to the pore, followed by (2) a step in which the protein translocates through the pore. It is this second step which is the ATP-dependent step of transport, since pore binding but not translocation was seen to occur in the absence of ATP.
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PMID:Nuclear transport in vitro. 261 52

A new glycoprotein of 31,500 dalton, which has a high affinity for ouabain, and is independent of (Na+-K+)-ATPase, was solubilized from transverse tubule membrane and junctional sarcoplasmic reticulum complexes (TTM-JSR) of cat cardiac muscle. This protein could be extracted only in small amounts from sarcolemma-plasma membrane (SLM-PL) fragments, suggesting that it indeed originates from the TTM-JSR.
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PMID:Solubilization and characterization of a ouabain-sensitive protein from transverse tubule membrane-junctional sarcoplasmic reticulum complexes (TTM-JSR) in cat cardiac muscle. 272 38

Polyclonal rabbit antibodies were raised against 4-acetamido-4'-isothiocyanostilbene-2,2'-disulphonic acid (SITS), an inhibitor of a variety of anion transport proteins. These antibodies specifically recognize SITS-reacted erythrocyte band 3 in immunoprecipitations and Western blots. In Western blots of SITS-reacted membrane proteins derived from vesicles of the electric organ of Torpedo californica (known to express a SITS-sensitive Cl- channel) the antibodies recognized two major species of approximately 93 kDa and approximately 105 kDa. The approximately 93 kDa protein was identified as the alpha-subunit of the Na,K-ATPase. The approximately 105 kDa protein (designated sp105) is a glycoprotein which binds to wheat-germ agglutinin and concanavalin A and is present as a disulphide-linked homodimer under non-reducing conditions. A partial amino acid sequence and a polyclonal antibody were used to clone the corresponding cDNA. sp105 is encoded in electroplax by two abundant mRNAs of approximately 6 and approximately 6.8 kb. A hybridizing mRNA of approximately 5 kb was over 200-fold and over 500-fold less abundant in brain and heart respectively. Sequence analysis of the cDNA predicted a novel protein of 697 amino acids containing eight potential N-linked glycosylation sites. Analysis of hydrophobicity indicated the presence of at least one, and possibly three, putative membrane-spanning domains. When expressed from the Sp6 message in Xenopus laevis oocytes, the protein was inserted into membranes, glycosylated and processed to form a dimer. However, no increase in 36Cl uptake or in membrane conductance could be detected. We found no effect of hybrid depleting the specific message on expression of the Torpedo electroplax Cl- channel in oocytes. Thus we conclude that this novel electroplax membrane protein is probably not a functional part of the chloride channel.
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PMID:Primary structure of a novel 4-acetamido-4'-isothiocyanostilbene-2,2'-disulphonic acid (SITS)-binding membrane protein highly expressed in Torpedo californica electroplax. 277 1

The microsomal (H+,K+)-ATPase systems from dog and pig fundic mucosa were purified to homogeneity and partially characterized. The method involves sodium dodecyl sulfate (SDS) (0.033% w/v) extraction of the microsomal non-ATPase proteins under appropriate conditions followed by sucrose density gradient centrifugation. Two distinct membrane bands of low (buoyant density = 1.08 g/mL) and high (buoyant density = 1.114 g/mL) densities having distinct enzymatic and chemical composition were harvested. The low-density membrane was highly enriched in Mg2+- or Ca2+-stimulated ATPase and 5'-nucleotidase activities but totally devoid of (H+,K+)-ATPase and K+-p-nitrophenylphosphatase activities. The latter two activities were found exclusively in the high-density membrane. SDS-polyacrylamide gel electrophoresis revealed the high-density membranes to consist primarily of a major 100-kilodalton (kDa) protein and a minor 85-kDa glycoprotein, the former being the catalytic subunit of the (H+,K+)-ATPase. The amino acid composition of the pure dog (H+,K+)-ATPase revealed close similarities with that from pig. The N-terminal amino acid was identified to be lysine as the sole residue. Similar to the high-density membrane-associated pure (H+,K+)-ATPase, the low-density membranes containing high Mg2+-ATPase activity also contained a 100-kDa peptide and a 85-kDa glycopeptide in addition to numerous low molecular weight peptides. Also, similar to the pure (H+,K+)-ATPase, the Mg2+-ATPase-rich fraction produced an E approximately P unstable to hydroxylamine and partially (about 25%) sensitive to K+ but having a slow turnover. The levels of E approximately P produced by the pure (H+,K+)-ATPase- and Mg2+-ATPase-rich fractions were 1400 and 178 pmol/mg of protein, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Purification and partial characterization of the (H+,K+)-transporting adenosinetriphosphatase from fundic mucosa. 282 83

Highly specific antibodies against vital enzymes of the collecting ducts were used to study the appearance of cell type specific enzyme profiles in developing rat kidneys. (Na+K)-ATPase, the abundant enzyme of principal cells, could be detected early in utero in most collecting duct cells. However, the characteristic basolateral polarization of this enzyme did not appear until the first hours after birth. After this, the relative amount of (Na+K)-ATPase immunoreactive cells along collecting ducts decreased steadily, to reach the amount found in adult rat kidneys by the 30th postnatal day. Carbonic anhydrase immunoreactivity characteristic for intercalated cells was not detectable in fetal kidneys, but appeared soon after birth, with steadily increasing numbers of cells that were positive. Interestingly, immunoreactive band 3 glycoprotein (anion channel protein of erythrocytes) did not appear until the 5th day of life, with only a slowly increasing number of cells positive for this probe. These results, showing the sequential appearance of cell type-specific enzyme reactivities along collecting ducts, likely reflect a similar pattern of functional development of the respective main cell types. These results may provide an explanation for physiologic neonatal acidosis, as the enzyme profile associated with proton secretion was seen to appear slowly during the first weeks of life in a distinct manner.
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PMID:Ontogeny of cell type-specific enzyme reactivities in kidney collecting ducts. 282 8

The cellular and subcellular distribution of 5'-nucleotidase in tissues of the electric ray Torpedo marmorata has been investigated by means of an antiserum raised against the native enzyme purified from the electric organ. As revealed by immunohistochemistry the enzyme is associated with the surface of the axons of the electric nerves and of spinal nerves. Using the post-embedding colloidal gold technique at the electron-microscopical level 5'-nucleotidase could be located at the plasma membrane of the Schwann cells including the myelin and the fine processes covering the terminal axon ramifications. Also the perineurial sheath of the axons inside the electric organ is 5'-nucleotidase positive. The plasma membrane of the axon and the terminal axon region or the postsynaptic membrane do not contain 5'-nucleotidase. Immunoprecipitation studies using polyacrylamide beads suggest that the ecto-Ca2+- or -Mg2+-adenosine 5'-triphosphatase previously ascribed to synaptosomes of the Torpedo electric organ is not associated with the same membranes as 5'-nucleotidase. Within the electric organ the dorsal plasma membrane of the electroplaque cell, blood capillaries and the connective tissue layer surrounding the columns of electroplaque cells also bind the antibodies. In central nervous tissue solely blood vessels show immunofluorescence. Within the electric lobe both the surface of the electromotor neurons as well as the myelinated axons giving rise to the electric nerve are negative. This also applies to the axons of the optic nerve suggesting that the antiserum is Schwann cell specific, and does not bind to a potential oligodendroglial 5'-nucleotidase. In peripheral tissue the surface of skeletal muscle fibres as well as that of individual myofibrils bind the anti-5'-nucleotidase antibodies. Our results demonstrate that the Schwann cell plasma membrane, including myelin, contains 5'-nucleotidase and that one can distinguish by means of a specific antiserum between Schwann cell and oligodendroglia plasma membranes. The functional significance of the association of 5'-nucleotidase with Schwann cells along the entire surface of axons including the synaptic region as well as with other parts of the electric tissue is discussed regarding its catalytic activity and also the possibility that this surface glycoprotein may be involved in mediating cellular interactions.
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PMID:Monospecific antiserum against 5'-nucleotidase from Torpedo electric organ: immunocytochemical distribution of the enzyme and its association with Schwann cell membranes. 283 6


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