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)

In this review, the cell envelope of the human pathogenic yeast Candida albicans includes the plasma membrane and the mannoproteins, enzymes, beta-glucans, and chitin of the wall. The organization of the wall is complex and ultrastructural studies show distinct "layers". Mannoprotein is distributed throughout the wall but is concentrated on the exterior surface and adjacent to the plasma membrane. The mannoproteins contain the antigenic determinants of the yeast cells. The major structural components of the wall are beta-1,3- and beta 1,6-glucans, and these two linkages are present in almost equal amounts. Chitin is concentrated at the bud scar, but small amounts are located over the entire wall where it appears to be linked to beta-1,6-glucan. Chemical bonding both within and between wall components confers rigidity on the wall and restricts movement of molecules into and out of the cell. Soluble enzymes are retained within the wall matrix, but a number of enzymes and proteins are excreted. The plasma membrane of C. albicans is similar to that isolated from other fungi and contains the proton pump ATPase and enzymes involved in biosynthesis of the wall such as chitin synthase and beta-1,3-glucan synthase.
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PMID:Cell envelope of Candida albicans. 331 22

1,3-beta-D-glucan synthase [also known as beta(1-->3) glucan synthase] is a multi-enzyme complex that catalyzes the synthesis of 1,3-beta-linked glucan, a major structural component of the yeast cell wall. Temperature-sensitive mutants in the essential Rho-type guanosine triphosphatase (GTPase), Rho1p, displayed thermolabile glucan synthase activity, which was restored by the addition of recombinant Rho1p. Glucan synthase from mutants expressing constitutively active Rho1p did not require exogenous guanosine triphosphate for activity. Rho1p copurified with beta(1-->3)glucan synthase and associated with the Fks1p subunit of this complex in vivo. Both proteins were localized predominantly at sites of cell wall remodeling. Therefore, it appears that Rho1p is a regulatory subunit of beta(1-->3)glucan synthase.
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PMID:Identification of yeast Rho1p GTPase as a regulatory subunit of 1,3-beta-glucan synthase. 860 5

The callose synthase (UDP-glucose: 1,3-beta-D-glucan 3-beta-D-glucosyl transferase; EC 2.4.1.34) enzyme (CalS) from pollen tubes of Nicotiana alata Link et Otto is responsible for developmentally regulated deposition of the cell wall polysaccharide callose. Membrane preparations from N. alata pollen tubes grown in liquid culture were fractionated by density-gradient centrifugation. The CalS activity sedimented to the denser regions of the gradient, approximately 1.18 g.ml-1, away from markers for Golgi, endoplasmic reticulum and mitochondria, and into fractions enriched in ATPase activity and in membranes staining with phosphotungstic acid at low pH. This suggests that pollen-tube CalS is localised in the plasma membrane. Callose synthase activity from membranes enriched by downward centrifugation was solubilised with digitonin, which gave a 3- to 4-fold increase in enzyme activity, and the solubilised activity was then enriched a further 10-fold by product entrapment. The complete procedure gave final CalS specific activities up to 1000-fold higher than those of pollen-tube homogenates. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that several polypeptides co-fractionated with CalS activity through purification, with a polypeptide of 190 kDa being enriched in product-entrapment pellets.
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PMID:Membrane fractionation and enrichment of callose synthase from pollen tubes of Nicotiana alata Link et Otto. 964 Jun 64

Permeabilization of yeast and other fungal cells by osmotic shock enabled the in situ assays of intracellular plasma membrane-bound enzymes, such as beta-1,3-glucan synthase, chitin synthase, and Na(+)/K(+) ATPase as well as the soluble, cytoplasmic enzymes, such as lactate dehydrogenase and alpha-glucosidase. The permeabilization was accomplished by rapid changes in osmolarity of the washing buffer at 0 degrees C whereby 0.5-3.5 M glycerol, sorbitol, and/or mannitol and/or 1 M KCl could be used as the osmolytes. No appreciable leakage of intracellular proteins occurred during the permeabilization procedure. The described procedure caused practically complete cell permeabilization while avoiding treatments with organic solvents, detergents, and other xenobiotics currently used for the permeabilization of microbial cells.
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PMID:In situ assays of fungal enzymes in cells permeabilized by osmotic shock. 1131 15

Elongation of subapical segments of maize (Zea mays) roots was greatly inhibited by (2)H(2)O in the incubation medium. Short-term exposure (30 min) to (2)H(2)O slightly reduced O(2) uptake and significantly increased ATP levels. (2)H(2)O inhibited H(+) extrusion in the presence of both low (0.05 mm) and high (5 mm) external concentrations of K(+) (about 30 and 53%, respectively at 50% [v/v] (2)H(2)O). Experiments on plasma membrane vesicles showed that H(+)-pumping and ATPase activities were greatly inhibited by (2)H(2)O (about 35% at 50% [v/v] (2)H(2)O); NADH-ferricyanide reductase and 1,3-beta-glucan synthase activities were inhibited to a lesser extent (less than 15%). ATPase activities present in both the tonoplast-enriched and submitochondrial particle preparations were not affected by (2)H(2)O. Therefore, the effect of short incubation time and low concentration of (2)H(2)O is not due to a general action on overall cell metabolism but involves a specific inhibition of the plasma membrane H(+) -ATPase. K(+) uptake was inhibited by (2)H(2)O only when K(+) was present at a low (0.05 mm) external concentration where absorption is against its electrochemical potential. The transmembrane electric potential difference (E(m)) was slightly hyperpolarized by (2)H(2)O at low K(+), but was not affected at the higher K(+) concentrations. These results suggest a relationship between H(+) extrusion and K(+) uptake at low K(+) external concentration.
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PMID:Effects of deuterium oxide on growth, proton extrusion, potassium influx, and in vitro plasma membrane activities in maize root segments. 1665 24

Detergent-solubilized plasma membrane proteins from pea (Pisum sativum L.) stem tissue were separated by isoelectric focusing (IEF) using a Bio-Rad Rotofor cell, with the goal of identifying protein(s) involved in beta-1,3-glucan synthase (GS-II) activity. Ordinary IEF procedures result in membrane protein precipitation. Inclusion of 10% glycerol mitigates this problem in digitonin-solubilized preparations, but not in those solubilized in 3-[(-cholamidopropyl)dimethylammonio]-1-propanesulfonate. Loss of GS-II activity during IEF is minimized by improved cooling of the Rotofor cell. GS-II focuses at pH 5.1. Antiserum against a 55 kilodalton (kD) polypeptide that was recognized from other evidence as involved in GS-II activity, detects this polypeptide in exact correspondence with the GS-II activity peak. A presumptive P-type ATPase, detected using an antibody against corn root plasma membrane 97 kD ATPase, focuses at pH 5.3. In this digitonin/glycerol medium, most of the membrane proteins focus within the relatively narrow pH range of 4.5 to 6, compared to pH 5.5 to 8.5 for IEF in the presence of 9 molar urea, 2% Nonidet P-40 (NP-40), and 5% mercaptoethanol, a medium that inactivates GS-II. This latter medium increases the apparent isoelectric point (pl) values of the abovementioned 55 and 97 kD polypeptides to 5.8 and 7.3, respectively. In the digitonin/glycerol medium, membrane polypeptides apparently focus at pH values lower than their true pls, because of adhering negatively charged phospholipids, which can be at least partially removed by the detergent NP-40 in the presence of urea. These results provide independent evidence that the 55 kD polypeptide is associated with the GS-II activity and indicate that inclusion of urea and a strong nonionic detergent such as NP-40 is necessary if membrane proteins are to be focused at pH values near their true pls.
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PMID:Isoelectric Focusing of Plant Plasma Membrane Proteins : Further Evidence that a 55 Kilodalton Polypeptide Is Associated with beta-1,3-Glucan Synthase Activity from Pea. 1666 30

The NaGSL1 gene has been proposed to encode the callose synthase (CalS) enzyme from Nicotiana alata pollen tubes based on its similarity to fungal 1,3-beta-glucan synthases and its high expression in pollen and pollen tubes. We have used a biochemical approach to link the NaGSL1 protein with CalS enzymic activity. The CalS enzyme from N. alata pollen tubes was enriched over 100-fold using membrane fractionation and product entrapment. A 220 kDa polypeptide, the correct molecular weight to be NaGSL1, was specifically detected by anti-GSL antibodies, was specifically enriched with CalS activity, and was the most abundant polypeptide in the CalS-enriched fraction. This polypeptide was positively identified as NaGSL1 using both MALDI-TOF MS and LC-ESI-MS/MS analysis of tryptic peptides. Other low-abundance polypeptides in the CalS-enriched fractions were identified by MALDI-TOF MS as deriving from a 103 kDa plasma membrane H+-ATPase and a 60 kDa beta-subunit of mitochondrial ATPase, both of which were deduced to be contaminants in the product-entrapped material. These analyses thus suggest that NaGSL1 is required for CalS activity, although other smaller (<30 kDa) or low-abundance proteins could also be involved.
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PMID:Proteomic and biochemical evidence links the callose synthase in Nicotiana alata pollen tubes to the product of the NaGSL1 gene. 1766 22

A procedure is described in which vacuoles are isolated from leaf tissue of lettuce (Lactuca sativa L.). After incubation in an enzyme solution, the vacuoles are directly extracted from the leaf tissue by osmotic shock using a phosphate buffer. In this method no protoplasts are released from the leaf tissue. This procedure avoids the problems of separating vacuoles from protoplasts with similar density. To evaluate the purity of the vacuoles, the activity of glucan synthetase 11 (EC 2.4.1.34), NAD(P) H-cytochrome c reductase (EC 1.6.99.3) and malate dehydrogenase (EC 1.1.1.37) was measured. To measure vanadate- and nitrate-sensitive ATPase activity (EC 3.6.1.8) vesicles were prepared from the vacuoles and ATP-dependent vesicle acidification was measured as acridine orange quenching. Nitrate inhibited the quenching, while addition of vanadate had no effect. It was concluded that the vacuoles were not contaminated with plasma membranes. To evaluate the viability of the vacuoles [(14) C]-malate uptake was measured. The vacuoles showed a constant rate of [(14) C]-malate uptake during 45 min. This rate was maximal at pH 6.8.
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PMID:Direct isolation of vacuoles from leaf tissue of lettuce (Lactuca sativa) retaining protoplasts within the leaves. 2108 81