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:3.1.3.1 (alkaline phosphatase)
47,916 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have investigated the possible role of a cAMP-mediated protein-phosphorylation event(s) as the key regulatory mechanism in beta-adrenoreceptor-stimulated activation of mannosylphosphodolichol (Man-P-Dol) synthase (GDP-mannose:dolichyl-phosphate O-beta-D-mannosyltransferase, EC 2.4.1.83) in rat parotid acinar cells. Microsomal membranes isolated from these cells pretreated with 10 microM isoproterenol for 60 min showed approximately 40-80% enhanced Man-P-Dol synthase activity compared to the untreated controls. This change in enzyme activity was not associated with a significant alteration in apparent Km for GDP-mannose, but the Vmax was enhanced 2-fold. When microsomal membranes isolated from control cells were phosphorylated in vitro by a cAMP-dependent protein kinase, an increase in Man-P-Dol synthase activity, similar to that with membranes from isoproterenol-treated cells, was observed (i.e., a moderate change in Km for GDP-mannose but a 2-fold higher Vmax). Furthermore, treatment of in vitro phosphorylated microsomal membranes by alkaline phosphatase led to a substantial reduction in Man-P-Dol synthase activity. Increased Man-P-Dol synthesis (approximately 30-40%) was also observed in bovine brain and hen oviduct microsomal membranes after in vitro protein phosphorylation. In aggregate, these results strongly suggest that agents that increase cAMP in cells may modulate protein N-glycosylation in those cells by activating this key glycosyltransferase of the dolichol cascade by a cAMP-dependent protein kinase-mediated protein phosphorylation/dephosphorylation cycle.
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PMID:cAMP-mediated protein phosphorylation of microsomal membranes increases mannosylphosphodolichol synthase activity. 281 74

The role of N-linked oligosaccharide side chains in the biogenesis and function of Na+-coupled transporters in renal luminal brush-border membrane (BBM) is not known. We examined the question of how in vivo inhibition by alkaloid swainsonine of alpha-mannosidase, a key enzyme in processing of glycoproteins in the Golgi apparatus, affects Na+/H+ antiport and Na+/Pi symport as well as activities of other transporters and enzymes in rat renal BBM. Administration of swainsonine to thyroparathyroidectomized rats, control or treated with 3,5,3'-triiodothyronine, markedly decreased the rate of Na+/H+ antiport, but had no effect on the rate of Na+/Pi symport across renal BBM vesicles (BBMV). Moreover, administration of swainsonine did not change activities of Na+ gradient, ([extravesicular Na+] greater than [intravesicular Na+])-dependent transport of D-glucose, L-proline, or the amiloride-insensitive 22Na+ uptake by BBMV; the activities of the BBM enzymes alkaline phosphatase, gamma-glutamyltransferase, or leucine aminopeptidase in BBMV were also not changed. The in vitro enzymatic deglycosylation of BBM by incubating freshly isolated BBMV with bacterial endoglycosidase F also resulted in a decreased rate of Na+/H+ antiport, but not Na+-coupled symports of Pi, L-proline, and D-glucose, or the activities of the BBM enzymes were not significantly affected. Similar incubation with endoglycosidase H was without effect on any of these parameters. Both the modification of BBMV glycoproteins by administration fo swainsonine in vivo as well as the in vitro incubation of BBMV with endoglycosidase F resulted in a decrease of the apparent Vmax of Na+/H+ antiport, but did not change the apparent Km of this antiporter for extravesicular Na+ and did not increase H+ conductance of BBM. Taken together, our findings suggest that intact N-linked oligosaccharide chains of the biantennary complex type in renal BBM glycoproteins are required, directly or indirectly, for the transport function of the Na+/H+ antiporter inserted into BBM of renal proximal tubules.
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PMID:Role of N-linked oligosaccharides in the transport activity of the Na+/H+ antiporter in rat renal brush-border membrane. 284 30

Surface protein mutants of the invasive Salmonella species, S. choleraesuis, were generated using the transposon TnphoA. 626 alkaline phosphatase (PhoA+) fusion mutants were identified and screened for their ability to pass through (transcytose) polarized epithelial monolayers of Madin Darby canine kidney (MDCK) cells grown on membrane filters. Forty two mutants were unable to pass through this barrier. All of these transcytosis mutants were unable to adhere to or invade MDCK monolayers, yet these mutations were not in the genes encoding type 1 pili or mannose-resistant haemagglutination (MRHA). These transcytosis mutants could be grouped into six classes. Class 1 mutants had altered lipopolysaccharide (LPS) O side-chain structures while Class 2 mutants had defects in their LPS core. Mutants belonging to Classes 5 and 6 did not decrease the transepithelial electrical resistance of polarized MDCK cell monolayers, in contrast to the parental strain and the other mutants (Classes 1, 2, 3 and 4). Mutants belonging to Class 1 were less virulent in mice, while Class 2 (defective core) and Classes 4 and 5 (normal LPS) mutant strains were avirulent in mice. Mutants from Classes 3 and 6 were as virulent in mice as S. choleraesuis. These results suggest that the ability to pass through epithelial barriers may be an important virulence characteristic of Salmonella. These data indicate that bacterial adherence, internalization and monolayer transcytosis are closely linked events. It was also demonstrated that a mutant with decreased rates of intracellular replication still passed through the monolayer at rates similar to wild-type S. choleraesuis.
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PMID:Identification and characterization of TnphoA mutants of Salmonella that are unable to pass through a polarized MDCK epithelial cell monolayer. 285 Apr 43

alpha-Lactalbumin was isolated from milk of M. eugenii and its concentration in milk samples taken at various times during lactation (0-40 weeks post partum) was determined by single radial immunodiffusion using rabbit antiserum to the purified protein. The alpha-lactalbumin concentration remained almost constant throughout lactation even though the concentration of total lactose (free lactose plus lactose contained in oligosaccharides) fell to zero after 34 weeks post partum. This fall in lactose was accompanied by a rise in the free galactose and glucose concentrations and marked increases in UDP-galactose hydrolase, nucleotide pyrophosphatase, alkaline phosphatase and acid beta-galactosidase activities. It is suggested that the in vitro hydrolysis of UDP-galactose was due to nucleotide pyrophosphatase and that this enzyme may also play a role in vivo late in lactation by making UDP-galactose unavailable for the synthesis of lactose. Alternatively, lactose and lactose-containing oligosaccharides might be degraded by the acid beta-galactosidase during or after secretion.
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PMID:Changes in alpha-lactalbumin, total lactose, UDP-galactose hydrolase and other factors in tammar wallaby (Macropus eugenii) milk during lactation. 285 90

We reviewed charts of 38 Patients who developed cholestasis whilst receiving total parenteral nutrition (TPN). A standard protocol was followed for administration of TPN and included crystalline amino acid solution with lipid emulsion and dextrose as calorie sources. 5 of the 38 patients did not exhibit an increase in alkaline phosphatase. This may be related to the concomitant low levels of either serum zinc or magnesium or both. We also obtained a positive correlation between serum levels of zinc and magnesium and the peak activity of alkaline phosphatase (r = 0.49, p less than 0.01, r-0.55, p less than 0.01, respectively) associated with cholestasis. We hypothesize that the reason for this association is related to both zinc and magnesium being activators of alkaline phosphatase activity.
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PMID:Absence of increase of serum alkaline phosphatase activity with parenteral nutrition-associated cholestasis: possible consequence of hypozincemia and hypomagnesemia. 286 Oct 85

Addition of increasing amounts of benzyl alcohol progressively reduced the steady-state anisotropies of diphenylhexatriene and trimethylammoniumdiphenylhexatriene in brush-border membranes from rat kidney. The decrease in order of membrane lipids, equivalent for 50 mM benzyl alcohol to that produced by a rise in temperature of approx. 6 degrees C, had no effect on the activities of alkaline phosphatase or gamma-glutamyltranspeptidase. On the other hand, benzyl alcohol markedly inhibited the D-glucose uptakes measured in the presence of a 100 mM sodium gradient. For concentrations less than 30 mM, benzyl alcohol reduced the Jmax without significant effects on Km, 22Na+ uptake or the vesicular volume of brush-border preparations. Comparable results were obtained substituting octanol for benzyl alcohol. Our data strongly suggest that, at constant temperature, the D-glucose carrier present in renal brush-border membranes is extremely sensitive to variations in membrane physical state.
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PMID:Effects of benzyl alcohol on enzyme activities and D-glucose transport in kidney brush-border membranes. 287 65

We have treated bovine lung heparan sulfate with alkaline [3H]borohydride to end label the chains with [3H]xylitol. After subsequent periodate oxidation-alkaline elimination products were separated by gel permeation and ion exchange chromatography. The linkage region fragment expected to have 2 galactoses and 1 [3H]xylitol residue appeared in the tetra-/trisaccharide region after gel filtration and was bound to the anion exchange resin. A similar negatively charged fragment, expected to have 2 galactoses, 1 xylose and 1 serine, was isolated after periodate oxidation-alkaline elimination of unlabeled heparan sulfate. The negative charge was due to the presence of alkaline phosphatase-labile phosphate ester. The molar ratio of galactose:phosphate:xylose was 2.17:1.19:1.00. The phosphate ester was associated with the xylose/[3H] xylitol moiety as indicated by the formation of phosphoxylose/-xylitol by beta-galactosidase digestion of the phosphorylated trisaccharide. Furthermore, orcinol reactivity disappeared after periodate oxidation of the dephosphorylated trisaccharide. The phosphate ester must be located to C-2 of xylose/xylitol as the 1-3H radioactivity could be released by periodate oxidation when it was preceded by alkaline phosphatase treatment. It is estimated that almost every chain of heparan sulfate carries 2-phosphoxylose. It would be of interest to know if glycosaminoglycan chains that are artificially initiated onto exogeneous beta-D-xylosides also acquire the 2-phosphoxylose moiety.
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PMID:Structure of the heparan sulfate-protein linkage region. Demonstration of the sequence galactosyl-galactosyl-xylose-2-phosphate. 293 48

A rat hepatocyte cell line was cultured in Higuchi's medium with fetal calf serum and insulin and labeled with 35SO2/4-. The cells were treated with a number of ligands to displace the heparan 35SO4 proteoglycan (HSPG) from the pericellular matrix. Maximum release was obtained with D-mannose-6-PO4 (50 mM), D-glucose-6-PO4 (50 mM), myo-inositol-2-PO4 (2-5 mM), myo-inositol hexaphosphate (2-5 mM), and DL-myo-inositol-1-PO4 (1-2 mM). D-myo-Inositol-1,3,4-(PO4)3 (1 mM) and L-myo-inositol-1-PO4 (2 mM) were intermediate in their ability to release the cell surface HSPG, whereas heparin (2 mg/ml), yeast phosphomannan (4 mg/ml), D-xylose-1-PO4 (50 mM), D-glucose-6-SO4 (50 mM), and myo-inositol hexasulfate (5 mM) were ineffective. When 35SO2/4- was added to cell cultures, the total cell surface HSPG increased linearly, but the percentage of the total cell surface [35SO4]HSPG that was released by myo-inositol-PO4 increased with time during the labeling period, reaching a maximum of 65% after 5 h. When cells were labeled for 12 h without insulin in the medium, the maximum amount of cell surface HSPG that was released by myo-inositol-PO4 was reduced to 30%. However, when cells labeled in the absence of insulin were treated with phosphatidylinositol-specific phospholipase C and then myo-inositol-PO4, the release of the cell surface [35SO4]HSPG was increased to 73%. When the [35SO4]HSPG that was released from the cell surface by treatment with myo-inositol-PO4 was added to cultures of unlabeled hepatocytes, it was taken up very rapidly and a portion of the internalized HSPG was converted to free heparan SO4 chains which appeared in the nucleus. Uptake was Ca2+- and Mg2+-independent. The amount of [35SO4]HSPG taken up was markedly reduced when the myo-inositol-PO4-releasable [35SO4]HSPG was pretreated with trypsin, thermolysin, alkaline borohydride, or alkaline phosphatase. When the cells were grown in inositol-deficient medium or in the presence of myo-inositol-PO4, the amount of heparan SO4 found in the nucleus was markedly reduced, and the cells no longer exhibited contact inhibition. These effects of myo-inositol deficiency on the growth and nuclear heparan SO4 were accentuated by addition of LiCl to the cultures to prevent phosphatidylinositol synthesis from the endogenous myo-inositol-PO4.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Involvement of phosphatidylinositol and insulin in the coordinate regulation of proteoheparan sulfate metabolism and hepatocyte growth. 295 71

In the present in-vitro study we investigated the possible role of the calmodulin-antagonistic drugs loperamide and calmidazolium in the regulation of transepithelial Ca2+ transport of human duodenum. Brush border membrane vesicles and basolateral membrane vesicles were simultaneously prepared from surgically resected pieces of morphologically intact human duodenum with a modified Percoll-gradient centrifugation method. Brush border and basolateral membrane vesicles were characterized using enzyme marker analysis and electron microscopy: alkaline phosphatase was enriched 20-fold in brush border membrane vesicles, whereas [Na+ + K+]-stimulated adenosine triphosphatase was enriched 15-fold in basolateral membrane vesicles. Calmodulin activity was determined by a specific radioimmunoassay after solubilizing brush border and basolateral membrane vesicles in 1% Triton X-100. In basolateral membrane vesicles, we found no calmodulin activity. In brush border membrane vesicles calmodulin activity was impaired by 50% after pre-incubation with loperamide or calmidazolium. We measured calcium, sodium, D-glucose and D-mannitol uptake with a rapid filtration technique. Before the transport experiments, brush border and basolateral membrane vesicles were pre-incubated with 5 microM loperamide or 5 microM calmidazolium for 60 min at 5 degrees C. In drug-pretreated, brush border membrane vesicles calcium uptake was significantly reduced after 1 min incubation (-25% +/- 5%, P less than 0.05); this effect was completely reversed in the presence of 5 microM calmodulin. In basolateral membrane vesicles, we found two Ca2+ transport systems: (1) Na+/Ca2+ exchange and (2) ATP-dependent Ca2+ transport. In basolateral membrane vesicles loperamide had no effect. Calmidazolium had no effect on Na+/Ca2+ exchange, but significantly inhibited ATP-dependent Ca2+ transport. This effect could not be reversed by calmodulin.
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PMID:Effect of two potent calmodulin antagonists on calcium transport of brush border and basolateral vesicles from human duodenum. 297 85

Parathyroid hormone (PTH) and cAMP inhibit sodium, water, and bicarbonate reabsorption in the proximal tubule. We wished to determine whether these agents directly inhibit proximal tubular Na+/H+ exchange. A suspension of rabbit proximal tubules was prepared by enzymatic digestion and Ficoll gradient centrifugation. Oxygen consumption at 37 degrees C was stable over 60 min, averaged 20 nmol X mg protein-1 X min-1, and was inhibited 60% by ouabain. Over 96% of cells excluded trypan blue. From this suspension, brush border membrane vesicles were isolated. The vesicles were enriched 12.7 times in alkaline phosphatase relative to a cortical homogenate and demonstrated pH gradient-stimulated, amiloride-sensitive Na+/H+ countertransport and sodium-phosphate and sodium-D-glucose cotransport. When the tubule suspension was exposed to PTH or dibutyryl cAMP, the activity of Na+/H+ countertransport in the resultant brush border vesicles was inhibited. Neither PTH nor dibutyryl cAMP affected the amiloride-insensitive component of sodium transport or sodium-phosphate or sodium-D-glucose cotransport. The effect of PTH on Na+/H+ counter-transport could not be explained by an alteration in fluidity of the brush border membrane. These experiments demonstrate that PTH and dibutyryl cAMP directly inhibit Na+/H+ countertransport in the brush border membrane of the rabbit proximal tubule.
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PMID:Parathyroid hormone and dibutyryl cAMP inhibit Na+/H+ exchange in renal brush border vesicles. 298 85


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