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)

De novo synthesis of two periplasmic enzymes in Escherichia coli, alkaline phosphatase and acid hexose phosphatase, have been studied in the presence and absence of new phospholipid synthesis. Alkaline phosphatase synthesis was initiated by a temperature shift in a strain carrying a phoA amber mutation and a temperature-sensitive suppressor mutation; acid hexose phosphatase was studied after relief of catabolite repression. Glycerol auxotrophs (gpsA) were used to control phospholipid synthesis. Synthesis of both enzymes proceeded at a normal rate for 0.5 to 1.0 generation of growth, although it was then curtailed. It is concluded that secretion of these enzymes is not obligatorily coupled to new net phospholipid synthesis.
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PMID:Enzyme secretion in Escherichia coli: synthesis of alkaline phosphatase and acid hexose phosphatase in the absence of phospholipid synthesis. 78 56

Incubation of rat adipocytes with 1 microM-noradrenaline caused a decrease in both the N-ethylmaleimide-sensitive (microsomal) and N-ethylmaleimide-insensitive (mitochondrial) glycerol phosphate acyltransferase activities measured in homogenates from freeze-stopped cells. The effects of noradrenaline on glycerol phosphate acyltransferase activity were apparent over a wide range of concentrations of glycerol phosphate and palmitoyl-CoA. The effect of noradrenaline was reversed within cells by the subsequent addition of insulin or propranolol. Inclusion of albumin in homogenization buffers abolished the effect of noradrenaline on the N-ethylmaleimide-sensitive activity. The effect of noradrenaline on the N-ethylmaleimide-insensitive (mitochondrial) activity was, however, not abolished by inclusion of albumin in buffers for preparation of homogenates from freeze-stopped cells. Inclusion of fluoride in homogenization buffers did not alter the observed effect of noradrenaline. The inactivating effect of noradrenaline persisted through the subcellular fractionation procedures used to isolate adipocyte microsomes (microsomal fractions). The effect of noradrenaline on mitochondrial glycerol phosphate acyltransferase did not persist through subcellular fractionation. Noradrenaline treatment of cells significantly decreased the Vmax. of glycerol phosphate acyltransferase in isolated microsomes without changing the activity of NADPH-cytochrome c reductase. Glycerol phosphate acyltransferase activity in microsomes from noradrenaline-treated cells is unstable, being rapidly lost on incubation at 30 degrees C. Bivalent metal ions (Mg2+, Ca2+) or post-microsomal supernatant protected against this inactivation. Glycerol phosphate acyltransferase activity in microsomes from noradrenaline-treated cells could not be re-activated by incubation with either alkaline phosphatase or phosphoprotein phosphatase-1. Addition of cyclic AMP-dependent protein kinase catalytic subunits to adipocyte microsomes incubated with [gamma-32P]ATP considerably increased the incorporation of 32P into microsomal protein, but did not cause inactivation of glycerol phosphate acyltransferase. These findings provide no support for the proposal that inactivation of adipocyte microsomal glycerol phosphate acyltransferase by noradrenaline is through a phosphorylation type of covalent modification.
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PMID:Regulation by noradrenaline of the mitochondrial and microsomal forms of glycerol phosphate acyltransferase in rat adipocytes. 635 49

3-Oxo-5 alpha-steroid: NAD+ delta 4-oxidoreductase ("NADH-5 alpha-reductase", EC 1.3.1.?) is rapidly inactivated in the presence of 17 beta-hydroxy-4-androsten-3-one (testosterone). This activation is prevented by increasing the phosphate concentration. When the enzyme assay is carried out in Tris-HCl, only a small activity (1.7 nmol X min-1 X mg-1) is observed which may be further decreased by addition of phosphatases. Addition of the phosphatase inhibitor dextran sulphate or ATP, Mg++ and c-AMP results in a significant increase of activity (228% and 273%, respectively) compared with the Tris-HCl control. Glycerol 2-phosphate and glycerol 3-phosphate have a stabilizing effect on 3-oxo-5 alpha-steroid: NAD+ delta 4-oxidoreductase by decreasing the Km towards the substrate testosterone from 1.2 X 10(-5) mol/l to 3.3 X 10(-6) mol/l. V remains unchanged. Half maximal velocity (testosterone reduction) is achieved with 20 mumol/l glycerol 2-phosphate and glycerol 3-phosphate. Addition of c-AMP dependent protein kinase (EC 2.7.1.37) to a microsomal preparation pretreated with alkaline phosphatase (EC 3.1.3.1) results in a significant increase of 3-oxo-5 alpha-steroid: NAD+ delta 4-oxidoreductase activity compared with the control.
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PMID:Rat liver 3-oxo-5 alpha-steroid delta 4-dehydrogenase. Modulation of enzyme activity by changes in phosphorylation state. 652 91

A new DNA hybridization technique, based on chromatographic migration of DNA on a nitrocellulose strip passing through an immobilized probe area, is described. The new paper chromatography hybridization assay (PACHA) is faster and simpler to use than the conventional dot hybridization assay. In this assay, an aliquot of biotinylated, PCR-amplified target DNA is applied to one end of a nitrocellulose strip. The DNA migrates to the opposite end of the strip by capillary forces and hybridizes to a specific DNA probe immobilized in a reaction zone (RZ), located in the middle of the strip. Unhybridized DNA migrates away from the RZ. The biotinylated hybrid is visualized by a color reaction employing a streptavidin-alkaline phosphatase (SA-AP) conjugate and a specific chromogenic substrate. The new PACHA technique allows for detection of as little as 1-5 pg of specific human papilloma virus 16 (HPV16) DNA in 25 min of hybridization. In this system, the hybridization efficiency is controlled by the flow velocity of the hybridization solution (HS) and by the volume of the amplified labeled DNA migrating across the immobilized probe. Glycerol (30%) or polyvinyl pyrrolidone (PVP) (1%) reduces the flow rate by a factor of 2.5-3 and increases the sensitivity of the assay by a factor of 5.2 for glycerol and 2.6 for PVP. This novel method ensures efficient hybridization to multiple probes and appears to be superior to currently available solid-phase hybridization techniques.
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PMID:A novel rapid hybridization technique: paper chromatography hybridization assay (PACHA). 829 6

An acid phosphatase from a heavy-metal-accumulating strain of a Citrobacter sp. was resolved into two forms on the basis of their nonbinding (phosphatase I) or binding (phosphatase II) behaviour on the cation-exchange resin SP-Sephadex C50. Both holoenzymes had a molecular mass of 103-108 kDa as determined by Superose Q-6 column chromatography in the presence of 150 mM KCl and a subunit molecular mass of 27 kDa as determined by SDS-PAGE; the enzyme was tetrameric. Both enzymes had a pI approximately 9.0 and were immunologically cross-reactive. There were minor differences in amino acid composition and in peptide maps following tryptic digest. The pH optimum for phosphatases I and II was 5.5 and 6.25, respectively; phosphatase II alone retained activity at pH values up to 9.0. Phosphatase I was more resistant to mechanical shear, gamma-irradiation, high temperature, and toxins (F- and formaldehyde). Glycerol increased the thermostability of both enzymes, particularly the more thermosensitive phosphatase II. Phosphatase II had a lower Km and a lower Vmax for glycerol 2-phosphate hydrolysis. The production of enzyme isoforms is a phenomenon similar to that described previously for the alkaline phosphatase of Escherichia coli, where the isoforms relate to precursive and final processed forms of the enzyme. Acid phosphatase is physiologically distinct, with a role that is still obscure but that may relate to cellular stress responses.
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PMID:Purification and characterization of acid-type phosphatases from a heavy-metal-accumulating Citrobacter sp. 944 88

Surfactant-like particle (SLP) is a phosphatidylcholine (PC)-rich membrane produced in the small intestine, and its secretion is increased by fat feeding. In Caco-2 cells known to produce SLP, preincubation with [(3)H]palmitate labelled the SLP and was used as a marker for newly secreted membrane. SLP-associated PC and protein (d=1.07-1.08 g/ml in a linear non-equilibrium NaBr gradient) were secreted in parallel with triacylglycerols (TG) and at a rate about twice the control rate in response to feeding cells with an oleate/egg PC mixture. Cholesterol and apolipoprotein A-I identified only a small peak corresponding to high-density lipoprotein (HDL), but the largest peak corresponded with SLP (d=1.07-1.08). Palmitate incorporation into PC showed a similar small peak migrating at the density of HDL, but most labelled PC secreted from the cells was due to SLP. PC secretion, alkaline phosphatase activity, and newly synthesized immunoprecipitated SLP proteins from conditioned serum-free media migrated together at a density of >/=1.21 g/ml in a lipoprotein NaBr step gradient, and represented SLP. Glycerol incorporated into TG migrated at a peak density of 1.12 g/ml, consistent with HDL secretion from cells incubated in serum-free media. These data confirm that the secreted PC in SLP is distinct from lipoprotein particles. Incorporation of [(3)H]palmitate into the PC fraction of either whole cell homogenate or isolated brush border membranes was not affected by oleate/egg PC feeding. Both Pluronic L-81, an inhibitor of chylomicron secretion, and BMS-197636-02, a microsomal triglyceride transfer protein inhibitor, blocked the secretion of both TG and PC. Elevation of intracellular cAMP levels that stimulate surfactant secretion from type II pneumocytes caused a 50% reduction in SLP and TG secretion from Caco-2 cells. These results confirm the SLP response to fat feeding found in vivo, further supporting a role for SLP in TG secretion from the enterocyte, and show that the regulation of SLP secretion differs from that of pulmonary surfactant.
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PMID:Regulation of surfactant-like particle secretion by Caco-2 cells. 1128 80

Laboratory friendly, cryopreservation procedures with respect to cryopreservation formulations and cryopreservation temperatures were attempted, in the present study to ensure perennial availability of cultured mantle cells of bivalve (Paphia malabarica). Screening of cryopreservative formulations with different concentrations of DMSO, Propylene glycol and Glycerol was carried out for cryopreservation of freshly dissociated cells of Paphia malabarica. Out of these cryopreservative formulations, 10% DMSO, 10% Propylene glycol and 15% Glycerol were selected for cryopreservation of the mantle cells pooled from 1-day old primary culture and cell line after 3 passages at the end of different cryopreservation periods. Cryopreservative formulation with 15% glycerol, served as a best cryoprotectant for the cryopreservation of cells sourced from freshly dissociated cells as well as from primary cultures and cell cultures after three passages of mantle cells of Paphia malabarica, retaining metabolic activity of resurrected cells. Both, cell cultures established from uncryopreserved cells as well as cryopreserved cells showed similar alkaline phosphatase and carbonic anhydrase activities thus indicating retention of their biomineralization capacity even after cryopreservation at low and ultralow temperatures.
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PMID:Cryopreservation of cultured mantle cells of Paphia malabarica for perennial availability. 2962 63