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)

In Reuber rat hepatoma cells (R-Y121B), alkaline phosphatase activity increased without de novo enzyme synthesis (Sorimachi, K., and Yasumura, Y. (1986) Biochim. Biophys. Acta 885, 272-281). The enzyme was partially purified by butanol extraction from the particulate fractions. The incubation of the extracted alkaline phosphatase with the cytosol fraction induced a large increase in enzyme activity (5-10-fold of control). The dialyzed cytosol was more effective than the undialyzed cytosol during an early period of incubation at 37 degrees C. This difference between the dialyzed and the undialyzed cytosol fractions was due to endogenous Na+. For maximal activation of the enzyme, both Mg2+ above 1 mM and Zn2+ at low concentrations (below 0.01 mM) were needed, although Zn2+ at high concentrations (above 0.1 mM) showed an inhibitory effect. Zn2+ and Mg2+ alone slightly increased alkaline phosphatase activity. This activation of the enzyme was temperature dependent and was not observed at 0 or 4 degrees C. Polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate showed that the increase in alkaline phosphatase activity did not involve the fragmentation of the enzyme and that 65Zn2+ bound to it during enzyme activation with 65Zn2+ and Mg2+. The cytosol fraction not only supplied Zn2+ to the nascent enzyme but also increased the maximal enzyme activity more than did direct addition of metal ions. Ferritin and metallothionein contributed to the activation of alkaline phosphatase with the metal ions. Since the binding of Zn2+ and Mg2+ to the nascent alkaline phosphatase is disturbed in Reuber rat hepatoma cells (R-Y121B), the apoenzyme is accumulated inside the cells. The binding of Zn2+ and Mg2+ to the apoenzyme readily takes place in the cell homogenates accompanied by an increase in catalytic activity without new enzyme synthesis.
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PMID:Activation of alkaline phosphatase with Mg2+ and Zn2+ in rat hepatoma cells. Accumulation of apoenzyme. 380 40

Age-related changes in enzyme activities, protein electrophoretic patterns and lipid composition of kidney-brush-border membranes were studied in 10-20- and 30-month-old male and female Wistar rats. Polyacrylamide gel electrophoresis of membrane proteins revealed very little changes with increasing age, whether males or females were considered. The Km of three hydrolases - maltase, L-aminopeptidase and alkaline phosphatase - were not affected by age while the Vmax of maltase and alkaline phosphatase, but not of L-aminopeptidase, decreased from 10 to 30 months. The phospholipid to protein ratio which remained constant between 10 and 20 months, rose in both sexes from 20 to 30 months. In males, the cholesterol content of the membrane increased faster than that of phospholipid and the cholesterol over phospholipid ratio was then greater at 30 months than at 10 months, while in females this ratio remained unchanged in the course of aging. The fatty acid composition of the brush-border remained more or less constant with age in female rat whereas in the male, a 10% decrease in the proportion of arachidonic acid from 10 to 30 months was responsible for a lower unsaturation index.
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PMID:Age-related changes in enzyme activities, protein content and lipid composition of rat kidney brush-border membrane. 388 47

A literature review and ten new cases of benign transient hyperphosphatasemia of infancy are presented, with special attention paid to isoenzyme studies. Polyacrylamide gel electrophoresis, heat denaturation, and binding of alkaline phosphatase to anti-human alkaline phosphatases showed that the sources of the elevated alkaline phosphatase levels are normal bone and liver and not the small intestine. The data also suggest that the following criteria be present for a diagnosis of transient hyperphosphatasemia: (1) an age of less than 5 years, (2) variable symptoms, (3) no bone or liver disease noted on physical examination or (4) from laboratory investigations, (5) isoenzyme analysis showing elevations in both bone and liver activity, and (6) a return to normal serum alkaline phosphatase activity values within four months.
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PMID:Isoenzyme studies in transient hyperphosphatasemia of infancy. Ten new cases and a review of the literature. 401 98

Polyacrylamide gel electrophoresis of Loxosceles reclusa venom demonstrated that only one of seven or eight major (plus three or four minor) protein components caused necrosis in guinea pig skin. Sephadex gel filtration separated the venom into three major peaks, the second peak of which contained the dermonecrotic activity. Hyperimmunization of rabbits with increasing doses of venom from L. reclusa produced potent precipitating antisera, and the rabbits became resistant to lesion development. Ouchterlony-type immunodiffusion and immunoelectrophoretic studies revealed six to seven distinct precipitation lines, one of which stained intensely for esterase activity. Immunohistochemical techniques failed to detect any protease, lipase, catalase, acid phosphatase, alkaline phosphatase, or amylase activity in the venom. The spreading activity of recluse spider venom in guinea pig skin was inhibited as much as 71% by antivenom. Venom preincubated with antivenom was unable to incite lesions in guinea pig skin. Passive immunization of guinea pigs 18 h before an injection of venom conferred venom resistance upon the animals. Local injections of antivenom immediately after intradermal injections of venom markedly reduced the dermal lesion. Heparin reduced the local and systemic effects of venom when preincubated with whole venom or when administered systemically before an intradermal injection of venom. Treatment of whole venom with the chelating agent ethylenediaminetetraacetate did not inhibit its necrotic activity. Transfer studies from a 24-h lesion indicated that the necrotic activity was localized and remained active in tissue for at least 24 h but not for 5 days. No lesions developed when high concentrations of venom were intradermally injected into the skin of sacrificed guinea pigs, indicating that an interaction of body constituents and venom is essential for the development of a lesion.
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PMID:Immunological studies of Brown recluse spider venom. 414 Jan 61

Polyacrylamide-gel electrophoresis and Bio-Gel P-300 molecular-sieve chromatography of mouse duodenal alkaline phosphatase demonstrates its molecular heterogeneity, which, in a kinetic sense, is manifest also in the differential relative velocities of the heterogeneous forms of the enzyme with two substrates, phenylphosphate and beta-glycerophosphate. Different treatments that eliminate most of the electrophoretic and chromatographic variability of the enzyme also decrease the velocities with both substrates so that the molar ratio of hydrolysis of one substrate relative to the other is also altered to a low but stable value. Concomitant with these changes, lipids and peptides are dissociated from the enzyme. The lipids are tentatively identified as a sterol and phospholipids. The peptides have an average composition of four to six amino acids and appear to be strongly electropositive. The conditions of dissociation suggest that their binding to the enzyme is non-covalent and predominantly based on hydrophobic and ionic bonding. The concept of lipid and peptide association would suggest prima facie differential molecular weights as a factor in the observed electrophoretic and chromatographic heterogeneity. However, the molecular forms of the enzyme with differences in elution volume equivalent to more than one-half the void volume of the Bio-Gel P-300 column, or even enzyme fractions dissociated from the lipids and peptides compared with undissociated portions, do not show any differences in sedimentation on sucrose-density-gradient centrifugation. This may be because the alterations in molecular weight owing to binding of small molecules are too small to be detected by this method. Alternatively, since lipids are involved, the binding may alter the partial specific volume in such a way that the buoyant density is not significantly altered.
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PMID:Molecular heterogeneity of mouse duodenal alkaline phosphatase. Association of lipids and peptides. 445 9

Polyacrylamide gel electrophoresis was used to investigate the relation of the soluble thiamine triphosphatase activity of various rat tissues to other phosphatases. This technique separated the thiamine triphosphatase of rat brain, heart, kidney, liver, lung, muscle and spleen from alkaline phosphatase (EC 3.1.3.1), acid phosphatase (EC 3.1.3.2) and other nonspecific phosphatase activities. In contrast, the hydrolytic activity for thiamine triphosphate in rat intestine moved identically with alkaline phosphatase in gel electrophoresis. Thiamine triphosphatase from rat liver and brain was also separated from alkaline phosphatase and acid phosphatase by gel chromatography on Sephadex G-100. This gave an apparent molecular weight of about 30,000 and a Stokes radius of 2.5 nanometers for brain and liver thiamine triphosphatase. The intestinal thiamine triphosphatase activity of the rat was eluted from the Sephadex G-100 column as two separate peaks (with apparent molecular weights of over 200,000 and 123,000) which exactly corresponded to the peaks of alkaline phosphatase. The isoelectric point (pI) of the brain thiamine triphosphatase was 4.6 (4 degrees C). The partially purified thiamine triphosphatase from brain and liver was highly specific for thiamine triphosphate. The results suggest that, apart from the intestine, the rat tissues studied contain a specific enzyme, thiamine triphosphatase (EC 3.6.1.28). The specific enzyme is responsible for most of the thiamine triphosphatase activity in these tissues. Rat intestine contains a high thiamine triphosphatase activity but all of it appears to be due to alkaline phosphatase.
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PMID:The relation of the soluble thiamine triphosphatase activity of various rat tissues to nonspecific phosphatases. 627 68

The synthesis of N-[5-(hydroxyethyl)dithio-2-nitrobenzoylaminoethyl] acrylamide (I) is described. If the disulfide bond in this compound is reduced with thiol reagents, an intense yellow color develops (epsilon 412 V 13,600 at pH 7.4) due to essentially the same chromophore as 5-thio-2-nitrobenzoic acid, the reduced form of 5,5'-dithiobis(2-nitrobenzoic acid)(Ellman's reagent). Polyacrylamide gels were prepared that were crosslinked with N,N'-methylenebisacrylamide and which contained I as an integral part of the polymerized acrylamide chain. Acetylcholinesterase (from electric eel and human brain tissue slices) and alkaline phosphatase (from Escherichia coli and calf intestine) were subjected to electrophoresis and then the gels were immersed in an appropriate thiol-substrate buffer (acetylthiocholine and cysteamine-S-phosphate, respectively). A yellow band developed rapidly in the acrylamide gel at the site of enzyme activity. Electrophoresis on the mixed disulfide-polyacrylamide gel proved to be a rapid and sensitive technique to detect very small amounts of enzyme (approximately 0.02 fmol acetylcholinesterase) and should have wide application for detecting other enzymes that hydrolyze thiol substrates.
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PMID:Polyacrylamide gels which contain a novel mixed disulfide compound can be used to detect enzymes that catalyze thiol-producing reactions. 636 79

Subcutaneous implantation of demineralized diaphyseal bone matrix in allogeneic rats results in the local induction of endochondral bone differentiation. We have explored the potential of three dissociative extractants, 4 M guanidine hydrochloride (Gdn . HCl), 8 M urea/1 M NaCl, and 1% NaDodSO4 at pH 7.4, containing protease inhibitors to solubilize putative inductive molecules in the bone matrix. Extraction of bone matrix with any one of these extracts resulted in the loss of the bone inductive property. The solubilized extracts were then reconstituted with the residue by dialysis against water. The various reconstituted matrices were bioassayed for bone inductive potential by quantitation of alkaline phosphatase activity and 45Ca incorporation on day 12 after implantation. There was complete recovery of biological activity after reconstitution of the residues with each of the three extracts. Polyacrylamide gel electrophoresis of the extracts revealed similar protein profiles. Gel filtration of the 4 M Gdn. HCl extract on Sepharose CL-4B showed a heterogeneous broad peak. When fractions of that peak containing proteins less than 50,000 daltons were reconstituted with inactive 4 M Gdn . HCl-treated bone matrix and then implanted, new bone was induced. These observations demonstrate the dissociative extraction and successful biological reconstitution of bone inductive macromolecules in demineralized bone matrix.
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PMID:Dissociative extraction and reconstitution of extracellular matrix components involved in local bone differentiation. 695 Apr 1

The isoenzymes of acid and alkaline phosphatases and their histochemical localization were studied by polyacrylamide disc gel electrophoresis in four species of trematodes: Gigantocotyle explanatum from the liver and Gastrothylax crumenifer from the rumen of water buffalo (Bubalus bubalis) and Echinostoma malayanum and Fasciolopsis buski from the small intestine of the pig (Sus scrofa). Both acid and alkaline phosphatases were present in the tegument, gastrodermis, suckers, testes, ovary, eggs, vitellaria and uterus but alkaline phosphatase activity was demonstrated only in the parenchyma and excretory ducts. Polyacrylamide gel electrophoresis revealed two to four isoenzymes for both acid and alkaline phosphatase.
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PMID:Histochemical and electrophoretic studies on phosphatases of some Indian trematodes. 709 66

The reliability of histochemical determinations of the enzyme activity after thermal damage has been studied with the aid of two model systems. Polyacrylamide films and erythrocyte ghosts containing either beta-glucuronidase or alkaline phosphatase, were submitted to heating and the activities retained were assessed both biochemically and histochemically. For the enzymes studied, the results show that tissue alterations induced by heat can influence histochemical reaction procedures, and that with these model systems, factors which are important for the histochemical quantitation of enzyme activities in thermally damaged tissues can be evaluated quantitatively. Potentialities of these model systems in the study of evaluating thermal damage through histochemical enzyme activity determinations, are discussed.
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PMID:Histochemical model studies of enzyme activity after thermal damage. 714 91


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