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)

1. Proteins of fat-globule membrane from bovine milk were solubilized with the non-ionic detergent Triton X-100 in the presence of protease inhibitors. Approximately 25% of the total membrane protein was solubilized and the extracts were shown to contain a sample of most of the major membrane proteins and glycoproteins. 2. The solubilized proteins were separated in flat-beds of Ultrodex by electrofocusing and the pI values for the major proteins, glycoproteins and certain enzymes determined. Several of the proteins displayed marked heterogeneity indicating the existence of protein variants and isoenzymes. Principal pI values for the enzymes assayed were as follows: xanthine oxidase, 7.35--7.55; NADH2: iodonitrotetrazolium reductase, less than 4.5; 5'-nucleotidase, 7.15--7.4; alkaline phosphatase, 5.4--5.7; phosphodiesterase, 4.6--4.8; gamma-glutamyl transpeptidase, 4.4--4.55. 3. Fractions after electrofocusing were analyzed by 'fused rocket' immunoelectrophoresis and crossed immunoelectrophoresis after separation in polyacrylamide gels containing sodium dodecyl sulphate. Major antigens of the membrane include xanthine oxidase and glycoproteins of apparent molecular weights 67 000, 49 500 and 46 000. The latter two components share common antigenic determinants and could not be separated by gel filtration, ion-exchange chromatography, lectin-affinity chromatography or preparative electrofocusing.
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PMID:Separation of the proteins of bovine milk-fat-globule membrane by electrofocusing with retention of enzymatic and immunological activity. 610 13

Reaction of peroxides with 5-deazaflavin bound to glucose oxidase, lactate oxidase, or D-amino acid oxidase results in the formation of 5-deazaflavin 4a, 5-epoxide. The reaction of D-amino acid oxidase with m-chloroperoxybenzoate is an exception since the reagent reacts rapidly with the protein moiety to form m-chlorobenzoate which then binds noncovalently near the unmodified coenzyme. Epoxide bound to glucose oxidase is converted to deazaFAD X X in a reaction similar to that observed previously with oxynitrilase and glycolate oxidase. With lactate oxidase the epoxide is quite stable in the absence of light. With D-amino acid oxidase, denaturation of the protein is accompanied by the release of the epoxide into solution where it decomposes in a manner similar to that observed with model epoxide compounds at neutral pH. Reaction of deazaFAD X X with phosphodiesterase and alkaline phosphatase yields deazariboflavin X X. The same compound has been formed in model studies by exposing 5-deazariboflavin 4a,5-epoxide to alkaline conditions. Structural studies indicate that this reaction involves contraction of the pyrimidine ring to yield 4-ribityl-6,7-dimethyloxazolo[ 4,5-b ]quinolin-2(4H)-one. Model reaction studies are consistent with a mechanism initiated by alkaline hydrolysis of the pyrimidine ring at position 4 followed by two additional steps which proceed at neutral pH. A similar mechanism for the enzyme reactions appears likely since analogous intermediates are detected in the glycolate oxidase and the model reactions. The results suggest that position 4 of the coenzyme in oxynitrilase, glycolate oxidase, and glucose oxidase must be accessible to solvent and that the protein moiety must facilitate the initial hydrolysis of the pyrimidine ring since the enzyme reactions occur at neutral pH. Failure to observe formation of deazaFMN X X with lactate oxidase is attributed, at least in part, to the inaccessibility of the pyrimidine ring to solvent.
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PMID:Reaction of enzyme-bound 5-deazaflavin with peroxides. Pyrimidine ring contraction via an epoxide intermediate. 613 30

Pyrophosphate, p-nitrophenyl phosphate and a variety of pyrimidine and purine nucleotides are hydrolyzed by the solubilized membrane-bound enzymes of the brush border plasma membrane of Hymenolepis diminuta. The pH optima (or ranges) for hydrolysis of substrates are 8.0 (pyrophosphate), 8.8 (p-nitrophenyl phosphate), 8.4-8.9 (nucleoside monophosphates), and 7.1-8.1 (nucleoside triphosphates); all substrates, with the exception of nucleoside triphosphates, have a higher affinity for the solubilized enzyme at pH 7.4 than at their optimal pH for hydrolysis. ATP is degraded completely by the enzyme preparation to adenosine and inorganic phosphate, but since neither ADP nor ATP accumulate in the incubation medium it is not known whether ATP hydrolysis involves the sequential hydrolysis of terminal phosphate groups. Isoelectric focusing and various chromatographic procedures (gel permeation, ion-exchange and hydrophobic interaction chromatography) fail to separate the alkaline phosphatase, phosphodiesterase, 5'-nucleotidase, adenosine triphosphatase and ribonuclease activities associated with the solubilized membrane preparation. Additionally, inhibitor studies indicate that only a single enzyme with low substrate specificity is involved in the hydrolysis of nucleotides, p-nitrophenyl phosphate, pyrophosphate and hexose phosphate esters. Purines and pyrimidines and their nucleosides interact with the active site, and in some instances activity of the enzyme is stimulated by an unknown mechanism.
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PMID:Nucleotide hydrolysis by solubilized membrane-bound enzymes of the brush border plasma membrane of Hymenolepis diminuta. 613 88

Plasma membranes of vertebrate lens fiber cells contain large numbers of gap junctions that may provide pathways for metabolic cooperation. Characterization of fiber cell gap junctions is thus necessary to understand this function. In this study, plasma membrane fractions were isolated from bovine lens according to established techniques, but without urea, detergents, or proteolytic enzymes. Electron microscopy indicated that isolated plasma membranes with gap junctions form double-membrane vesicles, and gap junctions comprised approximately 35% of the total membrane area in the crude fraction. These vesicles were impermeable to cationized ferritin, suggesting that they were sealed, and may be useful for permeability studies. Treatment of the crude fraction with 2.5% beta-mercaptoethanol or dithiothreitol caused reversible separation of junctional membranes, suggesting that disulfide bonds may be important in maintaining gap junction structure. Fractions with varying proportions of gap junctions were isolated using linear sucrose density gradient centrifugation. The proportional area of gap junction membrane versus total membrane in the various fractions ranged from 10% to at least 51%. The following plasma membrane enzymes were assayed in all fractions: Mg++-ATPase, Ca++-ATPase, alkaline phosphatase, phosphodiesterase, 5'-nucleotidase, and Na+, K+-ATPase. There was no correlation between enzyme activity and gap junction enrichment. This suggests that these enzymes are not associated with fiber cell gap junctions.
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PMID:Biochemical and structural characterization of membrane fractions from bovine lens. 613 51

Four types of 1-beta-D-arabinofuranosylcytosine (ara-C) conjugates with poly-L-glutamic acid (PLGA) or poly-N5-(2-hydroxyethyl)-L-glutamine (PHEG) were prepared in an attempt to enhance the efficacy of the drug in simple dosage schedules. The conjugates were made by linking ara-C to the carboxyl groups of PLGA directly at N-4 of ara-C (ara-C:PLGA) or indirectly through the 2-aminoethylphosphoryl or 6-aminohexylphosphoryl side chain which had been introduced to C-5' of ara-C, 1-[5'-(2-aminoethylphosphoryl)-beta-D-arabinofuranosyl]cytosine: PLGA [araCMP(C2):PLGA and 1-[5'-(6-aminohexylphosphoryl)-beta-D-arabinofuranosyl]cytosine:++ +PLGA, respectively, or made by converting the remaining carboxyl groups in the PLGA conjugates to the 2-hydroxyethylamide groups [ara-C:PHEG, ara-CMP(C2):PHEG, 1-[5'-(6-aminohexylphosphoryl)-beta-D-arabinofuranosyl]cytosine:++ +PHEG]. Studies in vitro showed that the conjugates had decreased cytotoxicity against L1210 cells when compared with that of ara-C. Studies in vivo showed that all of the conjugates, except ara-CMP(C2):PLGA, had a greater antitumor activity than did ara-C in L1210 tumor-bearing BALB/c X DBA/2 F, (hereafter called CD2F1) mice (inoculum, 1 X 10(5) cells i.p. on Day 0) which were treated by a single i.p. injection of either the conjugates or the control ara-C on Day 1. The largest antitumor activity [increased life span (ILS) 170%] was observed with a dosage of 50 mg (equivalent ara-C per kg) of ara-C:PHEG. When CD2F1 mice which had been inoculated i.p. with 1 X 10(5) L1210 cells were treated with an i.p. injection of 12.5 or 25 mg (equivalent ara-C per kg) of ara-C:PHEG daily for 5 days starting from Day 1, 2 of 5 mice survived more than 42 days, and the ILS of the remaining mice was 153 and 184%. The injections of 3.2 mg (equivalent ara-C per kg) of ara-C:PHEG showed a moderate antitumor activity with an ILS of 113% which was similar to the ILS (119%) found when unconjugated ara-C (400 mg/kg) was used to treat tumor-bearing mice. In in vitro release experiments, ara-C was released slowly from ara-C:PLGA at pH 7.4, and ara-CMP(C2):PLGA was chemically stable but cleaved by phosphodiesterase, acid phosphatase, and alkaline phosphatase to give mainly 1-beta-D-arabinofuranosylcytosine 5'-monophosphate.
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PMID:Antitumor activity of 1-beta-D-arabinofuranosylcytosine conjugated with polyglutamic acid and its derivative. 619 62

The chemical synthesis of the tital bridged trinucleoside diphosphates 3e and 3f along with the corresponding dinucleoside phosphates 3c and 3d is described. Bridged nucleosides 3a and 3b gave on treatment with triethyl orthoformate in the presence of p-toluenesulfonic acid in dimethylformamide the cyclic orthoesters 2a and 2b. Condensation of 2a and 2b with N,2',5'-O-triacetylcytidine 3'-phosphate (1) using dicyclohexylcarbodiimide in pyridine afforded after deblocking and chromatographic separation products 3c-f. The latter were readily degraded with pancreatic RNase, but 3c and 3e were completely resistant toward snake venom phosphodiesterase whereas 3d and 3f were digested to the extent of 65 and 43%, respectively. The major product of degradation of 3f with phosphodiesterase was compound 3d resulting from the combined action of phosphodiesterase and contaminating phosphomonoesterase. The results are explained in terms of stacking of terminal bridge nucleoside units in 3c-f. The implications of these findings for the function of snake venom phosphodiesterase are discussed.
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PMID:Synthesis of dicytidylyl-(3'-5')-1,2-di(adenosin-N6-yl)ethane and dicytidylyl-(3'-5')-1,4-di(adenosin-N6-yl)butane: covalently joined terminals of two transfer ribonucleic acids and their behavior toward snake venom phosphodiesterase. 624 71

A phosphodiesterase activity is shown to copurify with the plasma membrane fraction prepared by the two-phase partition method. The enrichment in phosphodiesterase parallels that of alkaline phosphatase, which is thought to be a typical membranous enzyme. Up to 66% of the phosphodiesterase activity can be solubilized by a treatment with 0.2% Triton X-100. Higher doses were ineffective in solubilizing more activity. Analysis by native gel electrophoresis showed that an activity extracted by 2 M NaCl migrated at the same position as 'soluble' phosphodiesterase of cytosolic or extracellular origin. In contrast, the Triton-solubilized enzyme had an apparently higher molecular weight. When subjected to charge shift electrophoresis on agarose gels in the presence of an ionic detergent, the Triton-solubilized phosphodiesterase displayed a hydrophobic character. This behaviour contrasts with that of 'soluble' phosphodiesterases, the electrophoretic mobility of which is unaffected by the presence of an anionic detergent. The hydrophobic character of the membranous enzyme was lost after gentle hydrolysis by papain.
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PMID:The hydrophobic character of the membrane-bound phosphodiesterase from Dictyostelium discoideum. 626 Feb 58

The intracellular distribution of phosphodiesterase [EC 3.1.4.17] induced by cyclic adenosine 3',5'-monophosphate (cAMP) in Dictyostelium discoideum was studied. When cAMP-treated cells were homogenized and fractionated according to the method of de Duve et al. ((1955) Biochem, J. 60, 604), the specific activity of phosphodiesterase was highest in the light mitochondrial fraction. Peaks of specific activities of alkaline phosphatase (marker enzyme of membrane) and catalase (marker enzyme of peroxisomes) also appeared in the same fraction as phosphodiesterase. However, after centrifugation of the light mitochondrial fraction in a sucrose density gradient, the activity of phosphodiesterase was clearly separated with that of catalase (density 1.19 g/ml) and showed three peaks at lower density (1.10, 1.13, 1.17 g/ml) with good reproducibility. Some parts (1.13, 1.17 g/ml) of the activity in the gradient overlapped with alkaline phosphatase activity, but in the density fraction of 1.10 g/ml the activity of alkaline phosphatase was hardly detectable. When the light mitochondrial fraction was treated with Emulgen 108, or sonicated, phosphodiesterase was more easily solubilized than alkaline phosphatase and catalase, and was found in supernate after centrifugation at 20,000 X g for 30 min. In order to distinguish the locations of the three enzymes, the supernatant of the light mitochondrial fraction treated with Emulgen 108 was subjected to charge shift electrophoresis. The electrophoretic mobilities of phosphodiesterase and catalase were unaffected by ionic detergent. However, alkaline phosphatase shifted towards the anode in the presence of anionic detergent (sodium deoxycholate), and shifted towards the cathode in cationic detergent (cetyltrimethylammonium bromide), relative to nonionic detergent (Emulgen 108) alone. Thus, some part of the phosphodiesterase induced by cAMP may be associated with the plasma membrane, but the remainder is localized in some kind of intracellular particle of lower density. Moreover, the association with the membrane or particle is more easily dissociated than that of alkaline phosphatase, and the liberated phosphodiesterase is rather hydrophilic.
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PMID:Intracellular localization of phosphodiesterase induced by cyclic adenosine 3',5'-monophosphate in Dictyostelium discoideum. 626 72

The products derived from the degradation of the sixteen possible diribonucleoside monophosphates (NpN') by Fusarium phosphodiesterase-phosphomonoesterase were analyzed by means of thin layer chromatography. The analysis showed that NpN' was first cleaved into nucleoside N and 5'-nucleotide pN', which was then dephosphorylated to yield nucleoside N'. The dephosphorylation was fast when N' was adenosine or cytidine but slow when N' was guanosine or uridine. The cleavage reaction was followed by measuring the increase of absorbance due to hyperchromicity, and the kinetic constants, Km and kcat, were determined for the sixteen dinucleoside phosphates. The Km value was higher, for a given N, when N' was a pyrimidine nucleoside than when N' was a purine nucleoside. For a given N', uridine as N gave the highest Km value and adenosine gave the lowest one. The kcat value was the highest, for a given N, when N' was cytidine. For a given N', uridine as N gave by far the lowest kcat value. These results can be interpreted in terms of two binding sites on the enzyme with different base preferences. Comparison of kcat/Km values suggested that the base of nucleoside N plays an important role in determining whether a dinucleoside phosphate is a good substrate of the enzyme. The dinucleoside phosphates with uridine as N were found to be particularly poor substrates of the enzyme.
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PMID:Mode of hydrolysis of diribonucleoside monophosphates by phosphodiesterase-phosphomonoesterase of Fusarium moniliforme. 627 68

The isolated, brush-border membrane of Hymenolepis diminuta contained an enzyme which hydrolyzed phosphodiester bonds. This enzyme appeared to be a Type I phosphodiesterase (E. C. 3.1.4.1) (produces nucleoside 5'-phosphates) and had no activity against synthetic, Type II phosphodiesterase substrates (mononucleotides substituted at the 3' position). The effects of various potential inhibitors of enzymatic activity, and cation requirements of this enzyme, demonstrated a distinct difference between the phosphodiesterase and alkaline phosphatase activities of the isolated, brush-border membrane. SDS-polyacrylamide gel electrophoresis of the isolated membrane preparation, followed by localization of phosphodiesterase activity in the gels, indicated the enzyme had a molecular weight of approximately 87,000. Thus, the phosphodiesterase activity represents a previously undescribed, membrane-bound enzyme of the brush-border of Hymenolepis diminuta.
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PMID:Type I phosphodiesterase in the isolated, brush-border membrane of Hymenolepis diminuta. 627 42


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