Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.3.8 (phytase)
 1,997 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The degradation of phytate (inositol hexaphosphate) in rapeseed meal diet not containing phytase activity was studied in 15 growing ileum-fistulated pigs. Stomach and small intestinal degradation and total gastrointestinal degradation were compared. The effect of addition of calcium carbonate to the rapeseed meal diet at two levels (9.2 and 18.5 g/kg diet) was investigated. A commercial barley-wheat-soybean diet with intrinsic phytase activity was used as reference. Phytate and its hydrolysis products in diets, ileal digesta and feces were determined by HPLC ion-pair chromatography. Hydrolysis of phytate in the stomach and small intestine was 35-45% in pigs fed the rapeseed meal diet independent of calcium addition, and 65% in pigs fed the reference diet. Total gastrointestinal degradation of phytate in pigs fed the rapeseed diet was 97, 77 and 42% (P < 0.001) when calcium intakes were 4.5, 9.9 and 15 g/d, respectively; total gastrointestinal degradation was 72% in pigs fed the reference diet. The intestinal phytate degradation pattern, when rapeseed diet was fed, indicated the activity of an unspecific phosphatase, whereas that of the reference diet indicated intrinsic dietary phytase activity. We conclude that dietary supplementation of calcium carbonate decreases the phytate degradation in the colon of pigs, but not in the stomach and small intestine.
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PMID:High dietary calcium level decreases colonic phytate degradation in pigs fed a rapeseed diet. 846 57

Of all the sources of phytase that have been studied (plant, animal, and microorganisms), the highest yields are produced by a wild-type strain A. niger NRRL 3135 (12.7 mg P/hr/ml = 6.8 microns P/ml/min = 113.9 nKat/ml) in a mineral salt medium in which total phosphate (4 mg %) is limiting for growth and cornstarch and glucose are the carbon sources. Synthesis of the enzyme is repressed by phosphate in the wild-type strain. Aspergillus niger NRRL 3135 produces two phytases one with pH optima at 2.5 and 5.5 (phyA) and one with an optimum at pH 2.0 (phyB). It also produces a pH 6.0 optimum phosphatase that has no phytase activity. These three glycoproteins have been purified to homogeneity, characterized, sequenced, and cloned. The sequences have been compared to each other, other phytases, and to known phosphatases. Their homology has been determined. The active sites of phytases show remarkable homology to the active site residues of the members of a particular class of acid phosphatase (histidine phosphatase). The most conserved sequence is RHGXRXP. Phytase has been covalently immobilized on Fractogel TSK HW-75 F and glutaraldehyde-activated silicate. It has been immobilized on agarose. Losses of activity have been noted on immobilization but these may be minimized by future research. It should be possible to commercially produce and recover penta-, tetra-, tri-, di-, and monoinositol phosphates using immobilized phytase if markets develop for those products. Phytase (phyA) from A. niger NRRL 3135 has been cloned into an A. niger glucoamylase producing strain CBS 513.88 using a construct that has a glucoamylae promoter and an A. niger NRRL 3135 leader sequence, and that is devoid of phosphate repression. The yield of the secreted enzyme was increased 52-fold above that of wild-type A. niger NRRL 3135. The bioengineered organism produces 270 microns P/ml/min (4500 nKat/ml) which is approximately 7.9 g/liter in the medium. The yield of the secreted enzyme was increased 1440-fold above that of wild type CBS 513.88. Commercial preparations of the cloned enzyme are available. Phytase (phyA) has been cloned into tobacco and canola. The enzyme is localized in the seed and expressed at high levels. Feeding of the seed to animals has made the phytin-P in the commercial diets available to the animals. The efficacy of feeding phytase to monogastric animals (poultry and swine) has been established. The amount of enzyme that is necessary to be added to commercial diets has been titred for broilers, layers, turkeys, ducks, and swine. The units of enzyme required are related to the phytin-P content in the diet. The use of the enzyme as a feed additive has been cleared in 22 countries. If phytase were used in the diets of all of the monogastric animals reared in the U.S., it would release phosphorus that has a value of $1.68 x 10(8) per year. The FDA has approved the enzyme preparation as GRAS. The effect of feeding phytase to animals enables assimilation of the P found in feed ingredients and diminishes the amount of phosphate in the manure and subsequently entering the environment. The effect of feeding phytase to animals on pollution has been quantitatively determined. If phytase were used in the diets of all of the monogastric animals reared in the United States, it would preclude 8.23 x 10(7) kg P from entering the environment.
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PMID:Phytase. 886 87

The characterization of the multiple inositol polyphosphate phosphatase (MIPP) is fundamental to our understanding of how cells control the signalling activities of 'higher' inositol polyphosphates. We now describe our isolation of a 2.3 kb cDNA clone of a rat hepatic form of MIPP. The predicted amino acid sequence of MIPP includes an 18 amino acid region that aligned with approximately 60% identity with the catalytic domain of a fungal inositol hexakisphosphate phosphatase (phytase A); the similarity encompassed conservation of the RHGXRXP signature of the histidine acid phosphatase family. A histidine-tagged, truncated form of MIPP was expressed in Escherichia coli and the enzymic specificity of the recombinant protein was characterized: Ins(1,3,4,5,6)P5 was hydrolysed, first to Ins(1,4,5,6)P4 and then to Ins(1,4,5)P3, by consecutive 3- and 6-phosphatase activities. Inositol hexakisphosphate was catabolized without specificity towards a particular phosphate group, but in contrast, MIPP only removed the beta-phosphate from the 5-diphosphate group of diphosphoinositol pentakisphosphate. These data, which are consistent with the substrate specificities of native (but not homogeneous) MIPP isolated from rat liver, provide the first demonstration that a single enzyme is responsible for this diverse range of specific catalytic activities. A 2.5 kb transcript of MIPP mRNA was present in all rat tissues that were examined, but was most highly expressed in kidney and liver. The predicted C-terminus of MIPP is comprised of the tetrapeptide SDEL, which is considered a signal for retaining soluble proteins in the lumen of the endoplasmic reticulum; the presence of this sequence provides a molecular explanation for our earlier biochemical demonstration that the endoplasmic reticulum contains substantial MIPP activity [Ali, Craxton and Shears (1993) J. Biol. Chem. 268, 6161-6167].
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PMID:Molecular cloning and expression of a rat hepatic multiple inositol polyphosphate phosphatase. 935 36

Phytases hydrolyze phytic acid to less phosphorylated myo-inositol derivatives and inorganic phosphate. A thermostable phytase is of great value in applications for improving phosphate and metal ion availability in animal feed, and thereby reducing phosphate pollution to the environment. Here, we report a new folding architecture of a six-bladed propeller for phosphatase activity revealed by the 2.1 A crystal structures of a novel, thermostable phytase determined in both the partially and fully Ca2+-loaded states. Binding of two calcium ions to high-affinity calcium binding sites results in a dramatic increase in thermostability (by as much as approximately 30 degrees C in melting temperature) by joining loop segments remote in the amino acid sequence. Binding of three additional calcium ions to low-affinity calcium binding sites at the top of the molecule turns on the catalytic activity of the enzyme by converting the highly negatively charged cleft into a favorable environment for the binding of phytate.
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PMID:Crystal structures of a novel, thermostable phytase in partially and fully calcium-loaded states. 1065 18

Two assays were conducted to study the evolution of rye and barley phosphatases (phytase and acid phosphatase) and the degradation of its substrates (inositol phosphate esters) during seed germination. In this manner we could obtain a low-phytate, endogenous phosphatase rich ingredient to be used in animal nutrition. In the first assay, the seeds were soaked for 1 and 14 h and germinated for 3 and 5 days with and without the addition of gibberellic acid (GA3). In the second assay, the seeds were soaked for 1 h and germinated for 1, 3, and 5 days with GA3. Phytase (up to 5739 and 3151 U x kg(-1)) and acid phosphatase (up to 18288 and 3151 U x g(-1)) activities, and IP6 (6.09 and 6.01 mg x g(-1)), IP5 (0.48 and 0.48 mg x g(-1)), and IP4 (0.13 and 0.06 mg x g(-1)) were detected in ungerminated rye and barley, respectively. The germination process caused a significant increase of Phy and AcPh activities in rye (up to 112 and 213%) and barley (up to 212 and 634%) and a reduction in the phytate phosphorus content (up to 84 and 58%, respectively). Phytate phosphorus content was affected only by soaking time in the case of rye. Finally, during the course of germination, IP6 and IP5 were rapidly degraded in rye (88 and 79%) and barley (67 and 52%), and IP4 was only a short-living intermediate, which was increased during hydrolysis and degraded to IP3. In conclusion, a marked increase of Phy and AcPh activities in rye and barley with a concomitant decrease in phytate phosphorus content and an increase in the content of lower inositol phosphates were observed during the rye and barley germination.
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PMID:Effect of several germination conditions on total P, phytate P, phytase, and acid phosphatase activities and inositol phosphate esters in rye and barley. 1145 53

A new de novo synthesis of the enantiomeric pair D-myo-inositol 1,2,4-trisphosphate and D-myo-inositol 2,3,6-trisphosphate is described. Starting from enantiopure dibromocyclohexenediol, several C2 symmetrical building blocks were synthesized which gave access to D-myo-inositol 1,2,4,5-tetrakisphosphate and D-myo-inositol 1,2,3,6-tetrakisphosphate. Exploiting the high regiospecificity of two partially purified phosphohydrolases from Dictyostelium, a 5-phosphatase and a phytase, the inositol tetrakisphosphates were converted enzymatically to the target compounds. Their potential to modulate the activity of Ins3,4,5,6P4 1-kinase was investigated and compared with the effects of D-myo-inositol 1,3,4-trisphosphate.
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PMID:Regiospecific phosphohydrolases from Dictyostelium as tools for the chemoenzymatic synthesis of the enantiomers D-myo-inositol 1,2,4-trisphosphate and D-myo-inositol 2,3,6-trisphosphate: non-physiological, potential analogues of biologically active D-myo-inositol 1,3,4-trisphosphate. 1159 6

Phosphatase activities associated with the intestinal brush border membrane (BBM) of the rat were examined histochemically in relation to the characteristic environment of the intestine, where luminal pH fluctuates drastically between alkaline and acid pH ranges. Special attention was given to intestinal alkaline phosphatase (IALP) and phytase on the BBM. Whole body fresh-frozen sections of young rats and their rapidly frozen and freeze-substituted small intestines, embedded in Technovit 7100, were processed for the histochemical demonstration of phosphatase activity at three different pH values (9.2, 7.3, and 5.2), representing the deviation of luminal pH in vivo. Either an azo-dye method or lead-salt method was employed using naphthol AS-MX phosphate and ATP as substrate, respectively. With the azo-dye method, intense phosphatase reactions were demonstrated along the BBM at all three pH ranges. Phosphatase reactions of the BBM at pH 9.2 and 7.3 were abolished by L(+)-phenylalanine, heat pre-treatment, and EDTA chelation although some reaction remained at pH 7.3 after the treatment with EDTA or L(+)-phenylalanine. Phosphatase reactions of the BBM at pH 5.2 were resistant to L(+)-phenylalanine, L(+)-tartrate, PCMB and EDTA chelation, implying that the characteristics of the enzyme responsible for phosphohydrolysis at acid pH values differed from those at higher pH values. The lead-salt method in which ATP was used as substrate revealed intense reactions--which were dependent on Mg++ and stimulated by Ca++ and resistant to L(+)phenylalanine--to be localized along the BBM at alkaline and neutral pH values, but not at acid pH values. In vitro experiments showed progressive hydrolysis of naphthol AS-MX phosphate by purified phytase at pH 5.2, in a dose-dependent manner, and suggested the possible involvement of phytase in the phosphatase reactions of the BBM at acid pH. These data indicate that the phosphatase reactions at alkaline and neutral pH values, associated with the BBM of the rat intestine, represent IALP and Mg++/ Ca++-ATPase, while those at acid pH appear to correspond to phytase activity, something which has not been demonstrated by histochemical methods despite the availability of extensive data based on biochemical analyses.
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PMID:Phosphatase activities of rat intestinal enterocytes and their relation to diverse luminal pH, with special references to the possible localization of phytase along the brush border membrane. 1183 8

Escherichia coli phytase is a phosphatase that catalyzes the hydrolysis of phytic acid into inorganic phosphate and myo-inositol. Two crystal forms of this enzyme were obtained in the presence of heavy metals. Crystal forms I and II were obtained with the heavy atoms CdCl(2) and HgCl(2) and diffracted to 1.5 A and 2.25 A resolution, respectively. Hg(2+) and Cd(2+) both acted as molecular bridge(s), linking and stabilizing E. coli phytase in the unit cell, and played a crucial role in the crystallization of phytase by bridging neighbouring symmetry related molecules.
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PMID:Heavy metal-mediated crystallization of Escherichia coli phytase and analysis of bridging interactions. 1214 14

Three phytase (EC 3.1.3.26) isoforms from the roots of 8-d-old maize (Zea mays L. var Consul) seedlings were separated from phosphatases and purified to near homogeneity. The molecular mass of the native protein was 71 kD, and the isoelectric points of the three isoforms were pH 5.0, 4.9, and 4.8. Each of the three isoforms consisted of two subunits with a molecular mass of 38 kD. The temperature and pH optima (40[deg]C, pH 5.0) of these three isoforms, as well as the apparent Michaelis constants for sodium inositol hexakisphosphate (phytate) (43, 25, and 24 [mu]M) as determined by the release of inorganic phosphate, were only slightly different. Phytate concentrations higher than 300 [mu]M were inhibitory to all three isoforms. In contrast, the dephosphorylation of 4-nitrophenyl phosphate was not inhibited by any substrate concentration, but the Michaelis constants for this substrate were considerably higher (137-157 [mu]M). Hydrolysis of phytate by the phytase isoforms is a nonrandom reaction. D/L-Inositol-1,2,3,4,5- pentakisphosphate was identified as the first and D/L-inositol-1,2,5,6-tetrakisphosphate as the second intermediate in phytate hydrolysis. Phytase activity was localized in root slices. Although phosphatase activity was present in the stele and the cortex of the primary root, phytase activity was confined to the endodermis. Phytate was identified as the putative native substrate in maize roots (45 [mu]g P g-1 dry matter). It was readily labeled upon supplying [32P]phosphate to the roots.
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PMID:Maize Root Phytase (Purification, Characterization, and Localization of Enzyme Activity and Its Putative Substrate). 1222 56

The production of phytase by three feed-grade filamentous fungi ( Aspergillus ficuum NRRL 3135, Mucor racemosus NRRL 1994 and Rhizopus oligosporus NRRL 5905) on four commonly used natural feed ingredients (canola meal, cracked corn, soybean meal, wheat bran) was studied in solid substrate fermentation (SSF). A. ficuum NRRL 3135 had the highest yield [15 IU phytase activity/g dry matter (DM)] on wheat bran. By optimizing the supplementation of wheat bran with starch and (NH(4))(2)SO(4), phytase production increased to 25 IU/g DM. Optimization was carried out by Plackett-Burman and central composite experimental designs. Using optimized medium, phytase, phosphatase, alpha-amylase and xylanase production by A. ficuum NRRL 3135 was studied in Erlenmeyer flask and tray SSF. By scaling up SSF from flasks to stationary trays, activities of 20 IU phytase activity/g DM were reproducibly obtained.
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PMID:Optimization of phytase production by solid substrate fermentation. 1271 56


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