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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)

A phytase (EC 3.1.3.8) was extracted from rat intestinal bacterium, Klebsiella Sp. No. PG.-2, and purified 50-fold by ammonium sulphate fractionation, ion-exchange chromatography and gel filtration. The enzyme is inducible in nature. The pH optimum was at 6.0 for all the inositol phosphates studied and this characterized the enzyme as an acid phosphohydrolase. Of a range of potential substrates tested, only p-nitrophenyl phosphate alongwith the inositol phosphates was hydrolyzed. It exhibits a Km of 2.0 mM; temperature optimum of 37 degrees C and energy of activation 9,120 cal/mole for all the inositol phosphates studied. The activity was inhibited by Ag2+, Hg2+, Cu2+, fluoride and high substrate concentration.
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PMID:Phytase from Klebsiella Sp. No. PG-2: purification and properties. 216 21

Aspergillus carneus (Van Tiegh) Blockwitz synthesized moderate quantities of phytase in culture filtrate. Maximal enzyme yield was obtained after eight days incubation statically in a medium containing sucrose and K2HPO4 in a C/P ratio of 591.8/1 with 0.1% corn steep liquor (CSL) as the sole source of nitrogen. Substitution of (NH4)2SO4 with certain amino acids decreased phytase yields significantly. Addition of fish or soybean meal to the nitrogen-free medium failed to enhance phytase production. The enzyme was purified about 43-fold from the culture filtrate by precipitation with acetone, gel filtration through Sephadex G-75 and ion exchange chromatography on DEAE-cellulose. Activity of both crude and purified phytase was influenced greatly by changing of pH and reaction temperature: maximum activity of the crude enzyme occurred at 35 degrees C and pH 5.6, whereas that of the purified preparation at 40 degrees C and pH 5.6. The pure enzyme was found stable between pH 5.6-6.2. About 68% of the enzyme activity was lost by heating at 45 degrees C for 60 min. The pure phytase retained its activity over a long period when stored at 4 degrees C.
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PMID:Biosynthesis, purification and some properties of extracellular phytase from Aspergillus carneus. 217 69

Soybean acid phosphatase (orthophosphoric-monoester phosphohydrolase, EC 3.1.3.2) was completely separated from phytase (EC 3.1.3.8) isolated from cotyledons of germinating seeds and purified to homogeneity. A four-step purification regimen consisting of ammonium sulfate fractionation, and ion-exchange, affinity, and chromatofocusing gel chromatographies was employed to achieve a homogeneous preparation. Acid phosphatase activity appeared as a major band of the three forms of acid phosphatase identified on native gels. The purified enzyme had a molecular weight of 53,000 when electrophoresed on 8% sodium dodecyl sulfate-polyacrylamide gel electrophoresis and a molecular weight of 53,000 from its mobility in a Fracto-gel TSK HW-50F gel permeation column. The molar extinction coefficient of the enzyme at 278 nm was estimated to be 4.2 X 10(4) M-1 cm-1. The isoelectric point of the protein, as revealed by chromatofocusing, was about 6.7. The optimal pH for activity, like other plant acid phosphatases, was 5.0. While the enzyme failed to accommodate phytate as a substrate, the enzyme did exhibit a broad substrate selectivity. The affinity of the enzyme for p-nitrophenyl phosphate was high (Km = 70 microM), and activity was competitively inhibited by orthophosphate (Ki = 280 microM). The estimated catalytic turnover number (Kcat) of the enzyme for p-nitrophenyl phosphate was about 430 per second. Although the purified enzyme was stable at 0 degrees C and exhibited maximum catalytic activity at 60 degrees C, thermal inactivation studies indicated that the enzyme lost 100% activity after treatment at 68 degrees C for 10 min.
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PMID:Purification and characterization of acid phosphatase from cotyledons of germinating soybean seeds. 282 34

Low-phytate wheat bran was produced by enzymatic hydrolysis and extraction. Rat bioassay methods were utilized to determine bioavailability of iron and zinc in the low-phytate brans and to study the effect of dietary phytate/zinc molar ratio on zinc bioavailability when the phytate source was bran. Endogenous phytase activity hydrolyzed 80-100% of the phytate when wheat bran was incubated in water overnight. The relative biological values of the iron in raw bran and phytate-free bran were 98 and 113, respectively, compared to 100 for ferrous ammonium sulfate in a hemoglobin repletion assay. Low-phytate brans with phytate/zinc molar ratios of 8 or less were equivalent to zinc sulfate as dietary sources of zinc for growth of rats. Rats fed diets that contained wheat bran with zinc sulfate added to reduce the dietary phytate/zinc molar ratio from 40 or 50 to 20 grew at the same rate as rats fed a phytate-free diet, but femur zinc values were lower than those in the reference group. Gel filtration chromatography of extracts of raw and low-phytate brans suggested that zinc might be associated with phytate in wheat bran.
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PMID:Bioavailability to rats of iron and zinc in wheat bran: response to low-phytate bran and effect of the phytate/zinc molar ratio. 625 2

Differential agar media for the detection of microbial phytase activity use the disappearance of precipitated calcium or sodium phytate as an indication of enzyme activity. When this technique was applied to the study of ruminal bacteria, it became apparent that the method was unable to differentiate between phytase activity and acid production. Strong positive reactions (zones of clearing around microbial colonies) observed for acid producing, anaerobic bacteria, such as Streptococcus bovis, were not corroborated by subsequent quantitative assays. Experimentation revealed that acidic solutions generated false positive results on the selected differential medium. Empirical studies undertaken to find a solution to this limitation determined the false positive results could be eliminated through a two step counterstaining treatment (cobalt chloride and ammonium molybdate/ammonium vanadate) which reprecipitates acid solubilized phytate. This report discusses the application of the developed two step counterstaining treatment for the screening of phytase producing ruminal bacteria as well as its use in phytase zymogram assays.
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PMID:A novel staining method for detecting phytase activity. 1057 3

Phytases produced by numerous microorganisms and plants degrade phytic acid that has chelated with metal ions in food and feed. It is important to study phytase for the role of metal ions in nutrition of animals and humans as well as in the reduction of organic phosphate content of aqueous environment. This article reports on solid-state fermentation of phytase from a new substrate of cassava dregs. Large quantities of cassava dregs are produced in tropical areas as a byproduct of cassava starch processing. Protein and inorganic salts were found to be low in cassava dregs. Cassava dregs could be employed for phytase synthesis after the addition of a nitrogen source and mineral salts. Ammonium nitrate was the best nitrogen source among the nitrogen sources investigated, including beef extract, yeast extract, urea, ammonium nitrate, sodium nitrate, and ammonium sulfate. Sodium dodecyl sulfate promoted phytase production from cassava dregs. A maximum phytase yield of 6.73 U/g of dry mass was obtained. The obtained phytase was stable at feed-processing temperature, since 70% of initial enzyme activity was maintained after 30 min of treatment at 75 degrees C.
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PMID:Solid-state fermentation of phytase from cassava dregs. 1196 5

Modification of hemoglobin using inositol tetrakisphosphate (IP(4)) can improve the oxygen affinity of hemoglubin. Phytase was extracted from wheat bran and purified by ammonium sulphate fractionation, followed by Sephadex G-50 gel filtration and Mono Q chromatography. The purified phytase was used for hydrolysis of phytic acid under controled conditions. IP(4), as a major composition of the hydrolysate, was further purified on resin 714 column. The purified IP(4) was oxidized by periodate to obtain dialdehyde-IP(4). By the reaction between the aldehyde group of IP(4) derivative and the amino group of porcine hemoglobin (pHb), pHb-IP(4) conjugate was formed and was found to have better ability of O(2) binding and releasing than the native hemoglobin.
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PMID:Preparation of Inositol Tetrakisphosphate and Its Application in Modification of Porcine Hemoglobin. 1211 Sep 9

Five isonitrogenous diets (1-5) with 40% protein using oilcakes as protein sources were formulated and fed to Cirrhinus mrigala fingerlings maintained both under laboratory and field conditions. Water soaking of oilcakes for 24 h before incorporation in the diets helped in the reduction of antinutrient factors (phytase and tannins). Live weight gain in fish fingerlings fed on a diet containing groundnut oilcake (GNOC) was significantly (P < 0.05) enhanced in comparison to the other dietary treatments when examined at the end of a feeding schedule. Laboratory studies have further revealed that APD, PER, GPR and GER values were significantly (P < 0.05) enhanced, while those of feed conversion ratio were significantly (P < 0.05) reduced in fish fed on diet 1 containing GNOC. An analysis of water samples collected at two hourly interval from the aquaria revealed low levels of total ammonia (N-NH4+) excretion and reactive phosphate (O-PO4) production in fish fed on diet 1. Proximate carcass composition also revealed high accumulation of protein, fat, energy and phosphorus in fingerlings fed on a diet containing GNOC. Even in field studies a significant (P < 0.05) increase in mean fish weight gain and specific growth rate (SGR% d(-1)) was observed in fingerlings fed on diet 1, followed by canola (2), sunflower (3), mustard oilcake (4) and sesame (5). Water and sediment quality characteristics also correlated well with fish growth.
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PMID:Oilcakes as protein sources in supplementary diets for the growth of Cirrhinus mrigala (Ham.) fingerlings: laboratory and field studies. 1268 72

Phytases catalyze the hydrolysis of phytic acid (myo-inositol hexakisphosphate), the most abundant inositol phosphate in cells. Phytases are of great commercial importance because their use as food and animal feed supplement has been approved by many countries to alleviate environmental and nutritional problems. Although acid phytases have been extensively studied, information regarding alkaline phytases is limited. Alkaline phytases with unique catalytic properties have been identified in plants, however, there is no report on the purification or structural properties. In this paper, we describe the purification of alkaline phytase from plant tissue. The purification was challenging because of contamination from non-specific phosphatases and acid phytases and low endogenous concentration. The purification of alkaline phytase from pollen grains of Lilium longiflorum involved selective precipitation by heat and ammonium sulfate followed by anion exchange and chromatofocusing chromatography and, finally, gel electrophoresis. Alkaline phytase was purified approximately 3000-fold with an overall recovery of 4.2%. The native molecular mass was estimated to be in the range of 118+/-7 kDa by Ferguson plot analysis and Mr of denatured protein in the range of 52-55 kDa by SDS-PAGE suggesting that the enzyme is a homodimer. Separation by 2-D gel and matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometric analysis of separated proteins indicates the presence of multiple mass and charge isoforms with pI values between 7.3 and 8.3. To our knowledge, this is the first alkaline phytase to be purified from plant sources. The unique properties suggest that the enzyme has the potential to be useful as a feed and food supplement.
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PMID:Alkaline phytase from Lilium longiflorum: purification and structural characterization. 1619 25

Phytase (myo-inositol-1,2,3,4,5,6-hexakisphosphate phosphohydrolase, EC 3.1.3.26) catalyses the stepwise hydrolysis of phytic acid (myo-inositol hexakisphosphate). Phytases are of great commercial importance due to their usage as supplement of food and animal feed, which can cater to nutrition demands and alleviate environmental problems, has been approved by many countries. Although acid phytases have been extensively studied, information regarding the phytases from Citrobacter is limited. In the work presented, a phytase was separated from Citrobacter freundii. After steps of electrophoretic homogeneity by successive ammonium sulfate between 60% and 80% saturation precipitation, DEAE-Sepharose ion-exchange chromatography and gel filtration through Superdex HR 10/30, final gel elution resulted in a 41.3-fold purification and yield of 9.3%. Gel elution is an effective method to purify the protein which contaminated with a few other proteins. The purified preparations were used in subsequent characterization studies. Based on SDS-PAGE analysis, the molecular weight of the purified phytase was calculated to be approximately 45.0kDa in monomeric form. The pure enzyme has an optimum pH of 4.0 to approximately 4.5. It was found stable between pH5.0 to approximately 7.0, about 90% of the enzyme activity was retained at 37 degrees C for 60min. The phytase has an optimum temperature of 40 degrees C which was lower than that of other phytases from Aspergillus or E. coli (average 50 to approximately 60 degrees C) and was close to the temperature of gastrointestinal tract in animals (37 to approximately 40 degrees C). Thus the enzyme is a promising candidate for animal feed applications. Activity of the purified phytase was influenced by changing the reaction temperature. Data showed that the enzyme retained its activity over a long period when stored at 4 degrees C, whereas thermal inactivation studies indicated that the enzyme lost 100% activity after treatment at 60 degrees C for 4min. The Km values of the phytase for dodecasodium phytate at 37 degrees C was 0.85nmol/L with a Vmax 0.53IU/(mg x min). Phytase activity was strongly inhibited by SDS, Zn2+ and moderately inhibited by Cu2+, Cr3+, Fe2+ and Fe3+. Activity was not significantly affected by EDTA, K+, Mg2+ and Ca2+. The phytase has excellent resistance to trypsin, but not pepsin. The N-terminal amino acids sequence of the phytase protein was determined as QCAPEGYQLQQVLMM which exhibited about 80% homology to Glucose-1-phosphatases from E. coli, Shigella flexneri and Salmonella, whereas it did not show apparent sequence similarity with any other phytase listed in the databases. Initial characterization of the purified enzyme suggested that it is a potential candidate for use as an animal feed supplement.
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PMID:[Purification and properties of Citrobacter freundii phytase]. 1657 82


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