A fundamental thermostable cyanide-sensitive phosphoenolpyruvate acid carboxylase in photosynthetic and other organisms

1971 ◽  
Vol 49 (4) ◽  
pp. 631-643 ◽  
Author(s):  
David Pan ◽  
E. Roy Waygood
Keyword(s):  
Heat Treatment ◽  
Zea Mays ◽  
Acid Phosphatase ◽  
Plant Species ◽  
Phosphoenolpyruvate Carboxylase ◽  
Deae Cellulose ◽  
Bundle Sheath ◽  
Evolutionary Significance ◽  
Optimum Temperature ◽  
Optimum Ph

A thermostable 'phosphoenolpyruvate carboxylase' has been isolated from leaves of Zea mays different from phosphoenolpyruvate carboxylase (EC. 4.1.1.31) in that its optimum pH is 5.4, it does not liberate orthophosphate during the reaction, and it is inhibited by cyanide. The enzymic reaction has an optimum temperature of 70–75C and has been purified through steps including acidification to pH 4.6, heat treatment to 50C, and DEAE-cellulose and Sephadex G-200 column chromatography. Three fractions were active in the Sephadex eluate, but only fraction III was free from a thermostable acid phosphatase which catalyzes the liberation of orthophosphate from the substrate and the end product which is suggested to be a C4 phosphocarbonyl compound, although phosphohydroxypyruvate appears by either spontaneous or enzymic decarboxylation. The enzyme is assayed by the formation of a phenyl-hydrazone at 325 nm. The enzyme is localized and tightly bound in both the parenchyma bundle sheath and mesophyll chloroplasts, which are free from the thermostable acid phosphatase. Similar concentrations of the enzyme have been found in all plant species tested including C3 plants, ferns, bryophytes, algae, fungi, and even in calf liver. The enzyme must have considerable evolutionary significance.

scholarly journals Acid phosphatases and ribonucleases from Dactylis glomerata seeds. I. Chromatographic and electrophoretic heterogeneity of the enzymes

2015 ◽  
Vol 47 (4) ◽  
pp. 441-453 ◽  
Author(s):  
Eleonora Wieczorek ◽  
Janina Wiśniewska ◽  
Bronisława Morawiecka
Keyword(s):  
Acid Phosphatase ◽  
Sodium Acetate ◽  
Specific Activity ◽  
Deae Cellulose ◽  
Dactylis Glomerata ◽  
Molecular Forms ◽  
Acid Phosphatases ◽  
Sodium Acetate Buffer ◽  
Optimal Activity ◽  
Optimum Ph

Acid phosphatase and ribonuclease extracted with 0.1 M sodium acetate buffer, pH 5.1 from Dactylis glomerata seeds, and partially purified by means of 70% ethanol precipitation showed electrophoretic and Chromatographic heterogeneity. After chromatography on DEAE-cellulose acid phosphatase and ribonuclease were separated into four peaks. Nonadsorbing acid phosphatase on DEAE-cellulose (peak I) was separated into four peaks on CM-cellulose. The highest activity (11 units/mg) was found in fraction b (acid phosphatase Ib). The enzyme was activated by Mg<sup>2+</sup>, Ca<sup>2+</sup>, Li<sup>+</sup>, Cs<sup>+</sup>, K<sup>+</sup> ions and inhibited by Cu<sup>2+</sup>, Zu<sup>2+</sup>, F<sup>-</sup> and Mo<sup>-6</sup> at optimum pH 5.0. Strong absorbing ribonuclease on DEAE-cellulose (peak IV) was further separated on G-200 Sephadex into two molecular forms: RN-asa1 and RN-ase2. Ribonuclease l, a thermolabile enzyme with specific activity 807 units/mg, showed an optimal activity at pH 4.8-5.1.

scholarly journals Purification of NADP-Isocitrate dehydrogenase from Red Kidney beans (Phaseolus vulgaris rogue)

2010 ◽  
Vol 4 (1) ◽  
pp. 61-72
Author(s):  
Mukaram Shikara ◽  
Hiba Muneer Al-Khafagi ◽  
Wasnaa H. Faris
Keyword(s):  
Phaseolus Vulgaris ◽  
Isocitrate Dehydrogenase ◽  
Deae Cellulose ◽  
Nicotinamide Adenine Dinucleotide Phosphate ◽  
Optimum Temperature ◽  
Ammonium Sulphate Precipitation ◽  
Kidney Beans ◽  
Optimum Ph ◽  
Murashige Skoog ◽  
Red Kidney Beans

Nicotinamide adenine dinucleotide phosphate-dependent isocitrate dehydrogenase (NADP+-IDH; EC 1.1.1.42) was extracted from red kidney beans (Phaseolus vulgaris.) after the beans were placed into Murashige-Skoog medium and incubated under continuous white light (110 μmol photon m−2 s−1), then filtered, centrifuged and the supernatant was used for purification. The enzyme purified using ammonium sulphate precipitation, DEAE-cellulose and Matrex Bio red A (dye-ligand-chromatography) techniques, and exhibits several bands through electrophoresis, with one band corresponds to the IDH activity. Km values for the enzyme was 55.71± 4.56 x 10-6M. The enzyme has an optimum pH at 8.5, and optimum temperature at 30°C. The enzyme can be stable at RT (about 25°C) for 180min, but the activity disappears at 400min. Enzyme activity appears to be independent of divalent metals in deionized water, but the addition of Mg+2 and Mn+2 by 4.5 and 2-folds respectively. The purified enzyme was injected into white rabbits to raise an antiserum against NADP+-IDH. The specificity of the antiserum was assayed by its ability to decrease the NADP+-IDH activity present in the extract. NADP+-IDH activity decreased when the extract was incubated with increasing volumes of the antiserum obtained.

Purification and Properties of Ribonuclease From Buffalo Milk Whey

1977 ◽  
Vol 40 (6) ◽  
pp. 375-377 ◽  
Author(s):  
AZZA A. ISMAIL ◽  
N. S. AHMED ◽  
M. A. KHORSHID
Keyword(s):  
Deae Cellulose ◽  
Ribonuclease A ◽  
Buffalo Milk ◽  
Optimum Temperature ◽  
Isolation And Identification ◽  
Milk Whey ◽  
Purification And Properties ◽  
Optimum Ph ◽  
Ribonuclease B

A procedure was developed for isolation and identification of ribonuclease from buffalo milk whey. Ribonuclease was precipitated with (NH4)2SO4 between 65 and 90% saturation. The precipitate was dissolved, dialyzed, and fractionated on DEAE-cellulose. Two ribonuclease-rich fractions were collected, i.e. ribonuclease A and B. Ribonuclease A had an optimum pH of 7 .0, and ribonuclease B had an optimum pH of 8.6. Both had an optimum temperature at 38 C. The ribonucleases in the purified state were unstable to heat and their activity decreased as the time of exposure increased. Both enzyme fractions were sensitive to inhibitors. NaCl and NaN3 were stimulatory for ribonuclease A, while ribonuclease B was stimulated only by NaCl.

Purification and characterization of the extracellular amylases of the yeast Schwanniomyces alluvius

1984 ◽  
Vol 30 (9) ◽  
pp. 1163-1170 ◽  
Author(s):  
B. Simões-Mendes
Keyword(s):  
Deae Cellulose ◽  
Culture Fluid ◽  
Soluble Starch ◽  
Ph Stability ◽  
Starch Hydrolysis ◽  
Heat Inactivation ◽  
Optimum Temperature ◽  
Enzyme Preparations ◽  
Optimum Ph

The extracellular amylolytic system of a strain of the yeast Schwanniomyces alluvius consists of an α-amylase, a glucoamylase, and probably a debranching enzyme. Crude enzyme preparations were obtained by fractionation of the culture fluid, at the stationary phase of growth, with isopropanol. Purification was carried out by DEAE-cellulose chromatography. The glucoamylase had the following properties: molecular weight (MW), 117 000 ± 2300; optimum temperature, 50 °C; optimum pH, 4.5; range of pH stability, pH 4–6; final product of starch hydrolysis, glucose; ΔH≠ and ΔS≠ of heat inactivation, 39747 cal∙mol−1 and 274.3 cal∙deg−1∙mo−1; Km(30 °C, pH 4.5) for soluble starch, 22.22 g∙L−1. The α-amylase had the following properties: MW, 62 000 ± 500; optimum temperature, 40 °C; optimum pH, 6.3; range of pH stability, pH 4–7; final product of starch hydrolysis, maltose and glucose; ΔH≠ and ΔS≠ of heat inactivation, 36594 cal∙mol−1 and 256.9 cal∙deg−1 mol−1; Km (40 °C, pH 5.5), 2.7 g∙L−1.

Trends in the pollination ecology of the Orchidaceae: evolution and systematics

1992 ◽  
Vol 70 (3) ◽  
pp. 642-650 ◽  
Author(s):  
Raymond Louis Tremblay
Keyword(s):  
Plant Species ◽  
Evolutionary Ecology ◽  
Evolutionary Significance ◽  
Evolutionary Trend ◽  
Pollen Transfer ◽  
Orchid Species ◽  
Pollinator Species ◽  
Plant Taxon ◽  
Pollinator Specialization ◽  
The Cost

Reduction in the number of pollinator species per plant species is a mechanism that may lower the cost of pollen transfer. Using efficient pollinators may have an evolutionary significance. It is hypothesized that an evolutionary trend from many pollinators to few pollinators per plant species should be observable when species from ancestral versus recently derived monophyletic taxon are compared. Three different orchid phylogenetic sequences are used; two of the phylogenies show a reduction in the number of pollinator species per orchid species from the most ancestral to the most recently derived subfamilies. The third classification did not show this trend. It is thus possible to observe macroevolution of pollinator specialization of a monophyletic plant taxon. Key words: evolution, pollination, systematics, Orchidaceae, evolutionary ecology.

Immobilization and Characterization of Tannase and its Haze-removing

2009 ◽  
Vol 15 (6) ◽  
pp. 545-552 ◽  
Author(s):  
Erzheng Su ◽  
Tao Xia ◽  
Liping Gao ◽  
Qianying Dai ◽  
Zhengzhu Zhang
Keyword(s):  
Kinetic Parameter ◽  
High Activity ◽  
Green Tea ◽  
Storage Stability ◽  
Residual Activity ◽  
Black Tea ◽  
Optimum Temperature ◽  
Oolong Tea ◽  
Optimum Ph

Tannase was effectively immobilized on alginate by the method of crosslinking-entrapment-crosslinking with a high activity recovery of 76.6%. The properties of immobilized tannase were investigated. Its optimum temperature was determined to be 35 ° C, decreasing 10 °C compared with that of free enzyme, whereas the optimum pH of 5.0 did not change. The thermal and pH stabilities of immobilized tannase increased to some degree. The kinetic parameter, Km, for immobilized tannase was estimated to be 11.6 × 10-4 mol/L. Fe2+ and Mn2+ could activate the activity of immobilized tannase. The immobilized tannase was also applied to treat the tea beverage to investigate its haze-removing effect. The content of non-estern catechins in green tea, black tea and oolong tea increased by 52.17%, 12.94% and 8.83%, respectively. The content of estern catechins in green tea, oolong tea and black tea decreased by 20.0%, 16.68% and 5.04%, respectively. The anti-sediment effect of green tea infusion treated with immobilized tannase was significantly increased. The storage stability and reusability of the immobilized tannase were improved greatly, with 72.5% activity retention after stored for 42 days and 86.9% residual activity after repeatedly used for 30 times.

Temperature-induced alanine oxidation in a psychrotrophic Pseudomonas

1975 ◽  
Vol 21 (12) ◽  
pp. 2028-2033
Author(s):  
Prince K. Zachariah ◽  
John Liston
Keyword(s):  
Heat Treatment ◽  
Enzyme Synthesis ◽  
Optimum Temperature ◽  
Temperature Optimum ◽  
Ph Optimum ◽  
Sephadex Column ◽  
Oxidase System ◽  
Psychrotrophic Bacteria ◽  
Induction Of Enzymes ◽  
Elution Fraction

A psychrotrophic pseudomonad isolated from iced fish oxidized alanine at temperatures close to 0 °C and grew over the range 0 °C–35 °C. The rate of oxidation of alanine, measured manometrically, by cells grown at 2 °C was lower than that of cells grown at 22 °C. However, the consumption of oxygen after heat treatment at 35 °C for 35 min was reduced considerably by 2 °C grown cells. Alanine oxidase activity was tested in an extract from cells grown at 2 °C and 22 °C with alanine as the sole carbon, nitrogen, and energy source. Cells grown at 2 °C produced an alanine oxidase with a temperature optimum of 35 °C and pH optimum of 8, which lost about 80% activity by heat treatment at 40 °C for 30 min. There was no change in activity after dialysis at pH 7, 8, or 9. Extracts from cells grown at 22 °C contained an alanine oxidase system with an optimum temperature of 45 °C, a pH optimum above 8, and only about 30% reduction of activity after heat treatment. This enzyme activity was concentrated in the 0.5 M elution fraction from a Sephadex column, and dialysis reduced the activity at pH 7 and 8. Mesophilic enzyme synthesis apparently started around a growth temperature of 10 °C.The crude alanine oxidase systems of Pseudomonas aeruginosa derived from cells grown at 13 °C and 37 °C had a common optimum temperature of 45 °C. These data suggest that one mechanism of psychrophilic growth by psychrotrophic bacteria may be the induction of enzymes with low optimum temperatures in response to low temperature conditions.

Tonoplast and plasma membrane ATPases from maize lines of high or low vigour

1992 ◽  
Vol 2 (2) ◽  
pp. 105-111 ◽  
Author(s):  
S. Sánchez-Nieto ◽  
R. Rodríguez-Sotres ◽  
P. González-Romo ◽  
I. Bernal-Lugo ◽  
M. Gavilanes-Ruíz
Keyword(s):  
Zea Mays ◽  
Plasma Membrane ◽  
Acid Phosphatase ◽  
Atpase Activity ◽  
Kinetic Analysis ◽  
Atp Hydrolysis ◽  
Membrane Atpases ◽  
Substrate Concentrations ◽  
Tonoplast Atpase ◽  
Specific Inhibitors

AbstractThe effectiveness of ATPase in germinated seed may play an important role in the vigour of germination. The activities of tonoplast and plasma membrane ATPases in two maize (Zea mays L.) lines with different vigour of germination were determined. ATP hydrolysis was measured in microsomal fractions from coleoptiles along with the responses to specific inhibitors for the plasma membrane, tonoplast and mitochondrial ATPases as well as for acid phosphatase. Nitrate-sensitive ATPase activity was 1.5–3.0 times lower in the low-vigour line than in the high-vigour line. Kinetic analysis of ATP hydrolysis at different substrate concentrations revealed the existence of two enzymes in the microsomal fractions of the two lines. The Vmax of enzyme 1 in the low-vigour line was a third of that in the high-vigour line. This enzyme was identified as the nitrate-sensitive or tonoplast ATPase on the basis of measurements of ATP hydrolysis in the presence of specific inhibitors at high (8.12mm) and low (0.77mm) ATP concentrations.

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