Use of p-nitrophenyl phosphate for assay of soil phosphatase activity

1969 ◽  
Vol 1 (4) ◽  
pp. 301-307 ◽  
Author(s):  
M.A. Tabatabai ◽  
J.M. Bremner
Keyword(s):  
Phosphatase Activity ◽  
Nitrophenyl Phosphate ◽  
Soil Phosphatase

scholarly journals Structural analysis of human dual-specificity phosphatase 22 complexed with a phosphotyrosine-like substrate

2015 ◽  
Vol 71 (2) ◽  
pp. 199-205 ◽  
Author(s):  
George T. Lountos ◽  
Scott Cherry ◽  
Joseph E. Tropea ◽  
David S. Waugh
Keyword(s):  
Crystal Structure ◽  
Structural Analysis ◽  
Phosphatase Activity ◽  
Small Molecule ◽  
Molecular Basis ◽  
Protein Tyrosine Phosphatases ◽  
Simple Method ◽  
Dual Specificity Phosphatase ◽  
Dual Specificity ◽  
Nitrophenyl Phosphate

4-Nitrophenyl phosphate (p-nitrophenyl phosphate, pNPP) is widely used as a small molecule phosphotyrosine-like substrate in activity assays for protein tyrosine phosphatases. It is a colorless substrate that upon hydrolysis is converted to a yellow 4-nitrophenolate ion that can be monitored by absorbance at 405 nm. Therefore, the pNPP assay has been widely adopted as a quick and simple method to assess phosphatase activity and is also commonly used in assays to screen for inhibitors. Here, the first crystal structure is presented of a dual-specificity phosphatase, human dual-specificity phosphatase 22 (DUSP22), in complex with pNPP. The structure illuminates the molecular basis for substrate binding and may also facilitate the structure-assisted development of DUSP22 inhibitors.

A reference method for measurement of alkaline phosphatase activity in human serum.

1983 ◽  
Vol 29 (5) ◽  
pp. 751-761 ◽  
Author(s):  
N W Tietz ◽  
C A Burtis ◽  
P Duncan ◽  
K Ervin ◽  
C J Petitclerc ◽  
...  
Keyword(s):  
Alkaline Phosphatase ◽  
Human Serum ◽  
Phosphatase Activity ◽  
Alkaline Phosphatase Activity ◽  
Zinc Sulfate ◽  
Metal Ion ◽  
Reference Method ◽  
Reaction Conditions ◽  
Factors Affecting ◽  
Nitrophenyl Phosphate

Abstract We present an official AACC reference method for the measurement of alkaline phosphatase, the culmination of optimization experiments conducted by a group of independent laboratories. The details of this method and evaluation of factors affecting the measurement are described. A metal ion buffer has been incorporated that maintains optimal and constant concentrations of zinc(II) and magnesium(II) ions. Final reaction conditions are: pH (30 degrees C), 10.40 +/- 0.05; 2-amino-2-methyl-1-propanol buffer, 0.35 mol/L; 4-nitrophenyl phosphate, 16.0 mmol/L; magnesium acetate, 2.0 mmol/L; zinc sulfate, 1.0 mmol/L; and N-(2-hydroxyethyl)ethylenediaminetriacetic acid, 2.0 mmol/L.

Modeling of soil phosphatase activity in land use ecosystems and topsoil layers in the Brazilian Cerrado

2018 ◽  
Vol 385 ◽  
pp. 182-188 ◽  
Author(s):  
Marcos V.M. Leite ◽  
Lenka Bobuľská ◽  
Suéllen P. Espíndola ◽  
Maria R.C. Campos ◽  
Lucas C.B. Azevedo ◽  
...  
Keyword(s):  
Land Use ◽  
Phosphatase Activity ◽  
Brazilian Cerrado ◽  
Soil Phosphatase

Effects of temperature on measurement of alkaline phosphatase activity.

1985 ◽  
Vol 31 (2) ◽  
pp. 185-190 ◽  
Author(s):  
W H Copeland ◽  
D A Nealon ◽  
R Rej
Keyword(s):  
Alkaline Phosphatase ◽  
Phosphatase Activity ◽  
Kinetic Properties ◽  
Porcine Kidney ◽  
Human Origin ◽  
Arrhenius Behavior ◽  
Nitrophenyl Phosphate ◽  
Activity Measurements ◽  
Michaelis Constants ◽  
Effects Of Temperature

Abstract We examined the effects of temperature on the activity and steady-state kinetic properties of alkaline phosphatase (EC 3.1.3.1). Purified isoenzymes from human liver, intestine, and placenta were used, as was human serum, and the enzyme from porcine kidney. Phosphatase activity was estimated by two different assay techniques. For all isoenzymes, apparent Michaelis constants for the substrate 4-nitrophenyl phosphate decreased with increased temperature; Km at 37 degrees C was typically half that determined at 25 degrees C. All enzymes of human origin exhibited similar linear Arrhenius relationships over the range examined, 20-37 degrees C (Ea of 30-36 kJ X mol-1). The porcine kidney enzyme obeyed an Arrhenius relationship that was slightly, but significantly, different from the isoenzymes of human origin. Temperature relationships based upon Arrhenius behavior and individual activity measurements are presented. For human alkaline phosphatases, they differed by no more than 10%.

scholarly journals Inhibitory effect of okadaic acid on the p-nitrophenyl phosphate phosphatase activity of protein phosphatases

1991 ◽  
Vol 275 (1) ◽  
pp. 233-239 ◽  
Author(s):  
A Takai ◽  
G Mieskes
Keyword(s):  
Phosphatase Activity ◽  
Okadaic Acid ◽  
Protein Phosphatase ◽  
Protein Phosphatases ◽  
Inhibitory Effect ◽  
Enzyme Concentration ◽  
Alkaline Phosphatases ◽  
Nitrophenyl Phosphate ◽  
Type 1 Phosphatase

The phosphatase activities of type 2A, type 1 and type 2C protein phosphatase preparations were measured against p-nitrophenyl phosphate (pNPP), a commonly used substrate for alkaline phosphatases. Of the three types of phosphatase examined, the type 2A phosphatase exhibited an especially high pNPP phosphatase activity (119 +/- 8 mumol/min per mg of protein; n = 4). This activity was strongly inhibited by pico- to nano-molar concentrations of okadaic acid, a potent inhibitor of type 2A and type 1 protein phosphatases that has been shown to have no effect on alkaline phosphatases. The dose-inhibition relationship was markedly shifted to the right and became steeper by increasing the concentration of the enzyme, as predicted by the kinetic theory for tightly binding inhibitors. The enzyme concentration estimated by titration with okadaic acid agreed well with that calculated from the protein content and the molecular mass for type 2A phosphatase. These results strongly support the idea that the pNPP phosphatase activity is intrinsic to type 2A protein phosphatase and is not due to contamination by alkaline phosphatases. pNPP was also dephosphorylated, but at much lower rates, by type 1 phosphatase (6.4 +/- 8 nmol/min per mg of protein; n = 4) and type 2C phosphatase (1.2 +/- 3 nmol/min per mg of protein; n = 4). The pNPP phosphatase activity of the type 1 phosphatase preparation shows a susceptibility to okadaic acid similar to that of its protein phosphatase activity, whereas it was interestingly very resistant to inhibitor 2, an endogenous inhibitory factor of type 1 protein phosphatase. The pNPP phosphatase activity of type 2C phosphatase preparation was not affected by up to 10 microM-okadaic acid.

Polyphosphate body and acid phosphatase localization in Nostoc sp.

1984 ◽  
Vol 30 (1) ◽  
pp. 8-15 ◽  
Author(s):  
John D. DuBois ◽  
Keith R. Roberts ◽  
Lawrence A. Kapustka
Keyword(s):  
Enzyme Activity ◽  
Acid Phosphatase ◽  
Phosphatase Activity ◽  
Acid Phosphatase Activity ◽  
Energy Stress ◽  
Nitrophenyl Phosphate ◽  
Carbon Compounds ◽  
Electron And Light Microscopy ◽  
Polyphosphate Bodies ◽  
Hydrolysis Of

Polyphosphate bodies and acid phosphatase activity were characterized in Nostoc sp. to determine if the hydrolysis of polyphosphate bodies occurs during dark (energy stress) periods. Electron and light microscopy were used to locate polyphosphate bodies. Acid phosphatase activity was measured using p-nitrophenyl phosphate as the substrate to determine net changes in the level of the enzyme activity. To induce energy stress, Nostoc sp. cells were kept in the dark for 72 h to deplete stored carbon compounds. Cells incubated in the light for 72 h (controls) showed acid phosphatase activity localized around the perimeter of polyphosphate bodies. When cells were incubated in the dark, acid phosphatase activity occurred throughout the polyphosphate body matrix. However, complete hydrolysis of the polyphosphate body did not occur and the rate of acid phosphatase activity was not affected.

Toxicity of As on Soil Neutral Phosphatase

2012 ◽  
Vol 260-261 ◽  
pp. 1195-1199
Author(s):  
Xin Ke ◽  
Ying Sun ◽  
Yun Zhang
Keyword(s):  
Phosphatase Activity ◽  
Contaminated Soils ◽  
Organic Phosphorus ◽  
Prostate Gland ◽  
Arsenic Content ◽  
Soil Bioremediation ◽  
Toxic Heavy Metals ◽  
Lower Organism ◽  
The Relationship ◽  
Soil Phosphatase

Toxicity of As on soil neutral phosphatase was studied through a series indoor incubation experiments. Results showed that the activity of soil neutral phosphatase was significantly inhibited by As pollution, and the peak inhibiting ratio was appeared at the sixth incubation day. While the activity of soil neutral phosphatase was also decreased with concentration of As increasing in the same incubation day. The relationship between the concentration of as and soil neutral phosphatase was fit by model ①y=c/(1+bx)and ②y=c(1+ax)(1+bx).The model② was more suitable to describe the relationship between the concentration of As and soil enzyme. This means the effect mechanism of As on the soil neutral phosphatase was part of inhibition. The Km of neutral phosphatase was increase by As concentration adding, while the Vmax was lowered. This kind of inhibition belongs to mix competition inhibition. Phosphatase widely exists in the biological world, from lower organism Escherichia coli, yeast to higher animals and plant tissues, body fluid and human liver, prostate gland are found to have phosphatase exist. It can catalyze the phosphate hydrolysis of elemental and inorganic phosphate release, important enzymes of biological phosphorus metabolism [1-2]. Soil phosphatase is an enzyme that has a major impact on agricultural production, producing an important role in the cycle of soil phosphorus. Research show that phosphatase in soil and aquatic systems of phosphorus, organic phosphorus pesticide contaminated soil bioremediation is very important, so it can be used as material for ecosystem beneficial or deleterious effects of indicator. Therefore, some can be as the instructions of the beneficial or harmful effects of ecosystem biology. Arsenic is widespread nature of toxic heavy metals often pollute the environment with the waste material, pesticides, fertilizers and other. According to the statistics in early 1990s, each year around the world due to human activities the importation of soil arsenic content of 0.52-1.2 million tons, its impact on soil ecosystem is one of the important topics. This experiment which proposed indoor simulation method analysis different concentrations of arsenic contaminated soils of neutral phosphatase activity, to explore the effects of different concentrations of arsenic on soil phosphatase activity of short-term toxicity effect. Analysis of the influence degree and duration, further analysis between the two possible mechanism, provides the basis for environmental protection and monitoring.

Responses of the phosphatase activity of the lichen Cladina rangiferina to various environmental factors including metals

1979 ◽  
Vol 57 (14) ◽  
pp. 1534-1540 ◽  
Author(s):  
I. Lane ◽  
K. J. Puckett
Keyword(s):  
Enzyme Activity ◽  
Environmental Factors ◽  
Phosphatase Activity ◽  
Enzyme Activities ◽  
Enzyme Concentration ◽  
Acidic Ph ◽  
Nitrophenyl Phosphate ◽  
Cladina Rangiferina ◽  
Reduced Activity ◽  
Menten Constant

The characteristics of phosphatase activity of Cladina rangiferina (L.) Harm, have been studied. Calculations of enzyme activities were based on the liberation of p-nitrophenol from p-nitrophenyl phosphate. The phosphatase activity was found to be linear both with increasing sample size (enzyme concentration) and increasing time, showed highest activity at acidic pH, and had a Michaelis–Menten constant of 8.9 × 10−3 M. The enzyme activity was maximal in the range 61 ± 10 °C, was independent of light, and was completely eliminated by boiling the thalli. Various cations and anions were tested for their effect; uranyl and vanadyl ions inhibited activity by 60% whereas copper, nickel, and silver enhanced activity by 10%. The anions biselenite, cyanide, fluoride, molybdate, phosphate, and vanadate all greatly reduced activity (≥ 50%). Phosphatase activity was demonstrated in other lichen species.

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