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.

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.

scholarly journals Croatian produced unifloral honey characterized according to the protein and proline content and enzyme activities

2016 ◽  
Vol 60 (1) ◽  
pp. 39-48 ◽  
Author(s):  
Ivana Flanjak ◽  
Ivica Strelec ◽  
Daniela Kenjerić ◽  
Ljiljana Primorac
Keyword(s):  
Enzyme Activity ◽  
Acid Phosphatase ◽  
Protein Content ◽  
Glucose Oxidase ◽  
Phosphatase Activity ◽  
Enzyme Activities ◽  
Acid Phosphatase Activity ◽  
Black Locust ◽  
Proline Content ◽  
A Minor

Abstract In honey, the content of proteins, including the enzymes, is relatively low and has a minor nutritive significance. On the other hand, the proteins, including the enzymes, are usually used as honey quality evaluation parameters. This is because protein content and enzyme activities vary regarding the botanical origin of the honey. Since the results of protein content, glucose-oxidase, and acid phosphatase, for honeys produced in Croatia, are not available, four of the most abundant honey types produced in Croatia (black locust, sage, chestnut, and honeydew honey) are characterised according to the protein and proline content and enzyme activities. The characterisation was done to determine specificities and contribute to the characterisation of unifloral honeys. Dark honey types (honeydew and chestnut honey) had a higher proline content, and diastase, invertase, and glucose-oxidase activity than lighter sage and black locust honey. Black locust honey has a naturally low enzyme activity and showed the highest acid phosphatase activity among the analysed honey types, while honeydew honey, otherwise known to possess high proline content and enzyme activity, had a low protein content comparable to black locust honey. Statistically significant correlations were obtained between all analysed parameters, with the exception of acid phosphatase activity.

scholarly journals CYTOPHOTOMETRIC DETERMINATION OF ALKALINE PHOSPHATASE ACTIVITY OF INDIVIDUAL NEUTROPHILIC LEUKOCYTES WITH A BIOCHEMICALLY CALIBRATED MODEL SYSTEM

1968 ◽  
Vol 16 (11) ◽  
pp. 693-706 ◽  
Author(s):  
M. VAN DER PLOEG ◽  
P. VAN DUIJN
Keyword(s):  
Alkaline Phosphatase ◽  
Enzyme Activity ◽  
Phosphatase Activity ◽  
Alkaline Phosphatase Activity ◽  
Azo Dye ◽  
Enzyme Concentration ◽  
Model System ◽  
Coupling Method ◽  
Dye Coupling ◽  
Neutrophilic Leukocytes

A model system consisting of polyacrylamide films into which cell sonicate is incorporated was applied to investigate quantitative aspects of the cytochemical azo dye coupling method for alkaline phosphatase activity in neutrophilic leukocytes. The films were assayed in media containing naphthol AS-MX phosphate and 4-aminodiphenylamine diazonium sulfate. Optimal reaction conditions under which proportionality existed between the amount of reaction product and incubation time or enzyme concentration were determined. Since the enzyme activity in the films could also be measured chemically, using disodium phenyl phosphate as a substrate, a direct relation between the cytochemical and biochemical activity could be established. The azo dye coupling method was applied for the quantification of the enzyme activity. Air-dried microscopic preparations of exudate neutrophils were stained and the amount of dye formed in the cytoplasm of individual leukocytes was measured with a cytospectrophotometer based on the two-wavelength principle. By reference to the relation between biochemical and cytochemical activities in the model films, the cytochemically determined enzyme activity could be expressed in biochemical units. Independently, the average alkaline phosphatase activity per cell was determined by direct biochemical assay of the leukocyte suspension. The results showed that about 60% of the biochemically assayed activity in sonicate was demonstrated in the cell preparations by the cytochemical staining method. The discrepancy between the results of the two methods is discussed. The applicability of the model system for elucidation of the relationship between the results of cytochemical and biochemical enzyme determination in cells is stressed.

Effects of Hebeloma arenosa and phosphorus fertility on root acid phosphatase activity of red pine (Pinus resinosa) seedlings

1991 ◽  
Vol 69 (2) ◽  
pp. 380-383 ◽  
Author(s):  
Janet MacFall ◽  
Steven A. Slack ◽  
Jaya Iyer
Keyword(s):  
Enzyme Activity ◽  
Acid Phosphatase ◽  
Phosphatase Activity ◽  
Acid Phosphatase Activity ◽  
Fungal Growth ◽  
Pinus Resinosa ◽  
Ectomycorrhizal Fungus ◽  
Red Pine ◽  
Nitrophenyl Phosphate

The ectomycorrhizal fungus Hebeloma arenosa Burdsall, MacFall & Albers was assayed for surface-accessible acid phosphatase activity in vitro on roots of red pine (Pinus resinosa Ait.) seedlings. Hebeloma arenosa was grown in defined liquid media containing 0, 17, 34, 68, or 136 mg/L phosphorus for 4 weeks. When assayed for acid phosphatase activity with p-nitrophenyl phosphate, 7.3 μmol of orthophosphate were released per gram dry weight of fungal tissue. There was no effect of added P on enzyme activity, excluding the treatment with no added P in which there was negligible fungal growth. Red pine seedlings were grown in Sparta loamy fine sand amended with 0, 17, 34, 68, or 136 mg/kg P as superphosphate, with and without H. arenosa inoculum. Mycorrhizal roots had greater enzyme activity than nonmycorrhizal roots of seedlings grown in similarly P-amended soil. This was determined by the following three assays: orthophosphate release from two salts of myoinosital hexaphosphate (Na and KMg) and from p-nitrophenyl phosphate. It is suggested that greater acid phosphatase activity by roots mycorrhizal with H. arenosa is one mechanism for improved P nutrition through the formation of a pool of P released from sources unavailable for direct intake.

An Automated Method for Measuring Creatine Phosphokinase Activity in Serum

1970 ◽  
Vol 16 (4) ◽  
pp. 294-299 ◽  
Author(s):  
Ronald K Wright ◽  
Roy L Alexander
Keyword(s):  
Enzyme Activity ◽  
Enzyme Activities ◽  
Creatine Phosphokinase ◽  
Creatine Phosphate ◽  
Enzyme Concentration ◽  
Magnesium Ion ◽  
Manual Method ◽  
Automated Method ◽  
Automated Procedures

Abstract We describe a procedure for automating the determination of creatine phosphokinase (CPK) activity in serum by use of the AutoAnalyzer. Enzyme activity is determined by measuring the creatine phosphate formed from the CPK-catalyzed reaction of creatine with adenosine triphosphate (ATP). The sensitivity of the automated procedure was comparable to that of the manual method. By use of the most favorable concentrations of creatine, ATP, and magnesium ion in the substrate, a linear relationship was obtained between enzyme concentration and enzyme activities up to 600 mU, representing a sixfold improvement over that obtained by the manual method. The degree of correlation between results obtained by the manual and automated procedures is shown.

scholarly journals Early changes in enzyme activity during the development of and recovery from vitamin D deficiency in the rat

1966 ◽  
Vol 20 (3) ◽  
pp. 413-420 ◽  
Author(s):  
Elisabeth M. Cheesman ◽  
Alice M. Copping ◽  
Patricia M. Prebble
Keyword(s):  
Vitamin D ◽  
Enzyme Activity ◽  
Vitamin D Deficiency ◽  
Phosphatase Activity ◽  
Ribonucleic Acid ◽  
Enzyme Activities ◽  
Acid Content ◽  
Leucine Aminopeptidase ◽  
Recovery Period ◽  
Aminopeptidase Activity

1. Rats were given a rachitogenic diet with or without vitamin D for 1, 2, 3 or 4 weeks, and vitamin D was administered to a group of rats which had been deprived of vitamin D for 3 weeks. 2. Changes in enzyme activities of some tissues were followed histochemically. In general, changes in parathyroid preceded those in gut and kidney. Leucine aminopeptidase activity rose in the parathyroid, kidney and jejunum of the vitamin D-deficient rat as did the ribonucleic acid content of the parathyroid and the dehydrogenase activity of the gut. In contrast, the phosphatase activity of the gut and kidney fell in the vitamin D-deficient rat. All these changes were reversed during the 1-week recovery period. 3. A slower response of bones to both vitamin D deprivation and vitamin D dosage after deprivation was indicated by the changes in the percentages of ash in the bones and the changes in number and distribution of osteoclasts.

scholarly journals Purification and immunological quantification of rat liver lysosomal glycosidases

1989 ◽  
Vol 264 (1) ◽  
pp. 115-123 ◽  
Author(s):  
P C de Groen ◽  
G D LeSage ◽  
P S Tietz ◽  
N F LaRusso
Keyword(s):  
Enzyme Activity ◽  
Rat Liver ◽  
Lysosomal Enzyme ◽  
Enzyme Activities ◽  
Molecular Sieve ◽  
Polyacrylamide Gel Electrophoresis ◽  
Enzyme Concentration ◽  
Regulatory Control ◽  
Beta Galactosidase ◽  
Lysosomal Glycosidases

Although lysosomal enzyme activities are known to vary in response to numerous physiological and pharmacological stimuli, the relationship between lysosomal enzyme activity and enzyme concentration has not been systematically studied. Therefore we developed radioimmunoassays for two lysosomal glycosidases in order to determine lysosomal enzyme concentration. beta-Galactosidase and beta-glucuronidase were purified from rat liver 2780-fold and 1280-fold respectively, by using differential centrifugation, affinity chromatography, ion-exchange chromatography and molecular-sieve chromatography. Polyclonal antibodies to these enzymes were raised in rabbits, and two radioimmunoassays were established. Antibody specificity was shown by: (i) selective immunoprecipitation of enzyme activity; (ii) identical bands of purified enzyme on SDS/polyacrylamide-gel electrophoresis and immunoelectrophoresis; (iii) single immunoreactive peaks in molecular-sieve chromatography experiments. Sensitivities of the assays were such that 15 ng of beta-galactosidase and 45 ng of beta-glucuronidase decreased the ratio of bound to free radiolabel by 50%; minimal detectable amounts of immunoreactive enzymes were 2 ng and 10 ng respectively. The assays were initially used to assess the effects of physiological perturbations (i.e. fasting and age) on enzyme concentrations in rat liver; these experiments showed that changes in enzyme concentrations do not always correlate with changes in enzyme activities. This represents the first report of radioimmunoassays for lysosomal glycosidases. The results suggest that these radioimmunoassays provide useful technology for the study of regulatory control mechanisms of the concentrations of lysosomal glycosidases in mammalian tissues.

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.

scholarly journals The Role of Arg13 in Protein Phosphatase M tPphA from Thermosynechococcus elongatus

2012 ◽  
Vol 2012 ◽  
pp. 1-7 ◽  
Author(s):  
Jiyong Su ◽  
Karl Forchhammer
Keyword(s):  
Amino Acid ◽  
Protein Phosphatase ◽  
Arginine Residue ◽  
Side Chain ◽  
Wild Type ◽  
Catalytic Center ◽  
Nitrophenyl Phosphate ◽  
Reduced Activity ◽  
Amino Acid Variants

A highly conserved arginine residue is close to the catalytic center of PPM/PP2C-type protein phosphatases. Different crystal structures of PPM/PP2C homologues revealed that the guanidinium side chain of this arginine residue can adopt variable conformations and may bind ligands, suggesting an important role of this residue during catalysis. In this paper, we randomly mutated Arginine 13 of tPphA, a PPM/PP2C-type phosphatase from Thermosynechococcus elongatus, and obtained 18 different amino acid variants. The generated variants were tested towards p-nitrophenyl phosphate and various phosphopeptides. Towards p-nitrophenyl phosphate as substrate, twelve variants showed 3–7 times higher Km values than wild-type tPphA and four variants (R13D, R13F, R13L, and R13W) completely lost activity. Strikingly, these variants were still able to dephosphorylate phosphopeptides, although with strongly reduced activity. The specific inability of some Arg-13 variants to hydrolyze p-nitrophenyl phosphate highlights the importance of additional substrate interactions apart from the substrate phosphate for catalysis. The properties of the R13 variants indicate that this residue assists in substrate binding.

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