Індикаторна роль ферментів нітратного обміну в умовах екологічних змін стану навколишнього середовища

O.M. Vasilyuk; A.Y. Pakhomov

doi:10.15421/2018_239

Індикаторна роль ферментів нітратного обміну в умовах екологічних змін стану навколишнього середовища

Ukrainian Journal of Ecology ◽

10.15421/2018_239 ◽

2018 ◽

Vol 8 (1) ◽

pp. 483-486

Author(s):

O.M. Vasilyuk ◽

A.Y. Pakhomov

Keyword(s):

Enzyme Activity ◽

Nitrogen Metabolism ◽

Toxic Metal ◽

Total Activity ◽

Research Area ◽

Protective Properties ◽

Low Concentrations ◽

European Mole ◽

Glechoma Hederacea ◽

Metal Effect

The paper reflects analyzes of Cd impact on the total activity (nM pyruvic acid/ml s) of aspartate aminotransferase (AST, EC 2.6.1.1) nitrogen metabolism in Glechoma hederacea L. leaves subject (as model) which dominated in the research area (in natural floodplain oak with Stellaria holostea L.) in conditions of Cd pollution (as anthropogenic press) and digging activity by Mammalia (as biotic action, with Talpa europaea L., European mole, as model),) and their combine action. The Cd was introduced in the form of salts Cd(NO3)2 in the concentrations: 0.25, 1.25 and 2,5 g/m2, equivalent to the inclusion of Cd in 1,5 and 10 doses of MAC on experimental sites. When adding Cd, the content of doses (5 mg/kg soil MAC of Cd) was taken into account. It was founded the increasing of the AST activity on 26% (with adding the Cd salts at a dose of 1 МAС and digging activity by Talpa europaea L) according to control (1 MAC Cd), witch it proved the non-specific reaction on stress. With Cd concentration 5 and 10 MAC we observed the repression of the enzymes activity according to controls (5 and 10 MAC Cd) on 10% and 50% in accordance. The protective properties by T. europaea L. hadn’t positive results. The transferase enzyme activity according to another control (the area, is without pollution of Cd and digging activity by T. europaea L.) reflected the increasing AST enzyme activity from 166% tо 218% (in presence 1 and 5 MAC Cd) and reduction around 46% (in presence 10 MAC Cd). The digging activity by T. europaea L. lowered the toxic metal effect and the normalisation of the nitrogen metabolism by increasing the activity of AST from 55% to 266%, from 318% to 291% (AST, 1 та 5 MAC Cd). The digging activity by Mammalia did not contribute the metal toxic effect under the Cd 10 MAC. Thus, using the different representatives of zoocoenosis promotes improvement in the Steppe Dnieper at low concentrations of the factor has been revealed.

Вплив рийної функції Mammalia на активність аланінамінотрансферази в листках Glechoma hederacea в умовах кадмієвого забруднення

Ukrainian Journal of Ecology ◽

10.15421/2017_73 ◽

2017 ◽

Vol 7 (3) ◽

pp. 234-238

Author(s):

O. M. Vasilyuk ◽

A. Y. Pahomov

Keyword(s):

Environmental Changes ◽

Toxic Metal ◽

Total Activity ◽

Research Area ◽

Control Area ◽

Protective Properties ◽

European Mole ◽

Nitrate Metabolism ◽

Complex Regulation ◽

Glechoma Hederacea

The paper reflects analyzes of Cd impact on the total activity (nM pyruvic acid/ml s) of Alanine aminotransferase (ALT, EC 2.6.1.2) nitrogen metabolism in Glechoma hederacea L. leaves subject (as model) which dominated in the research area (in natural floodplain oak with Stellaria holostea L.) in conditions of Cd pollution (as anthropogenic press) and digging activity by Mammalia (as biotic action, with Talpa europaea L., European mole, as model), and their combine action. The Cd was introduced in the form of salts Cd(NO3)2 in the concentrations: 0.25, 1.25, and 2,5 g/m2, equivalent to the inclusion of Cd in 1,5 and 10 doses of MAC. The content of doses of MAC of Cd (5 mg/kg soil) adding took into account.It was found the increasing of the ALT activity on 88% (with adding the Cd salts at a dose of 1 МAС) and digging activity by Talpa europaea L. which proved the non-specific reaction on stress. We observed the repression of the enzymes according to controls (5 and 10 MAC Cd) with Cd concentration 5 and 10 MAC. The protective properties by T. europaea L. hadn’t positive results. The transferase enzyme activity according to another control (area without pollution of Cd and digging activity by T. europaea L.) reflected the inhibition of ALT on 78% tо 53% (in presence Cd 1 and 5 MAC). The digging activity by T. europaea L. promoted the toxic metal level and the normalisation of the nitrate metabolism from 25% tо 47% (ALT, 1 MAC Cd). The digging activity by Mammalia did not contribute the metal toxic effect and restoration of the natural functions of the plant organism under the Cd 10 MAC. The advisability for using the representatives of zoocenosis for the complex regulation of environmental changes in the conditions of the Ukrainian Steppe, if the antropogenic factor does not exceeds the maximum permissible significance have been founded.

Effect of Chromium(VI) Toxicity on Enzymes of Nitrogen Metabolism in Clusterbean (Cyamopsis tetragonoloba L.)

Enzyme Research ◽

10.1155/2014/784036 ◽

2014 ◽

Vol 2014 ◽

pp. 1-9 ◽

Cited By ~ 25

Author(s):

Punesh Sangwan ◽

Vinod Kumar ◽

U. N. Joshi

Keyword(s):

Heavy Metals ◽

Enzyme Activity ◽

Nitrogen Metabolism ◽

Potassium Dichromate ◽

Grain Filling ◽

Growth And Yield ◽

Growth Stages ◽

Cyamopsis Tetragonoloba ◽

Chromium Vi ◽

Low Concentrations

Heavy metals are the intrinsic component of the environment with both essential and nonessential types. Their excessive levels pose a threat to plant growth and yield. Also, some heavy metals are toxic to plants even at very low concentrations. The present investigation (a pot experiment) was conducted to determine the affects of varying chromium(VI) levels (0.0, 0.5, 1.0, 2.0, and 4.0 mg chromium(VI) kg−1 soil in the form of potassium dichromate) on the key enzymes of nitrogen metabolism in clusterbean. Chromium treatment adversely affect nitrogenase, nitrate reductase, nitrite reductase, glutamine synthetase, and glutamate dehydrogenase in various plant organs at different growth stages as specific enzyme activity of these enzymes decreased with an increase in chromium(VI) levels from 0 to 2.0 mg chromium(VI) kg−1 soil and 4.0 mg chromium(VI) kg−1 soil was found to be lethal to clusterbean plants. In general, the enzyme activity increased with advancement of growth to reach maximum at flowering stage and thereafter decreased at grain filling stage.

Serum Cholinesterase Variants: Examination of Several Differential Inhibitors, Salts, and Buffers Used to Measure Enzyme Activity

Clinical Chemistry ◽

10.1093/clinchem/17.3.183 ◽

1971 ◽

Vol 17 (3) ◽

pp. 183-191 ◽

Cited By ~ 30

Author(s):

Philip J Garry

Keyword(s):

Enzyme Activity ◽

Sodium Fluoride ◽

Phosphate Buffer ◽

Divalent Cations ◽

Kinetic Studies ◽

Competitive Inhibitor ◽

Serum Cholinesterase ◽

Low Concentrations

Abstract Dibucaine, used as a differential inhibitor with acetyl-, propionyl-, and butyrylthiocholine as substrate, clearly identified the "usual" and "atypical" serum cholinesterases. Succinylcholine was also used successfully as a differential inhibitor with butyrylthiocholine as substrate. Sodium fluoride, used as a differential inhibitor, gave conflicting results, depending on whether Tris or phosphate buffer was used in the assay. Mono- and divalent cations (NaCl, KCl, MgCl2, CaCl2, and BaCl2) activated the "usual" and inhibited the "atypical" enzyme at low concentrations. The "usual" enzyme had the same activity in 0.05 mol of Tris or phosphate buffer per liter, while the heterozygous and "atypical" enzymes showed 12 and 42% inhibition, respectively, when assayed in the phosphate buffer. Kinetic studies showed the phosphate acted as a competitive inhibitor of "atypical" enzyme. Km values, determined for "usual" and "atypical" enzymes, were 0.057 and 0.226 mmol/liter, respectively, with butyrylthiocholine as substrate.

5α-Reductase activity in stroma and epithelium of rat prostate and epididymis. A contribution to elucidation of the mechanism for development of hyperplastic growth of prostatic tissue

Acta Endocrinologica ◽

10.1530/acta.0.1030273 ◽

1983 ◽

Vol 103 (2) ◽

pp. 273-281 ◽

Cited By ~ 8

Author(s):

Ole Djøseland ◽

Nicholas Bruchovsky ◽

Paul S. Rennie ◽

Navdeep Otal ◽

Sian Høglo

Keyword(s):

Enzyme Activity ◽

Specific Activity ◽

Reductase Activity ◽

Major Change ◽

Growth Stimulation ◽

Total Activity ◽

Prostatic Tissue ◽

Ventral Prostate ◽

Total Enzyme Activity ◽

Abnormal Growth

Abstract. The 5α-reductase activity was assayed in homogenates of stroma and epithelium in the rat ventral prostate and epididymis. Samples consisting of 0.3 mg/ml tissue protein in TES buffer, pH 7.0 were incubated at 37°C for 30 min in the presence of 50 nm [1,2-3H]testosterone and a NADPH-generating system started with 5 × 10−4 m NADP. The yield of 5α-reduced metabolites, as established by using thin-layer chromatography, gave an estimate of enzyme activity. Whereas the specific activity of 5α-reductase was highest in prostatic stroma and epididymal epithelium, most of the total enzyme activity was associated with the epithelium in both the prostate and epididymis. The effect of dihydrotestosterone on specific activity of 5α-reductase was studied by administering the hormone to 7-day castrated rats. In prostate, the specific activity of both stromal and epithelium forms of the enzyme reached a maximum after 4 days of treatment. In epididymis only the epithelial form of 5α-reductase underwent a major change in specific activity, the latter peaking after 8–12 days of treatment. Furthermore, while the total activity of 5α-reductase in the prostatic tissue fractions could be induced by as much as 4-fold the normal control values, the epididymal enzyme could not be induced above the normal level either in the stroma or the epithelium. This may explain the relative resistance of epididymis to abnormal growth stimulation under the influence of hormones.

Corrigendum: Chloroplast Glutamine Synthetase, the Key Regulator of Nitrogen Metabolism in Wheat, Performs Its Role by Fine Regulation of Enzyme Activity via Negative Cooperativity of Its Subunits

Frontiers in Plant Science ◽

10.3389/fpls.2018.00466 ◽

2018 ◽

Vol 9 ◽

Cited By ~ 1

Author(s):

Edit Németh ◽

Zoltán Nagy ◽

Attila Pécsváradi

Keyword(s):

Enzyme Activity ◽

Glutamine Synthetase ◽

Nitrogen Metabolism ◽

Negative Cooperativity ◽

Fine Regulation ◽

Regulation Of Enzyme Activity

Analysis of the forms of acetylcholinesterase from adult mouse brain

Biochemical Journal ◽

10.1042/bj1470205 ◽

1975 ◽

Vol 147 (2) ◽

pp. 205-214 ◽

Cited By ~ 39

Author(s):

E D Adamson ◽

S E Ayers ◽

Z A Deussen ◽

C F Graham

Keyword(s):

Enzyme Activity ◽

Mouse Brain ◽

Gel Electrophoresis ◽

Polyacrylamide Gel Electrophoresis ◽

Gel Filtration ◽

Acetylcholinesterase Activity ◽

Total Activity ◽

Polyacrylamide Gel ◽

Soluble Extract ◽

Molecular Sizes

The solubilization of 80% of the acetylcholinesterase activity of mouse brain was performed by repeated 2h incubations of homogenates at 37 degrees C in an aqueous medium. Analysis of the soluble extract by gel filtration on Sephadex G-200 showed that up to 80% of the enzyme activity was eluted in a peak which was estimated to consist of molecules of about 74000mol.wt. This peak was called the monomer form of the enzyme. After 3 days at 4 degrees C, the soluble extract was re-analysed and was eluted from the column in four peaks of about 74000, 155000, 360000 and 720000 mol.wt. Since the total activity of the enzyme in these peaks was the same as that in the predominantly monomer elution profile of fresh enzyme, we concluded that the monomer had aggregated, possibly into dimers, tetramers and octomers. Extracts of the enzyme were analysed by polyacrylamide-gel electrophoresis and the resulting multiple bands of enzyme activity on gels were shown to separate according to their molecular sizes, that is by molecular sieving. All these forms had similar susceptibilities to the inhibitors eserine, tetra-isopropyl pyrophosphoramide and compound BW 284c51 [1,5-bis-(4-allyldimethylammoniumphenyl)pentan-3-one dibromide]. Thus the forms of the enzyme in mouse brain which can be detected by gel filtration and polyacrylamide-gel electrophoresis may all be related to a single low-molecular-weight form which aggregates during storage. This supports similar suggestions made for the enzyme in other locations.

Changes in Ouabain-Sensitive Adenosine Triphosphatase Activity in Gills of Coho Salmon (Oncorhynchus kisutch) During Parr–Smolt Transformation

Journal of the Fisheries Research Board of Canada ◽

10.1139/f76-007 ◽

1976 ◽

Vol 33 (1) ◽

pp. 54-62 ◽

Cited By ~ 36

Author(s):

M. A. Giles ◽

W. E. Vanstone

Keyword(s):

Enzyme Activity ◽

Coho Salmon ◽

Incubation Temperature ◽

Sea Water ◽

Oncorhynchus Kisutch ◽

Sharp Decrease ◽

Total Activity ◽

Sodium Ions ◽

Relative Contribution ◽

Sodium Activation

The effects of incubation temperature, pH, sodium, potassium, and ATP concentration, and ouabain on the activity of Na+–K+-activated ATPase of the gills of seawater-adapted juvenile coho salmon (Oncorhynchus kisutch) were determined. The temperature and pH optima were 40 C and 7.4, respectively. The apparent Km for ATP at equimolar Mg++ concentration was 0.2 mM at Na+ and K+ concentrations of 100 and 20 mM, respectively. Maximal enzyme activity for Na+ concentration of 10.50 and 100 mM occurred at K+ concentrations of 12.5, 15.0, and 20.0 mM, respectively. The Ki for ouabain was 2 × 10−6 M and 7 × 10−6 for K+ concentrations of 10 and 20 mM, respectively.A large portion (up to 60%) of the ouabain-sensitive ATPase activity in freshwater fish was activated by sodium ions in the absence of potassium ions (Na+-activation). Exposure to sea water resulted in a large increase in total ouabain-sensitive activity and a sharp decrease in the proportion of sodium activation. These changes occurred within 14 days after transfer to full strength sea water.On a seasonal basis, total ouabain-sensitive enzyme activity in juvenile freshwater coho was low (less than 5 μmol Pi/mg N per h) to the end of November, increased to a peak (over 125 μmol Pi/mg N per h) in mid-January, and subsequently declined by late February. A slow, steady rise in activity occurred during the smoking period of March and April and the relative contribution of sodium ions to the total activity declined in this period.

Inactivation of glucose 6-phosphate dehydrogenase during germination and outgrowth of Bacillus cereus T endospores

Biochemical Journal ◽

10.1042/bj1480259 ◽

1975 ◽

Vol 148 (2) ◽

pp. 259-268 ◽

Cited By ~ 4

Author(s):

M Orlowski ◽

M Goldman

Keyword(s):

Enzyme Activity ◽

Specific Activity ◽

Proteolytic Enzymes ◽

Polyacrylamide Gel Electrophoresis ◽

Total Activity ◽

Cellular Protein ◽

Metabolic Energy ◽

Supernatant Fraction ◽

Stage Of Development ◽

Glucose 6 Phosphate Dehydrogenase

The specific activity and total activity of glucose 6-phosphate dehydrogenase (EC 1.1.1.49) under conditions of complete cell breakage fall 10-20-fold during a 3h period of spore germination and outgrowth. The spores must germinate (lose refractility), but do not have to undergo outgrowth, for the loss of activity to occur. Glucose 6-phosphate dehydrogenase activity from cells as any stage of development is completely stable in extracts at 4 degrees C or 30 degrees C. All of the enzyme activity is found in a soluble (50000g supernatant) fraction and remains completely soluble throughout development. Soluble protein and total cellular protein remain constant for about 2h. Proteinases could not be detected or protein turnover demonstrated during the morphogenetic process. Phenylmethanesuophony fluoride and o-phenanthroline, inhibitors of proteolytic enzymes, do not prevent glucose 6-phosphate dehydrogenase inactivation when added to whole cells. Mixing experiments show no inhibitor of glucose 6-phosphate dehydrogenase to be present in late-stage cells. The enzyme is not excreted into the culture medium. Chloramphenicol and rifampicine immediately stop protein synthesis and development but not the inactivation of glucose 6-phosphate dehydrogenase. NaN3, 2,4-dinitrophenol or anaerobiosis immediately stop development and prevent the loss of enzyme activity. A requirement for metabolic energy is therefore probable. Extracts of spores pre-labelled with L[14C]leucine were made at various stages of morphogenesis and subjected to polyacrylamide-gel electrophoresis. Glucose 6-phosphate dehydrogenase, which was identified by a specific stain, did not lose 14C label, and therefore may not be degraded during the inactivation process.

EGTA-sensitive and -insensitive forms of particulate adenylate cyclase in rat cerebral cortex: regulation by divalent cations and GTP

Canadian Journal of Physiology and Pharmacology ◽

10.1139/y85-166 ◽

1985 ◽

Vol 63 (8) ◽

pp. 1007-1016 ◽

Cited By ~ 4

Author(s):

P. V. Sulakhe

Keyword(s):

Enzyme Activity ◽

Cerebral Cortex ◽

Adenylate Cyclase ◽

Gray Matter ◽

Rank Order ◽

Divalent Cations ◽

Rat Cerebral Cortex ◽

Chloride Salt ◽

Low Concentrations ◽

Stimulation Of

Interactions of several divalent cations (Mn2+, Ca2+, Co2+, Sr2+, and Zn2+) with EGTA-inhibitable adenylate cyclase were investigated in washed membranes (particles) isolated from the gray matter of rat cerebral cortex. The EGTA-inhibitable (called sensitive) enzyme activity was assayed in the presence of Triton X-100 since this detergent caused a marked increase (up to 20-fold) in the enzyme activity. The effects of various divalent metals (all added as chloride salt) indicated the presence of two distinct sites called site I and site II. At low concentrations (less than micromolar) Mn2+, Co2+, and Ca2+ increased (up to 10-fold) the enzyme activity to the same extent and appeared to act via binding to site I (high affinity site). The rank order of affinity was Mn2+ ≥ Co2+ > Ca2+. Zn2+ showed the highest affinity and Sr2+ the lowest towards binding to site I; both these metals increased the enzyme activity to lesser extents than Mn2+, Co2+, or Ca2+. GTP was not required for the stimulation of this enzyme by low concentrations of Ca2+. The interaction of Mn2+ with site II (low affinity site) caused further increase in the enzyme activity, whereas Co2+, Ca2+, and Sr2+ were inhibitory at concentrations >10 μM. Isolated fraction contained loosely and tightly associated pools of calmodulin. Myelin basic protein, but not calcineurin, inhibited the EGTA-sensitive adenylate cyclase activity. The EGTA-insensitive enzyme activity was increased by norepinephrine by mechanisms that depended on GTP and was inhibited by Ca2+. The stimulation of the EGTA-insensitive enzyme modulated the Mg2+ requirement such that Mg2+ binding to the low affinity site (site II) apparently occurred with higher affinity. The likely significance of these results is discussed with regard to (i) the presence of two classes of adenylate cyclase in rat cerebral cortex gray matter and (ii) the regulation of their activities by calmodulin-requiring and GTP-requiring mechanisms.

Coproporphyrinogenase in tobacco (Nicotiana tabacum L.)

Biochemical Journal ◽

10.1042/bj1170215 ◽

1970 ◽

Vol 117 (2) ◽

pp. 215-220 ◽

Cited By ~ 41

Author(s):

W. P. Hsu ◽

G. W. Miller

Keyword(s):

Enzyme Activity ◽

Nicotiana Tabacum ◽

Differential Centrifugation ◽

Enzyme Protein ◽

Sodium Amalgam ◽

Low Concentrations ◽

Nicotiana Tabacum L ◽

Km Value ◽

Ammonium Sulphate Fractionation ◽

Final Purification

1. Coproporphyrinogenase was extracted and purified from tobacco (Nicotiana tabacum L.). Enzyme activity was mainly located in mitochondria rather than in chloroplasts. The enzyme was purified by differential centrifugation, ammonium sulphate fractionation, calcium phosphate gel adsorption and dialysis. A 69-fold final purification was obtained. 2. An apparent Km value of 3.6×10−5m was found, the value being largely dependent on the amount of coproporphyrin III recovered after reduction with sodium amalgam to coproporphyrinogen III. Protoporphyrin formation was linear up to 3h and decreased with further incubation. The enzyme activity increased with the concentration of enzyme protein up to 30μg/ml of solution. 3. Enzyme activity was greatly enhanced by increasing Fe2+ concentrations up to 0.5mm, beyond which inhibition occurred. Co2+ and Mn2+ were also found to activate at low concentrations (0.1mm) and inhibit at higher concentrations (5mm). Fe3+ and Cu2+, both at 0.1mm, and o-phenanthroline and EDTA, each at 1mm, were found to be inhibitory.