scholarly journals Modulation of guanine deaminase

1973 ◽  
Vol 131 (4) ◽  
pp. 683-687 ◽  
Author(s):  
K. Sree Kumar ◽  
A. Sitaramayya ◽  
P. S. Krishnan
Keyword(s):  
Rat Brain ◽  
Substrate Concentration ◽  
Mouse Liver ◽  
Deae Cellulose ◽  
Supernatant Fraction ◽  
Saturation Curve ◽  
Guanine Deaminase ◽  
Sigmoidal Kinetics ◽  
Substrate Saturation

1. Guanine deaminases purified from the 15000g supernatant fraction of iso-osmotic sucrose homogenates of rat and mouse liver and brain were tested for the influence of GTP and allantoin. 2. The suffixes A and B were assigned to the isoenzyme fractions eluted from DEAE-cellulose with the lower and the higher molarity of eluent respectively. Isoenzyme A from rat liver, the activity of which showed a sigmoid dependence on substrate saturation, was activated by GTP and inhibited by allantoin. Isoenzyme B, which had a hyperbolic substrate-saturation curve, was not influenced by GTP or allantoin. 3. Isoenzyme A from rat brain, the activity of which had a sigmoid dependence on substrate concentration, was stimulated by GTP. Isoenzyme B, which showed classical Michaelis–Menten kinetics, was inhibited by allantoin. 4. Mouse liver guanine deaminase was not influenced by either GTP or allantoin. 5. Isoenzyme A from mouse brain, which had a hyperbolic substrate-saturation curve, was not influenced by GTP or allantoin but isoenzyme B, with sigmoidal kinetics, was inhibited by allantoin. 6. Mg2+ activated, or inhibited or did not have an effect on guanine deaminase, depending on the source of the enzyme. 7. The bearing of the above findings on the possible regulation of guanine deaminase activity in vivo is discussed.

scholarly journals Guanine deaminase in rat liver and mouse liver and brain

1972 ◽  
Vol 128 (5) ◽  
pp. 1079-1088 ◽  
Author(s):  
K. S Kumar ◽  
A. Sitaramayya ◽  
P. S. Krishnan
Keyword(s):  
Rat Brain ◽  
Rat Liver ◽  
Inhibitory Activity ◽  
Substrate Concentration ◽  
Mouse Liver ◽  
Deae Cellulose ◽  
The Other ◽  
Guanine Deaminase ◽  
Sigmoidal Kinetics ◽  
Km Value

1. The guanine deaminase in rat liver supernatant preparations was resolved into two fractions, A and B, on DEAE-cellulose columns. The two differed in electrophoretic mobility and in various properties. The most noteworthy distinction between A and B components was that the enzyme A activity showed a sigmoid dependence on substrate concentration whereas the enzyme B showed classical Michaelis–Menten kinetics. The Km value of enzyme A for guanine was 5.3μm and that of enzyme B 20μm. 2. The entire guanine deaminase activity of mouse liver was contained in the 15000g supernatant of iso-osmotic homogenates. 3. A reinvestigation of the behaviour of rat brain 15000g supernatant guanine deaminase isoenzymes revealed that one enzyme had sigmoidal kinetics and the other enzyme showed a hyperbolic response. 4. Of the guanine deaminase in mouse brain iso-osmotic sucrose homogenate 80% was recovered in the 15000g supernatant and the rest from the particles. The supernatant guanine deaminase was resolvable into two fractions on DEAE-cellulose columns. One enzyme showed sigmoidal kinetics whereas the other showed a hyperbolic response to increasing substrate concentration; the Km values for the reaction with guanine were respectively 5 and 66μm. 5. The particulate fractions of mouse liver and brain were devoid of any overt inhibitory activity.

Pyruvate kinase from the earthworm Allolobophora calliginosa: modification of the enzyme during anaerobiosis

1991 ◽  
Vol 69 (1) ◽  
pp. 251-254
Author(s):  
Athanasios I. Papadopoulos ◽  
Basile Michaelidis ◽  
Isidoros Beis
Keyword(s):  
Pyruvate Kinase ◽  
Deae Cellulose ◽  
Relative Activity ◽  
The Body ◽  
E Coli ◽  
In Vitro Incubation ◽  
Sigmoidal Kinetics ◽  
Independent Protein

The relative activity of pyruvate kinase from the body-wall muscle of the earthworm Allolobophora calliginosa was found to drop dramatically within 6 h of exposure to N2, whereas the opposite was observed during recovery. Two forms of pyruvate kinase (designated as peak I and peak II) were separated chromatographically on DEAE-cellulose and eluted at 50 and 150 mM of KCl, respectively. They displayed different kinetic behaviour with respect to substrate phosphoenolpyruvate; peak I exhibited Michaelis–Menten kinetics whereas peak II showed sigmoidal kinetics. The ratio of the enzyme units (peak I/peak II) decreased from 3.38 under normoxic conditions to 0.09 under anoxic conditions. In vitro incubation of the aerobic form of pyruvate kinase in the presence of ATP and Mg++ resulted in a reduction of the enzyme activity by 64%, suggesting the presence of an endogenous cyclic-nucleotide-independent protein kinase capable of phosphorylating pyruvate kinase. After in vitro incubation, alkaline phosphatase from E. coli increased the depressed activity of anaerobic pyruvate kinase, indicating that the enzyme molecule is phosphorylated in vivo during exposure to anoxia.

scholarly journals Aspartate transcarbamoylase from Phaseolus aureus. Partial purification and properties

1972 ◽  
Vol 129 (3) ◽  
pp. 571-581 ◽  
Author(s):  
B. L. Ong ◽  
J. F. Jackson
Keyword(s):  
Gel Filtration ◽  
Deae Cellulose ◽  
Aspartate Transcarbamoylase ◽  
Partial Purification ◽  
Saturation Curve ◽  
Phaseolus Aureus ◽  
Phosphate Binding ◽  
Carbamoyl Phosphate ◽  
Sequential Reaction ◽  
Substrate Saturation

1. Aspartate transcarbamoylase from 4-day-old radicles of Phaseolus aureus was purified 190-fold by (NH4)2SO4 fractionation, DEAE-cellulose and DEAE-Sephadex chromatography and Sephadex-gel filtration. The partially purified enzyme, which required Pi for maximum stability, had an apparent molecular weight of 83000±5000. 2. Uridine nucleotides were found to inhibit the activity; UMP was the most potent inhibitor, followed by UDP and UTP. No other nucleotide was found to affect the enzyme, nor could UMP inhibition be overcome by adding another nucleotide. Aspartate gives a hyperbolic substrate-saturation curve, both with and without UMP. The nucleotide inhibitor is non-competitive with respect to this substrate. Carbamoyl phosphate also yields a hyperbolic substrate-saturation curve in the absence of feedback inhibitor, but when UMP is added a sigmoidal pattern results, and the inhibition is competitive with carbamoyl phosphate. 3. The degree of inhibition by UMP is not affected by p-chloromercuribenzoate, urea, mild heat pretreatment or change in pH over the range 8.5–10.5, but is affected by temperature. 4. The aspartate analogue, succinate, both activates and inhibits the reaction, depending on the concentrations of aspartate and succinate used. 5. Kinetic studies with the partially purified enzyme showed that the Km for carbamoyl phosphate (0.091 mm) is much lower than that for aspartate (1.7mm). A sequential reaction mechanism was inferred from product-inhibition kinetics, with carbamoyl phosphate binding to the enzyme before aspartate, and the product, carbamoylaspartate, being released ahead of Pi. Initial-velocity studies gave a set of parallel reciprocal plots, compatible with an essentially irreversible step occurring before the binding of aspartate.

Evidence for glucocorticosteroid receptors in the erythroid cell line of fetal rat liver

1983 ◽  
Vol 96 (2) ◽  
pp. 311-319 ◽  
Author(s):  
P. Mayeux ◽  
C. Billat ◽  
J. M. Felix ◽  
R. Jacquot
Keyword(s):  
Activated Charcoal ◽  
Fetal Liver ◽  
Deae Cellulose ◽  
Erythroid Cell ◽  
Supernatant Fraction ◽  
Major Peak ◽  
Fetal Rat ◽  
Fetal Rat Liver ◽  
Macromolecular Component

A role for glucocorticosteroids in the evolution of the rat fetal liver erythropoiesis in vivo has been proposed; accordingly suppressible binding of [3H]dexamethasone by intact liver erythroid cells at 4 °C has been described. The nature of this binding was further investigated in the present paper. Suspensions of cells of the erythroid line were prepared from liver erythropoietic tissue obtained from fetuses aged 15 or 16 days. Suppressible binding of [3H]dexamethasone was studied on these suspensions. After incubation at 4 °C, approximately 89 per cent of this binding was located in the cytosol (100 000 g supernatant fraction), was excluded from Sephadex G-50 columns and was not adsorbed on activated charcoal; 11·3 ± 2·6 (s.d.) per cent (n = 4) of the suppressible binding was present in the nucleus. When cells prelabelled with [3H]dexamethasone at 4 °C were warmed to 37 °C, 52 ± 10 per cent (n = 5) of the suppressible binding was rapidly transferred to the nucleus. The suppressible radioactivity present in the cytosols at 4 °C was eluted from diethylaminoethyl (DEAE)–cellulose columns, in the presence of molybdate, as a single peak (peak II) at phosphate concentrations between 200 and 250 mmol/l. When these cytosols were warmed at 25 °C before chromatography, the radioactivity was eluted from DEAE-cellulose as a major peak (peak I) at phosphate concentrations between 50 and 70 mmol/l, and as a smaller peak corresponding to peak II. The presence of molybdate during the incubation at 25 °C prevented the formation of peak I. The suppressible binding present in the cytosols of cells incubated at 37 °C was eluted from DEAE–cellulose columns in two equivalent peaks corresponding to peaks I and II. It was concluded that [3H]dexamethasone was bound to a macromolecular component sharing the properties generally ascribed to the receptor of glucocorticosteroids.

FRACTIONATION OF MOUSE LIVER MICROSOMAL ESTERASES

1965 ◽  
Vol 43 (1) ◽  
pp. 97-104 ◽  
Author(s):  
C. Carruthers ◽  
E. Heins ◽  
A. Baumler
Keyword(s):  
Esterase Activity ◽  
Mouse Liver ◽  
Liver Homogenate ◽  
Deae Cellulose ◽  
Supernatant Fraction ◽  
Nonionic Detergent ◽  
Liver Homogenates ◽  
Two Peaks ◽  
Total Esterase Activity ◽  
Glycyl Glycine

Mouse liver microsomal esterases were fractionated on DEAE-cellulose after the solution of these enzymes in a solution containing 0.1 M glycyl glycine buffer, pH 7.0, and 5 × 10−4 M Lubrol W, a nonionic detergent. Elution of the enzymes from the DEAE-cellulose was accomplished by using NaCl in the glycyl glycine – Lubrol W solution. Microsomes isolated from sucrose and glycerol homogenates have three esterase peaks in common and two peaks not in common. The glycerol supernatant fraction from whole liver homogenate, which contains about 50% of the total esterase activity, and the sucrose supernatant fraction, which contains about 6% of the total esterase activity, have four common esterase peaks, one of which is different from both the microsomes isolated from glycerol and sucrose liver homogenates. Incorporation of Lubrol W in the solution that was employed for the elution of the esterases from the DEAE-cellulose was necessary to prevent extensive loss or inactivation of these enzymes. The nature of the changes induced by glycerol on liver esterases is briefly discussed.

scholarly journals Studies on guanine deaminase and its inhibitors in rat tissue

1967 ◽  
Vol 102 (3) ◽  
pp. 691-704 ◽  
Author(s):  
S. Kumar ◽  
V. Josan ◽  
K.C.S. Sanger ◽  
K.K Tewari ◽  
P. S. Krishnan
Keyword(s):  
Deae Cellulose ◽  
Mitochondrial Fraction ◽  
Enzymic Activity ◽  
Cerebral Hemispheres ◽  
Supernatant Fraction ◽  
Whole Brain ◽  
Guanine Deaminase ◽  
Kidney Enzyme ◽  
Rat Tissue ◽  
Triton X

1. In kidney, but not in rat whole brain and liver, guanine-deaminase activity was localized almost exclusively in the 15000g supernatant fraction of iso-osmotic sucrose homogenates. However, as in brain and liver, the enzymic activity recovered in the supernatant was higher than that in the whole homogenate. The particulate fractions of kidney, especially the heavy mitochondria, brought about powerful inhibition of the supernatant guanine-deaminase activity. 2. In spleen, as in kidney, guanine-deaminase activity was localized in the 15000g supernatant fraction of iso-osmotic sucrose homogenates. However, the particulate fractions did not inhibit the activity of the supernatant. 3. Guanine-deaminase activity in rat brain was absent from the cerebellum and present only in the cerebral hemispheres. The inhibitor of guanine deaminase was located exclusively in the cerebellum, where it was associated with the particles sedimenting at 5000g from sucrose homogenates. 4. Homogenates of cerebral hemispheres, the separated cortex or the remaining portion of the hemispheres had significantly higher guanine-deaminase activity than homogenates of whole brain. The enzymic activity of the subcellular particulate fractions was nearly the same. 5. Guanine deaminase was purified from the 15000g supernatant of sucrose homogenates of whole brain. The enzyme separated as two distinct fractions, A and B, on DEAE-cellulose columns. 6. The guanine-deaminase activity of the light-mitochondrial fraction of whole brain was fully exposed and solubilized by treatment with Triton X-100, and partially purified. 7. Tested in the form of crude preparations, the inhibitor from kidney did not act on the brain and liver supernatant enzymes and the inhibitor from cerebellum did not act on kidney enzyme, but the inhibitor from liver acted on both brain and kidney enzyme. 8. The inhibitor of guanine deaminase was purified from the heavy mitochondria of whole brain and liver and the 5000g residue of cerebellum, isolated from iso-osmotic homogenates. The inhibitor appeared to be protein in nature and was heat-labile. The inhibition of the enzyme was non-competitive. 9. Kinetic, immunochemical and electrophoretic studies with the preparations purified from brain revealed that the enzyme from light mitochondria was distinct from enzyme B from the supernatant. A distinction between the two forms of supernatant enzyme was less certain. 10. Guanine deaminase isolated from light mitochondria of brain did not react with 8-azaguanine or with the inhibitor isolated from heavy mitochondria.

Pyruvate kinase from Aspergillus niger: a regulatory enzyme in glycolysis?

1984 ◽  
Vol 30 (1) ◽  
pp. 16-22 ◽  
Author(s):  
Bibiana Meixner-Monori ◽  
Christian P. Kubicek ◽  
Max Röhr
Keyword(s):  
Citric Acid ◽  
Aspergillus Niger ◽  
Pyruvate Kinase ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Acid Accumulation ◽  
Sigmoidal Kinetics ◽  
Atp Inhibition ◽  
Very High

Pyruvate kinase from the filamentous, citric acid producing fungus Aspergillus niger was purified about 100-fold by ammonium sulfate precipitation, DEAE-cellulose chromatography, and gel filtration. The addition of fructose-1,6-diphosphate was necessary to prevent loss of activity during purification. The enzyme purified in the presence of fructose-1,6-diphosphate (FDP) exhibits hyperbolic kinetics with respect to phosphoenolpyruvate (PEP) and ADP. Monovalent cations activated the enzyme (K+, NH4+). FDP neither activated nor inhibited the enzymatic activity from extracts freshly prepared in the absence of exogenous FDP; ATP showed a weak activation. In contrast the enzyme from crude extracts which had been stored in the presence of glycerol for 3 days showed activation by FDP or a metabolite thereof and inhibition by ATP. In the absence of FDP sigmoidal kinetics were obtained with respect to PEP, which became hyperbolic kinetics after addition of FDP. ATP inhibition turned into slight ATP activation in the presence of FDP. However, it was possible to reactivate inactivated pyruvate kinase (after dialysis in the absence of FDP) by adding FDP to the enzyme assay. From these results and because of the very high affinity of pyruvate kinase for FDP (Ka < 0.1 μM), it is concluded that the enzyme probably has FDP bound to the protein in vivo. The significance of this hypothesis to the regulation of glycolysis in A. niger, with special reference to the mechanism of citric acid accumulation, is discussed.

Sigma receptors mediate potent neuroprotection in vivo and inhibit neuronal depolarisation and swelling in rat brain slices

2005 ◽  
Vol 25 (1_suppl) ◽  
pp. S468-S468
Author(s):  
Jennifer K Callaway ◽  
Christine Molnar ◽  
Song T Yao ◽  
Bevyn Jarrott ◽  
R David Andrew
Keyword(s):  
Rat Brain ◽  
Brain Slices ◽  
Sigma Receptors

In vivo diffusion tensor imaging of the neonatal rat brain development

2013 ◽  
Vol 44 (S 01) ◽  
Author(s):  
M Breu ◽  
D Reisinger ◽  
D Wu ◽  
Y Zhang ◽  
A Fatemi ◽  
...  
Keyword(s):  
Diffusion Tensor Imaging ◽  
Brain Development ◽  
Rat Brain ◽  
Diffusion Tensor ◽  
Neonatal Rat ◽  
Neonatal Rat Brain
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