GLYCYL-RNA SYNTHETASE OF RAT LIVER: PARTIAL PURIFICATION AND EFFECTS OF SOME METAL IONS ON ITS ACTIVITY

1963 ◽  
Vol 41 (5) ◽  
pp. 1123-1133 ◽  
Author(s):  
M. J. Fraser
Keyword(s):  
Heat Treatment ◽  
Metal Ions ◽  
Ammonium Sulphate ◽  
Rat Liver ◽  
Partial Purification ◽  
Ammonium Sulphate Fractionation ◽  
Stimulation Of

Glycyl-RNA synthetase has been purified 40-fold from a 105,000 × g supernatant of an homogenate of rat liver by successive precipitation at pH = 5.0, heat treatment at 55 °C for 3.0 minutes in the presence of 1.0 mM ATP, and ammonium sulphate fractionation. The purified fractions catalyzed glycine-dependent ATP-32PP exchange. The effect of some metal ions on glycine activation was studied. Activation occurred in the presence of either Mg++or Mn++. The apparent stimulation of glycine activation by Co++was found to be an artifact.

GLYCYL-RNA SYNTHETASE OF RAT LIVER: PARTIAL PURIFICATION AND EFFECTS OF SOME METAL IONS ON ITS ACTIVITY

1963 ◽  
Vol 41 (1) ◽  
pp. 1123-1133 ◽  
Author(s):  
M. J. Fraser
Keyword(s):  
Heat Treatment ◽  
Metal Ions ◽  
Ammonium Sulphate ◽  
Rat Liver ◽  
Partial Purification ◽  
Ammonium Sulphate Fractionation ◽  
Stimulation Of

Glycyl-RNA synthetase has been purified 40-fold from a 105,000 × g supernatant of an homogenate of rat liver by successive precipitation at pH = 5.0, heat treatment at 55 °C for 3.0 minutes in the presence of 1.0 mM ATP, and ammonium sulphate fractionation. The purified fractions catalyzed glycine-dependent ATP-32PP exchange. The effect of some metal ions on glycine activation was studied. Activation occurred in the presence of either Mg++or Mn++. The apparent stimulation of glycine activation by Co++was found to be an artifact.

PARTIAL PURIFICATION AND PROPERTIES OF PROLYL-RNA SYNTHETASE OF RAT LIVER

1963 ◽  
Vol 41 (10) ◽  
pp. 2123-2140 ◽  
Author(s):  
M. J. Fraser ◽  
D. B. Klass
Keyword(s):  
Exchange Reaction ◽  
Rat Liver ◽  
Liver Homogenate ◽  
Competitive Inhibitor ◽  
Partial Purification ◽  
Appreciable Effect ◽  
Inorganic Pyrophosphate ◽  
Ammonium Sulphate Fractionation ◽  
Ph Range ◽  
Rna Formation

Prolyl-RNA (prolyl ribonucleic acid) synthetase has been purified 30-fold from a 105,000 × g supernatant of a rat liver homogenate by precipitation at pH 5.0, heat treatment at 55 °C for 3.0 minutes in the presence of 1.0 mM ATP (adenosine triphosphate), and by ammonium sulphate fractionation. The enzyme catalyzed proline-dependent ATP-32PP (PP, inorganic pyrophosphate) exchange and the formation of prolyl hydroxamate and of prolyl-RNA. Although the enzyme did not catalyze the formation of hydroxyprolyl-RNA, it catalyzed a slight hydroxyproline-dependent ATP-32PP exchange and the formation of a small amount of hydroxyprolyl hydroxamate which was much less than the amount of prolyl hydroxamate formed under the same conditions. The enzyme is thus not quite specific for proline activation, but is specific for amino acyl-RNA formation. It is probably concerned in protein biosynthesis.In the proline-dependent ATP-32PP exchange reaction the enzyme showed optimum activity in the pH range 6.2–8.2 and no activity at pH 5.0. The apparent Kmfor proline in this reaction was found to be 0.43 mM. Mg++was required for activity. Prolyl-RNA formation was optimal at pH 8.0. The apparent Kmfor proline in this reaction was found to be 2.5 μM. The effects of some proline analogues on proline activation were studied. Hydroxyproline and thioproline were found to inhibit both proline-dependent ATP-32PP exchange and prolyl-RNA formation. Thioproline was a competitive inhibitor of the exchange reaction and showed a KIof 0.95 mM. Pyrrolidone carboxylate had no appreciable effect on proline activation.

scholarly journals Glutathione S-aralkyltransferase

1969 ◽  
Vol 115 (5) ◽  
pp. 985-991 ◽  
Author(s):  
E. Boyland ◽  
L. F. Chasseaud
Keyword(s):  
Ammonium Sulphate ◽  
Rat Liver ◽  
Benzyl Chloride ◽  
Animal Species ◽  
Rat Kidney ◽  
Acid Precipitation ◽  
Heat Inactivation ◽  
Catalysed Reaction ◽  
Glutathione S Transferases ◽  
Ammonium Sulphate Fractionation

1. The name ‘glutathione S-aralkyltransferase’ is proposed for the enzyme catalysing the reaction of benzyl chloride with GSH. 2. Results from heat-inactivation studies, ammonium sulphate-fractionation and acid-precipitation experiments, and studies of the distribution of activities in rat liver, in rat kidney and in the livers of other animals indicate that glutathione S-aralkyltransferase differs from glutathione S-alkyltransferase, S-aryltransferase, S-epoxidetransferase and an S-alkenetransferase. 3. The distribution of these enzymes in the livers of the animal species examined was different. 4. Glutathione S-alkyltransferase, S-aralkyltransferase and the S-alkenetransferase that are present in rat liver supernatant were inhibited by GSSG, and the nature of the inhibition varied in each case. 5. 3,5-Di-tert.-butyl-4-hydroxybenzyl acetate reacts spontaneously with GSH, but the rat liver-supernatant-catalysed reaction of GSH with this and other aralkyl esters was weak. 6. A probable function of the glutathione S-transferases is the protection of cellular constituents from strong electrophilic agents.

PARTIAL PURIFICATION AND PROPERTIES OF PROLYL-RNA SYNTHETASE OF RAT LIVER

1963 ◽  
Vol 41 (1) ◽  
pp. 2123-2140 ◽  
Author(s):  
M. J. Fraser ◽  
D. B. Klass
Keyword(s):  
Exchange Reaction ◽  
Rat Liver ◽  
Liver Homogenate ◽  
Competitive Inhibitor ◽  
Partial Purification ◽  
Appreciable Effect ◽  
Inorganic Pyrophosphate ◽  
Ammonium Sulphate Fractionation ◽  
Ph Range ◽  
Rna Formation

Prolyl-RNA (prolyl ribonucleic acid) synthetase has been purified 30-fold from a 105,000 × g supernatant of a rat liver homogenate by precipitation at pH 5.0, heat treatment at 55 °C for 3.0 minutes in the presence of 1.0 mM ATP (adenosine triphosphate), and by ammonium sulphate fractionation. The enzyme catalyzed proline-dependent ATP-32PP (PP, inorganic pyrophosphate) exchange and the formation of prolyl hydroxamate and of prolyl-RNA. Although the enzyme did not catalyze the formation of hydroxyprolyl-RNA, it catalyzed a slight hydroxyproline-dependent ATP-32PP exchange and the formation of a small amount of hydroxyprolyl hydroxamate which was much less than the amount of prolyl hydroxamate formed under the same conditions. The enzyme is thus not quite specific for proline activation, but is specific for amino acyl-RNA formation. It is probably concerned in protein biosynthesis.In the proline-dependent ATP-32PP exchange reaction the enzyme showed optimum activity in the pH range 6.2–8.2 and no activity at pH 5.0. The apparent Kmfor proline in this reaction was found to be 0.43 mM. Mg++was required for activity. Prolyl-RNA formation was optimal at pH 8.0. The apparent Kmfor proline in this reaction was found to be 2.5 μM. The effects of some proline analogues on proline activation were studied. Hydroxyproline and thioproline were found to inhibit both proline-dependent ATP-32PP exchange and prolyl-RNA formation. Thioproline was a competitive inhibitor of the exchange reaction and showed a KIof 0.95 mM. Pyrrolidone carboxylate had no appreciable effect on proline activation.

Effects of 5-5'-Diphenyl-2-thiohydantoin (DPTH) and Thyroxine on the Ultrastructure and Chemical Composition of Rat Liver

1971 ◽  
Vol 29 ◽  
pp. 570-571
Author(s):  
E. A. Elfont ◽  
R. B. Tobin ◽  
D. G. Colton ◽  
M. A. Mehlman
Keyword(s):  
Chemical Composition ◽  
Rat Liver ◽  
Future Study ◽  
Mode Of Action ◽  
Liver Mitochondria ◽  
Rat Liver Mitochondria ◽  
Effective Inhibitor ◽  
Biochemical Effects ◽  
Glycerophosphate Dehydrogenase ◽  
Stimulation Of

Summary5,-5'-diphenyl-2-thiohydantoin (DPTH) is an effective inhibitor of thyroxine (T4) stimulation of α-glycerophosphate dehydrogenase in rat liver mitochondria. Because this finding indicated a possible tool for future study of the mode of action of thyroxine, the ultrastructural and biochemical effects of DPTH and/or thyroxine on rat liver mere investigated.Rats were fed either standard or DPTH (0.06%) diet for 30 days before T4 (250 ug/kg/day) was injected. Injection of T4 occurred daily for 10 days prior to sacrifice. After removal of the liver and kidneys, part of the tissue was frozen at -50°C for later biocheailcal analyses, while the rest was prefixed in buffered 3.5X glutaraldehyde (390 mOs) and post-fixed in buffered 1Z OsO4 (376 mOs). Tissues were embedded in Araldlte 502 and the sections examined in a Zeiss EM 9S.Hepatocytes from hyperthyroid rats (Fig. 2) demonstrated enlarged and more numerous mitochondria than those of controls (Fig. 1). Glycogen was almost totally absent from the cytoplasm of the T4-treated rats.

Characterization and Partial Purification of Heterodisperse Polysomal RNAs in Normal and Neoplastic Cells

1972 ◽  
Vol 58 (2) ◽  
pp. 71-94
Author(s):  
Ada Sacchi ◽  
Gianni Chinali ◽  
Susetta Pons ◽  
Michela Galdieri ◽  
Piero Cammarano
Keyword(s):  
Size Distribution ◽  
Density Gradient ◽  
Rat Liver ◽  
Sucrose Density Gradient ◽  
Partial Purification ◽  
Messenger Rnas ◽  
Alkaline Ph ◽  
Sedimentation Profile ◽  
Molecular Weights ◽  
Phenol Extraction

The size distribution of cytoplasmic messenger RNAs (m-RNA) has been studied in rat liver and in monodifferentiated cells (mouse reticulocytes and myelomas). It has been found that the RNA which exhibits a « rapid turnover » and a polydisperse profile of radioactivity is refractory to phenol extraction. This property has been exploited to selectively isolate m–RNA from the phenol residue by means of an extraction at an alkaline pH. The sucrose density gradient profiles of m–RNA isolated from monodifferentiated cells show monodisperse peaks having the sedimentation coefficients expected on the basis of the molecular weights of monocistronic messages for α and β chains of hemoglobin (reticulocytes) and L and H chains of immunoglobulin (myelomas). The sedimentation profile of cytoplasmic m–RNA associated with rat liver polysomes shows a much broader distribution, with sedimentation coefficients ranging from 8 S to 28 S.

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