bluuebunny/crossref_metadata_2025 huggingface dataset:
Metabolic phases during the development of granulation tissue
Authors: Lampiaho Kirsti , Kulonen E.
Published: August 2015

1. The metabolism of incubated slices of sponge-induced granulation tissue, harvested 4–90 days after the implantation, was studied with special reference to the capacity of collagen synthesis and to the energy metabolism. Data are also given on the nucleic acid contents during the observation period. Three metabolic phases were evident. 2. The viability of the slices for the synthesis of collagen was studied in various conditions. Freezing and homogenization destroyed the capacity of the tissue to incorporate proline into collagen. 3. Consumption of oxygen reached the maximum at 30–40 days. There was evidence that the pentose phosphate cycle was important, especially during the phases of the proliferation and the involution. The formation of lactic acid was maximal at about 20 days. 4. The capacity to incorporate proline into collagen hydroxyproline in vitro was limited to a relatively short period at 10–30 days. 5. The synthesis of collagen was dependent on the supply of oxygen and glucose, which latter could be replaced in the incubation medium by other monosaccharides but not by the metabolites of glucose or tricarboxylic acid-cycle intermediates.
The biosynthesis of phosphatidic acid and phosphatidylinositol in mammalian pancreas
Authors: Prottey C. , Hawthorne J. N.
Published: August 2015

1. The labelling of guinea-pig pancreas phospholipids in vivo after intraperitoneal injection of [32P]orthophosphate is described. 2. Acyl-CoA synthetase activity in pancreas homogenates has been studied. There is no absolute requirement for added fatty acids, indicating an adequate supply of endogenous fatty acids in these preparations. 3. Phosphatidic acid is formed in guinea-pig pancreas preparations by two distinct routes, namely the acylation of l-3-glycerophosphate and the phosphorylation of 1,2-diglyceride. Phosphatidic acid formed by either mechanism is converted into phosphatidylinositol by guinea-pig pancreas in vitro. 4. The enzymes of pancreas that convert phosphatidic acid into phosphatidylinositol via CDP-diglyceride have been characterized. 5. Addition of bovine serum albumin is necessary in assaying certain of these enzymes.
Subcellular distribution of cytochrome c in rat liver. Methods for its extraction and purification
Authors: González-Cadavid N. F. , Campbell P. N.
Published: August 2015

1. A method for the extraction and purification of cytochrome c from rat liver is described. The method depends on multiple chromatography on Amberlite IRC-50 with elution with ammonium phosphate buffers of differing ionic composition and pH, interspersed with gel filtration with Sephadex G-25. Conditions leading to denaturation are avoided and the product is chromatographically pure. 2. The method may be used for the quantitative analysis of cytochrome c either in unfractionated liver or in subcellular fractions. 3. Two pools of cytochrome c were detected, one extractable at pH4·0 with distilled water and the other extracted from the residues of the first extraction with 0·15m-sodium chloride. 4. For subcellular distribution studies the liver was homogenized in 0·3m-sucrose and a nuclear fraction (washed thoroughly to remove trapped mitochondria), a mitochondrial fraction, a heavy microsomal fraction, a standard microsomal fraction and the cell sap were isolated. The mitochondrial fraction was subfractionated further by density-gradient centrifugation. Each fraction was analysed for protein, RNA, DNA, succinate–neotetrazolium oxidoreductase and glucose 6-phosphatase. 5. A total of 123μg. of cytochrome c was obtained/g. wet wt. of rat liver. 6. Values for the percentage subcellular distribution of cytochrome c are: nuclear fraction, 24·4; mitochondrial fraction, 57·2; heavy microsomal fraction, 5·2; standard microsomal fraction, 10·6; cell sap, 2·7. 7. Three out of the eight mitochondrial subfractions separated by gradient centrifugation contained 76% of the cytochrome c and 85% of the succinate–neotetrazolium oxidoreductase present in the mitochondrial fraction. 8. In unfractionated liver 94% of the cytochrome c was extracted at pH4·0 with water whereas in most of the subcellular fractions the corresponding value was approx. 75–80%.
Purification and properties of a soluble reduced nicotinamide–adenine dinucleotide (phosphate) dehydrogenase from the hepatopancreas of Octopus vulgaris
Authors: Di Prisco G. , Casola L. , Giuditta A.
Published: August 2015

1. The oxidation of NADH and NADPH catalysed by the soluble supernatant from the hepatopancreas of Octopus vulgaris is due to a single enzyme, which has been purified approximately 100-fold. The enzyme reacts rapidly with potassium ferricyanide, and more slowly with 2,6-dichlorophenol-indophenol. No activity is obtained with oxygen, cytochrome c, lipoic acid, vitamin K1, vitamin K3, ubiquinone-30, p-benzoquinone, 2-p-iodophenyl-3-p-nitrophenyl-5-phenyltetrazolium chloride or methylene blue. 2. GSH, cysteine and mercaptoethanol stimulate the enzymic activity up to fivefold. GSSG is without any apparent effect. When stimulated by GSH the enzyme becomes sensitive to dicoumarol, which produces an inhibition competitive with respect to the activator. 3. The purified enzyme contains an acid-removable flavine component, which has been identified as FMN by spectrofluorimetry and chromatography in three solvent systems. After acid ammonium sulphate treatment the enzymic activity is lost, but it can be almost fully restored by incubation with FMN. FAD produces only a partial reactivation.
Studies on the enzymic degradation of L-serine O-sulphate by a rat liver preparation
Authors: Thomas J H , Tudball N.
Published: August 2015

1. The enzyme system of rat liver responsible for the degradation of l-serine O-sulphate was purified 300-fold and the optimum conditions for the activity were determined. 2. Inorganic sulphate, pyruvate and ammonia were found to be the products of enzyme action on lserine O-sulphate, being formed in equivalent amounts under all conditions examined. No free l-serine was detected as a product of enzyme action. 3. The enzyme preparation was free from other serine-metabolizing systems such as O-phospho-l-serine phosphatase and l-serine dehydratase. 4. The enzyme has a very narrow substrate specificity and is inactive towards a wide variety of related sulphate esters and amino acids. 5. Pyridoxal 5′-phosphate is capable of catalysing the non-enzymic breakdown of l-serine O-sulphate in the presence of metal salts to yield inorganic sulphate, pyruvate and ammonia as products. 6. The possible role of pyridoxal 5′-phosphate as a coenzyme in the enzymic degradation of l-serine O-sulphate is discussed.
Further properties and possible mechanism of action of adenosine 5′-triphosphate–d-glucose 6-phosphotransferase from rat liver
Authors: Parry M. J. , Walker D G
Published: August 2015

1. Magnesium ions are the most effective bivalent ions in the glucokinase reaction. 2. The molecular weight of rat hepatic glucokinase is 48000–49000 as assessed by gel filtration on Sephadex G-100. 3. Anomalous kinetic behaviour at low glucose concentrations appears to be due to the formation during the purification procedure of fragments possessing modified catalytic properties, but is unlikely to be of physiological significance. 4. Extension of previous studies (Parry & Walker, 1966) suggests that glucokinase catalyses a reaction of the random Bi Bi type similar to that of yeast hexokinase. 5. The inhibitory effects of various thiol reagents suggest that a thiol group may be involved at or near the binding site of the acceptor molecule.
Studies on amino acid decarboxylases in Escherichia coli
Authors: Lawson Alexander , Quinn A. G.
Published: August 2015

The isolation of highly active preparations of glutamate decarboxylase, arginine decarboxylase and histidine decarboxylase from Escherichia coli is described. These preparations showed strict specificity, and were in each case resolved into apo- and co-enzyme as shown by the significant restoration of activity that took place on addition of pyridoxal 5-phosphate.
Studies on glycogen synthesis in pigeon liver homogenates. Incorporation of hexose into glycogen
Authors: Nigam V. N. , Fridland A.
Published: August 2015

Liver homogenates of avian species, but not of mammals, form glycogen from glucose, mannose, fructose and galactose. Incorporation of labelled glucose, fructose and mannose, but not of labelled galactose, into glycogen is diluted isotopically by unlabelled glucose. Except for fructose, glycogen formation from other substrates by pigeon liver homogenates compares favourably with that from the same substrates in pigeon liver slices. Optimum conditions for glycogen synthesis from glucose by pigeon liver homogenate are: medium of incubation, 0·175m-sucrose–45mm-potassium chloride−15mm-glycylglycine buffer, pH 7·5; concentration of substrate, 15mm; concentration of tissue, less than 120mg./ml.; temperature of incubation, 37–43°; atmosphere, oxygen. Uncouplers of oxidative phosphorylation, Ca2+, EDTA, PPi, 2-deoxyglucose 6-phosphate and microsomal fraction of rat liver are inhibitory to glycogen synthesis from glucose. Starvation of pigeons for 24 and 48hr. leads to a slight stimulation of glycogen synthesis in their liver homogenates as compared with fed controls. Pigeon liver homogenates can be separated into subcellular fractions that on reconstitution can synthesize glycogen. All the enzymes of the glycogen pathway except soluble high-Km glucokinase are present in pigeon liver.
Similarities between the biochemical actions of cycasin and dimethylnitrosamine
Authors: Shank R. C. , Magee P. N.
Published: August 2015

1. Rats were given the hepatotoxin and carcinogen cycasin by stomach tube. In one experiment, rats whose RNA had previously been labelled with [14C]-formate were given the acetate ester of the aglycone form of cycasin, methylazoxymethanol, by intraperitoneal injection. 2. Incorporation of 14C from l-[U−14C]leucine into the proteins of some organs was measured in cycasin-treated rats. Cycasin inhibited leucine incorporation into liver proteins but not into kidney, spleen or ileum proteins. This inhibition was not evident until about 5hr. after cycasin administration, but once established it persisted for the next 20hr. 3. Methylation of nucleic acids was detected in some organs of rats treated with cycasin or methylazoxymethanol. The purine bases of RNA and DNA were isolated by acid hydrolysis followed by ion-exchange column chromatography. The resulting chromatograms showed an additional purine base that was identified as 7-methylguanine. It was shown that, in animals treated with the toxin, liver RNA was methylated to a greater extent than was either kidney or small-intestine RNA. Also, as a result of cycasin administration, liver DNA guanine was methylated to a greater extent than was RNA guanine. 4. These results are discussed in relation to comparable experiments with dimethylnitrosamine. It is suggested that cycasin and dimethylnitrosamine are metabolized to the same biochemically active compound, perhaps diazomethane, but that various tissues differ in their capacity to metabolize the two carcinogens.
An improved synthesis of isopentenyl pyrophosphate
Authors: Donninger C. , Popják G.
Published: August 2015

1. Isopentenol was converted into isopentenyl phosphate with phosphoryl chloride in ether containing pyridine. 2. The isopentenyl phosphate reacted in 2-methylpropan-2-ol–water with morpholine and dicyclohexylcarbodi-imide to give isopentenyl phosphoromorpholidate. 3. The isopentenyl phosphoromorpholidate, with inorganic phosphate in pyridine containing tributylamine, gave isopentenyl pyrophosphate. The yield of pyrophosphate from monophosphate was 80–85% and the yield of pyrophosphate from isopentenol 40–60%. [1−14C]-Isopentenyl pyrophosphate was prepared by this method. 4. The yield of isopentenyl pyrophosphate from isopentenyl phosphate was substantially improved, in comparison with the yields obtained by published methods via the phosphoramidate, by the use of the phosphoromorpholidate.
The protein–polysaccharide complex of bovine nasal cartilage. Studies on the protein core
Authors: Serafini-Fracassini A , Peters T. J. , Floreani L.
Published: August 2015

1. Chondromucoprotein from bovine nasal cartilage was purified by cetylpyridinium chloride or by bismuth nitrate in acetone. 2. Amino acid compositions of crude and purified preparations were compared and few differences were found, in spite of the decrease in protein content on purification. 3. Amino acid analysis of bismuth-purified material revealed the existence of four groups of amino acids. Within each group, the amino acids were present in approximately equimolar concentrations. 4. Amino end-group assay on the same material showed six α-DNP derivatives. 5. A molecular weight of 6·3×105 for the protein–polysaccharide complex was calculated from the latter analysis.
The metabolism of 7- and 12-methylbenz[a]-anthracene and their derivatives
Authors: Sims P
Published: August 2015

1. 7- and 12-Methylbenz[a]anthracene were converted by rat-liver homogenates into the corresponding hydroxymethyl derivatives, products that are probably the 8,9-dihydro-8,9-dihydroxy and the 5,6-dihydro-5,6-dihydroxy derivatives, and a number of phenolic products. 2. Both hydrocarbons were converted into glutathione conjugates; that from 7-methylbenz[a]anthracene was also formed, together with 5,6-dihydro-5,6-dihydroxy- and 5-hydroxy-benz[a]anthracene, from 5,6-epoxy-5,6-dihydro-7-methylbenz[a]anthracene. 3. 7- and 12-Hydroxymethyl-benz[a]anthracene were converted into products that are probably 8,9-dihydro-8,9-dihydroxy derivatives, and into phenols. 4. The preparation of a number of derivatives of the hydrocarbons is described. 5. The oxidation of the hydrocarbons with lead tetra-acetate was investigated.
A method for the selective extraction of histone fractions f2(a)1 and f2(a)2 from calf thymus deoxyribonucleoprotein at pH7
Authors: Johns E W
Published: August 2015

1. A new method has been developed for the specific extraction of histone fraction f2(a) from calf thymus deoxyribonucleoprotein at pH7 by using a mixture of ethanol and guanidinium chloride. 2. Fraction f2(a) has been separated into the subfractions f2(a)1 and f2(a)2 by acetone precipitation from acid solution, and at pH7. 3. Modifications of existing electrophoretic methods are described that enable these fractions to be more easily characterized.
Quantitative nucleic acid changes during phytohaemagglutinin-induced lymphocyte transformation in vitro. Dependence of the response on phytohaemagglutinin/serum ratio
Authors: Forsdyke D. R.
Published: August 2015

1. Pig and human blood lymphocytes have been grown in culture without replenishment of medium, and stimulated to transform by phytohaemagglutinin. Quantitative nucleic acid changes during this process have been used as an index of transformation. 2. On the first day, cells attach to glass; then they detach and continue transforming. 3. The degree of transformation is dependent on the phytohaemagglutinin/serum ratio, and is independent of cell concentration within the range 0·5×106−2·0×106 cells/ml. 4. At low phytohaemagglutinin/serum ratios there is no response; at high phytohaemagglutinin/serum ratios, inhibition appears after the cells have been cultured for a day.
Effect of mercurial compounds on structure-linked latency of lysosomal hydrolases
Authors: Verity M. Anthony , Reith A.
Published: August 2015

1. A partially purified lysosomal preparation was obtained from adult mouse livers by sucrose-density-gradient centrifugation of a large-granule fraction. 2. This lysosome-enriched subfraction was contaminated approx. 10% by mitochondrial cytochrome c oxidase and malate dehydrogenase. 3. Free acid phosphohydrolase and β-glucuronidase contributed less than 10% of the total (Triton X-100-solubilized) activity in contrast with approx. 30% free N-acetyl-β-d-glucosaminidase when assayed in an iso-osmotic incubation system. 4. Exposure of the lysosomal preparation to inorganic Hg2+ ions and organic mercurials (p-chloromercuribenzoate, phenylmercuric acetate) induced an irreversible loss of structure-linked latency with resulting enzyme activation. 5. Maximal activation was related to log [Hg2+] and pH. The response was all-or-none for individual particles; the dose–response curve portrayed the variation in particle resistance within the lysosomal population. 6. l-Cysteine and GSH totally prevented Hg2+ ion-induced hydrolase activation. Ascorbate provided approx. 50% protection. 7. The three lysosomal hydrolases were differentially activated at constant [Hg2+], suggesting a different pattern of binding, unique for each enzyme studied.
Metabolism of [1−14C]glyoxylate, [1−14C]-glycollate, [1−14C]glycine and [2−14C]-glycine by homogenates of kidney and liver tissue from hyperoxaluric and control subjects
Authors: Dean Betty M. , Watts R. W. E. , Westwick Wendy J.
Published: August 2015

1. The metabolism of [1−14C]glyoxylate to carbon dioxide, glycine, oxalate, serine, formate and glycollate was investigated in hyperoxaluric and control subjects' kidney and liver tissue in vitro. 2. Only glycine and carbon dioxide became significantly labelled with 14C, and this was less in the hyperoxaluric patients' kidney tissue than in the control tissue. 3. Liver did not show this difference. 4. The metabolism of [1−14C]glycollate was also studied in the liver tissue; glyoxylate formation was demonstrated and the formation of 14CO2 from this substrate was likewise unimpaired in the hyperoxaluric patients' liver tissue in these experiments. 5. Glycine was not metabolized by human kidney, liver or blood cells under the conditions used. 6. These observations show that glyoxylate metabolism by the kidney is impaired in primary hyperoxaluria.
A computer programme for the calculation of radioactivity data from liquid-scintillation counters fitted with external standards
Authors: Felts J M , Mayes P A
Published: August 2015

1. A method for transferring data from liquid-scintillation counters to automatic punched-card read-out is described. 2. A method of calibration of an external standard is presented that supplies sufficient information for the correction of quenched data by computer. 3. A computer programme in FORTRAN IV is described that calculates the data from one isotope, or two isotopes counted simultaneously, and that incorporates features for examining the reproducibility of the input data. 4. With the methods described, quench correction of samples containing either 3H or 14C was accurate until samples were quenched 70%. When the two isotopes were both present in one sample and counted simultaneously, the correction was accurate until the samples were quenched 60%. When quenching was greater than either 70 or 60% respectively, its correction was still possible but with slightly diminished accuracy and with greater variability.
Oxygen toxicity in Astasia
Authors: Bégin-Heick Nicole , Blum J. J.
Published: August 2015

1. Exposure of Astasia longa to oxygen+carbon dioxide (95:5) at atmospheric pressure leads to an inhibition of growth rate and of respiration. Growth resumes at the normal rate as soon as the oxygenation is discontinued, but respiration recovers more slowly. 2. Mitochondria prepared from cells exposed to oxygen+carbon dioxide (95:5) during growth have considerably decreased activities of succinate–cytochrome c oxidoreductase, NADH–cytochrome c oxidoreductase, succinate dehydrogenase and succinate oxidase activities as compared with mitochondria obtained from cells exposed to air+carbon dioxide (95:5). Cytochrome oxidase activity is not appreciably inhibited by exposure of the cells to 95% oxygen. 3. The mitochondrial fraction of Astasia contains rhodoquinone. The rhodoquinone concentration increases in cells exposed to 95% oxygen. The content of ergosterol-containing compounds also increases in the mitochondria of cells exposed to 95% oxygen. There is little change in the ubiquinone content of the mitochondrial fraction. The ubiquinone of Astasia appears to be ubiquinone-45.
Effect of thrombin on the radioactive nucleotides of human washed platelets
Authors: Ireland D M
Published: August 2015

Radioactive ATP and ADP were found in platelets after incubation of human platelet-rich plasma with either [8−14C]adenosine or [8−14C]ADP. Treatment of the labelled and washed platelets with thrombin indicated that, though considerable amounts of ATP and ADP were released to the supernatant, radioactive ATP and ADP remained predominantly in the cellular fraction. Breakdown of radioactive ATP took place to form mainly IMP and hypoxanthine, the latter compound appearing in the supernatant. The results indicate the presence of at least two pools of nucleotide in platelets. Evidence is given that the two pools contain approximately the same amounts of ATP plus ADP, and that the ratio of ATP to ADP in the pool released to the supernatant by the action of thrombin is about 0·7–0·8.
Effectors of rat-heart hexokinases and the control of rates of glucose phosphorylation in the perfused rat heart
Authors: England P J , Randle P J
Published: August 2015

1. The kinetic properties of the soluble and particulate hexokinases from rat heart have been investigated. 2. For both forms of the enzyme, the Km for glucose was 45μm and the Km for ATP 0·5mm. Glucose 6-phosphate was a non-competitive inhibitor with respect to glucose (Ki 0·16mm for the soluble and 0·33mm for the particulate enzyme) and a mixed inhibitor with respect to ATP (Ki 80μm for the soluble and 40μm for the particulate enzyme). ADP and AMP were competitive inhibitors with respect to ATP (Ki for ADP was 0·68mm for the soluble and 0·60mm for the particulate enzyme; Ki for AMP was 0·37mm for the soluble and 0·16mm for the particulate enzyme). Pi reversed glucose 6-phosphate inhibition with both forms at 10mm but not at 2mm, with glucose 6-phosphate concentrations of 0·3mm or less for the soluble and 1mm or less for the particulate enzyme. 3. The total activity of hexokinase in normal hearts and in hearts from alloxan-diabetic rats was 21·5μmoles of glucose phosphorylated/min./g. dry wt. of ventricle at 25°. The temperature coefficient Q10 between 22° and 38·5° was 1·93; the ratio of the soluble to the particulate enzyme was 3:7. 4. The kinetic data have been used to predict rates of glucose phosphorylation in the perfused heart at saturating concentrations of glucose from measured concentrations of ATP, glucose 6-phosphate, ADP and AMP. These have been compared with the rates of glucose phosphorylation measured with precision in a small-volume recirculation perfusion apparatus, which is described. The correlation between predicted and measured rates was highly significant and their ratio was 1·07. 5. These findings are consistent with the control of glucose phosphorylation in the perfused heart by glucose 6-phosphate concentration, subject to certain assumptions that are discussed in detail.
The composition of crystalline complexes of neurophysin-M with [8-arginine]-vasopressin and oxytocin
Authors: Hollenberg M D , Hope D B
Published: August 2015

Neurophysin-M, a methionine-containing protein that is the major constituent of neurophysin, has been crystallized as complexes with [8-arginine]-vasopressin. Three moles of vasopressin alone or 2 moles of vasopressin together with 1 mole of oxytocin are bound/mole of protein. An amorphous complex of the protein with oxytocin alone contains 2 moles of the hormone/mole of protein. Deamino-[8-arginine]-vasopressin, a highly active basic analogue of vasopressin, is not bound by neurophysin. The primary amino group of both vasopressin and oxytocin is necessary for binding with neurophysin.
The morphological site of synthesis of cytochrome c in mammalian cells (Krebs cells)
Authors: Freeman K. B. , Haldar D. , Work T. S.
Published: August 2015

In Krebs ascites-tumour cells, cytochrome c is segregated in the mitochondria and the level in microsomes could not be measured. At 22° in glucose–buffer Krebs cells synthesized a spectrum of proteins including cytochrome c. Mild osmotic shock in the presence of ribonuclease had little effect on incorporation of [14C]-leucine or [14C]valine into mixed mitochondrial protein but strongly inhibited synthesis of non-mitochondrial cytoplasmic proteins. Under these conditions, labelling of cytochrome c was also strongly inhibited. After pulse labelling of Krebs cells at 22° for 10min. the cytcchrome radioactivity found in mitochondria was higher than in microsomes. After addition of unlabelled amino acid as ‘chase’ there was 137% increase in radioactivity of cytochrome c but only a 3% increase in radioactivity of whole-cell protein. It is concluded that the peptide chain of cytochome c is synthesized on cytoplasmic ribosomes. Mitochondria therefore do not have the character of self-replicating entities, but are formed by the cooperative function of messenger RNA of cytoplasmic ribosomes and, possibly, of intramitochondrial messenger derived from the mitochondrial DNA.
The effect of ultraviolet irradiation on acid-soluble collagen
Authors: Davidson R. J. , Cooper D. R.
Published: August 2015

1. A study has been made of the effect of ultraviolet irradiation on the conformational changes taking place in cooled solutions of thermally denatured acid-soluble calf-skin collagen. 2. The increase in negative rotation and viscosity at 15° for irradiated and thermally denatured collagen solutions becomes less as the irradiation dose is increased. 3. The principal effect of ultraviolet irradiation is the fission of the primary collagen chains, eventually yielding chain lengths incapable of stabilizing a helical structure. 4. The effects of ultraviolet irradiation on acid-soluble collagen may be closely correlated with similar effects on neutral salt-soluble collagen.
Synthesis of ribonucleic acid in baby-hamster kidney cells in the presence of actinomycin D
Authors: Martin S J , Brown F.
Published: August 2015

1. RNA molecules with sedimentation values in sucrose gradients of 12–20s are synthesized in baby-hamster kidney cells even after prolonged incubation in medium containing 1μg. of actinomycin D/ml. 2. The rate of formation of this RNA is dependent on the age of the cultures and is greatest during the exponential phase of growth. 3. Growth of cells on nutritionally poor medium causes degradation and inhibits the synthesis of these RNA fractions. 4. Replacement of the nutritionally poor medium with a rich medium stimulates the synthesis of actinomycin-resistant RNA. This stimulation is blocked by cycloheximide. 5. The base composition of this RNA is characterized by low cytidine and high guanosine values.
Effect of virus infection on the stability and synthesis of actinomycin-resistant ribonucleic acid in baby-hamster kidney cells
Authors: Martin S J , Brown F.
Published: August 2015

1. The sucrose-gradient pattern of 32P-labelled RNA synthesized in actinomycintreated baby-hamster kidney cells infected with foot-and-mouth-disease virus depends greatly on the period of labelling. 2. Fractions are formed in infected cells that sediment at 12–20s and have the same base composition as similar fractions found in non-infected cells that have been treated with actinomycin. 3. In the presence of guanidine, which completely inhibits viral RNA synthesis, these fractions are labelled to a greater extent than in non-infected cells.
Influence of hormones on the nicotinamide nucleotide coenzymes of adipose tissue
Authors: McLean Patricia , Greenbaum A. L. , Brown J. , Greenslade K. R.
Published: August 2015

The concentrations of the oxidized and reduced forms of the nicotinamide nucleotides were measured in the epididymal fat pads of normal, alloxan-diabetic and hypophysectomized rats. In both alloxan-diabetic rats and hypophysectomized rats the weight of the adipose tissue fell, as did the total content of NADH and NADPH; in addition, NAD+ was decreased in the alloxan-diabetic group. Of these changes the most marked was in NADPH and this was the only significant difference when the results were expressed as nicotinamide nucleotides/mg. of tissue protein. The concentration of NADPH in the hypophysectomized rats was not altered by treatment with growth hormone but was restored to normal by treatment with thyroxine. These results are discussed in relation to the known effect of these hormonal conditions on lipid synthesis in adipose tissue.
The preparation and immunological properties of 131I-labelled adrenocorticotrophin
Authors: Landon J. , Livanou Theodora , Greenwood F. C.
Published: August 2015

1. A procedure is described for preparing 131I-labelled adrenocorticotrophin suitable for use in radioimmunoassay. 2. Adsorption of labelled and unlabelled adrenocorticotrophin at low concentrations occurs to various surfaces despite the presence of diluent protein. Adsorption and desorption errors are minimized by low pH and by the use of polystyrene vials. 3. Preparations with low initial damage are obtained if the radioiodination is performed rapidly and the separation of 131I-labelled adrenocorticotrophin from unchanged [131I]iodide is carried out on cellulose columns by using dilute acid. 4. The immunological activity of 131I-labelled α1–24-adrenocorticotrophin, but not of 131I-labelled porcine adrenocorticotrophin, decreases with increasing specific radioactivity. The involvement of tyrosine residues in the immunological specificity of the α1–24-adrenocorticotrophin only is suggested to explain this finding.
Regulation of polyribosome formation and protein synthesis in the uterus. Effect of ovariectomy and administration of oestradiol-17β on the amino acid-incorporation activity in vitro and the cytoplasmic concentration in vivo of polyribosomal preparation
Authors: Teng Ching-Sung , Hamilton Terrell H.
Published: August 2015

1. The hormonal regulation of cytoplasmic protein synthesis in the uterus is described. Polyribosomal preparation from uteri of normal or ovariectomized rats was isolated by procedure 3 and assayed for [14C]leucine-incorporation activity in the cell-free system, as described by Teng & Hamilton (1967). 2. Ovariectomy of normal animals caused, 3 weeks after surgery, a 50–60% increase in the amino acid-incorporation activity in vitro of uterine polyribosomal preparation, but a 90–95% decrease in the cytoplasmic concentration in vivo of the preparation. 3. Administration of 10μg. of oestradiol-17β to ovariectomized rats at zero time caused, 10–12hr. later, a 100% stimulation in amino acid-incorporation activity in vitro of the uterine polyribosomal preparation. From 12hr. to 36hr. after hormone administration, the activity in vitro of the preparation decreased. If a second dose of hormone was administered at 36hr., the activity in vitro of the preparation continued to decrease, and approached at 48hr. and 72hr. the lower activity observed for the preparation from normal animals. The cytoplasmic concentration of polyribosomal preparation increased by 600–700% under these experimental conditions. If a second dose of oestradiol-17β was not administered at 36hr., the initially elevated cytoplasmic concentration of the preparation decreased by 50% from 36hr. to 72hr., and the activity in vitro of the preparation was not fully depressed to the ‘normal’ value. 4. Pretreatment of ovariectomized animals with actinomycin D or cycloheximide abolished 80–90% of the stimulatory effects of hormone treatment on the amino acid-incorporation activity in vitro and cytoplasmic concentration in vivo of uterine polyribosomal preparation. 5. Two major conclusions are drawn from the results reported: that during early oestrogen action new polyribosomes having amino acid-incorporation properties different from those of the old ones appear and accumulate in the cytoplasm of the uterus; and that the regulation of cytoplasmic protein synthesis in the organ by oestrogen is of an indirect nature, with dual effects of the hormone on genetic transcription resulting in turn in a regulation of the rate and amount of genetic translation.
The disulphide bridges of bovine chymotrypsinogen B
Authors: Smillie L. B. , Hartley B S
Published: August 2015

The five cystine peptides of chymotrypsinogen B have been isolated as pairs of cysteic acid peptides by the diagonal electrophoretic technique of Brown & Hartley (1963, 1966). The sequences of these peptides have been shown to be very similar to those of chymotrypsinogen A, and indicate that both zymogens have the same pattern of disulphide bridges. The determination of the sequence of residues 14–21 in chymotrypsinogen B has shown that residue 18 differs in the chymotrypsinogens from the corresponding residue in trypsinogen. Eight differences between chymotrypsinogens A and B are found in sequences accounting for 72 of the 245 residues.
A diagonal paper-electrophoretic technique for studying amino acid sequences around the cysteine and cystine residues of proteins
Authors: Perham R N
Published: August 2015

1. A diagonal electrophoretic technique for studying the amino acid sequence around cysteine and cystine residues in proteins is described. The residues are first converted into S-aminoethylcysteine, and the protein is then treated with S-ethyl trifluorothioacetate, which trifluoroacetylates all the protein amino groups. The modified protein is digested enzymically and the resulting peptides are separated by paper electrophoresis. After exposure of the peptides on the paper to ammonia vapour, the electrophoresis is repeated, this time at right angles to the original direction. Peptides from which a trifluoroacetyl group is removed by the ammonia treatment will vacate the 45° diagonal formed by all other unaffected peptides owing to the exposure of an additional amino group and consequent increased electrophoretic mobility towards the cathode. Peptides containing lysine or S-aminoethylcysteine are readily purified by this technique. 2. The successful application of the technique to bovine insulin is described. 3. Various methods for distinguishing peptides containing lysine from those containing S-aminoethylcysteine in more complicated proteins are suggested and discussed.
Effect of alloxan-diabetes on multiple forms of hexokinase in adipose tissue and lung
Authors: McLean Patricia , Brown J. , Walters Eileen , Greenslade K.
Published: August 2015

Comparison has been made of the effect of alloxan-diabetes on the multiple forms of hexokinase (EC 2.7.1.1) in adipose tissue and lung. Types I and II hexokinase were distinguished in adipose tissue by their different stabilities to heat treatment, which made it possible to determine the activity of each form spectrophotometrically; additional confirmatory evidence was obtained from starch-gel electrophoresis. Type II hexokinase was markedly depressed in adipose tissue from alloxan-diabetic rats. Lung contained types I, II and III hexokinase, type I predominating. There was no significant change in the pattern of these multiple forms of hexokinase in lung from alloxan-diabetic rats. These results are discussed in relation to current ideas that the insulin-sensitivity of a tissue may be correlated with the content of type II hexokinase.
The intracellular distribution of free nucleotides in the tobacco leaf. Formation of adenosine 5′-phosphate from adenosine 5′-triphosphate in the chloroplasts
Authors: Keys A J
Published: August 2015

1. A method of studying the free nucleotides of leaf tissue is outlined and the need for a metal ion-binding agent for the complete extraction of certain nucleotides by aqueous ethanol is established. 2. The method was used to study the effect of illumination or darkening of tobacco plants on the free nucleotides present in the chloroplast and non-chloroplast tissue components. 3. When plants that had been in the dark for a prolonged period were given 30sec. of bright light there was a rapid phosphorylation of ADP to ATP in both chloroplast and non-chloroplast components of the tissue. 4. Where plants were moved into the dark from conditions suitable for rapid photosynthesis there was a rapid conversion of ATP into AMP and the AMP was formed only in the chloroplast fraction. In continuing darkness the AMP remained restricted mainly to the chloroplast fraction for at least 2min., but eventually its concentration fell to the low value that is typical of tobacco leaves during conditions of constant illumination. If the plants were returned to the light for 30sec. the AMP was rapidly rephosphorylated.
Effect of endotoxin on plasma albumin and fibrinogen synthesis rates in rabbits as measured by the [14C]carbonate method
Authors: Koj A. , McFarlane A. S.
Published: August 2015

1. Rates of synthesis of plasma albumin and fibrinogen were measured by the [14C]carbonate method in normal rabbits and in animals that received a single intravenous injection of Shigella endotoxin 14–48hr. earlier. 2. The accuracy of the method was improved by introducing refinements into procedures for measuring 14C radioactivities associated with both urea and proteins that are lost from the plasma during the synthesis interval. 3. The synthesis interval (time between injecting carbonate and measuring specific radioactivities of protein guanidine carbon in plasma) can be shortened with advantage to 3–4hr. 4. Injection of endotoxin markedly decreased the fractional rate of loss in the first few hours of injected radioiodine-labelled fibrinogen and to a smaller extent of similarly labelled albumin from the plasma. The absolute rate of synthesis of fibrinogen was increased in endotoxin-treated rabbits by more than 400% compared with normal animals, and the rate of synthesis of albumin was increased by about 60%.
Stimulation of fatty acid synthesis in vitro by gonadotrophin-induced testicular ribonucleic acid
Authors: Goswami Ajit , Skipper James K. , Williams William L.
Published: August 2015

RNA from testes of hypophysectomized rats treated with follicle-stimulating hormone and luteinizing hormone markedly stimulates in vitro the incorporation of acetate and malonate (as CoA derivatives) into polyunsaturated fatty acids. The system in vitro contains the components necessary for both protein and fatty acid synthesis. That the RNA is a hormone-induced messenger type that causes enzyme synthesis that then causes fatty acid synthesis is supported by the following observations: (1) the stimulation of RNA synthesis by follicle-stimulating hormone and luteinizing hormone is decreased by injection of the animals with actinomycin D; (2) puromycin in the system in vitro decreases the synthesis of polyunsaturated fatty acids; (3) the activity of the RNA preparation is destroyed by digestion with ribonuclease; in fact, the digest is inhibitory, which is a characteristic of messenger-RNA-mediated protein synthesis; (4) protein that might be denatured enzyme is virtually absent from the effective RNA preparations.
Further purification and characterization of the acid α-glucosidase
Authors: Auricchio F. , Bruni C. B. , Sica V.
Published: August 2015

1. Centrifugation of rat liver acid glucosidase, which had been purified by adsorption on dextran gel, on a density gradient of sucrose showed the enzyme to be impure. 2. Preliminary purification of the enzyme before the gel filtration improved the final degree of purity of this preparation. Disc gel electrophoresis of this preparation showed a single band of protein. 3. The sedimentation co-efficient and the molecular weight determined on a sucrose gradient were 4·9–5·1s and 76000–83000 respectively for the rat liver enzyme, and 5·6s and 97000 for the acid α-glucosidase purified by means of the same procedure from the human kidney. 4. The Michaelis constants of rat liver and human kidney enzyme were 4·7×10−3m and 13·6×10−3m respectively with maltose as substrate. 5. The enzyme from both tissues was inhibited by tris and by erythritol. The inhibition of the rat liver acid glucosidase by erythritol was competitive.
The chemical synthesis of glucosaminylphosphatidylglycerol. Comparison with a new phospholipid isolated from Bacillus megaterium
Authors: Gurr M I , Bonsen P. P. M. , Den Kamp J. A. F. OP , Van Deenen L. L. M.
Published: August 2015

1. We describe the synthesis of a glucosamine derivative of phosphatidylglycerol having the same structure as that of the natural compound isolated from Bacillus megaterium. 2. 2-O-(3,4,6-Tri-O-acetyl-2-deoxy-2-phthalimido-d-glucopyranosyl)-3-O-benzyl-1-iodo-sn-glycerol was prepared by a Königs–Knorr condensation between 3-O-benzyl-1-toluene-p-sulphonyl-sn-glycerol and 3,4,6-tri-O-acetyl-1-bromo-2-deoxy-2-phthalimido-d-glucopyranose followed by replacement of the toluene-p-sulphonyl group with iodine. The iodide was treated with the silver salt of 2-isolauroyl-1-oleoyl-sn-glycerol 3-(monobenzyl hydrogen phosphate) to form the fully protected phosphoglycolipid. 3. Removal of benzyl protecting groups by catalytic hydrogenolysis, phthaloyl group with hydrazine and acetyl groups with pH10 buffer furnished 2-O-(2-amino-2-deoxy-d-glucopyranosyl)-1-(2-isolauroyl-1-stearoyl-sn-glycero-3-phosphoryl)-sn-glycerol. 4. The synthetic and natural compounds appeared identical when compared by chromatography and by identification of hydrolysis products from chemical and enzymic degradations.
Studies on ferrochelatase. The enzymic formation of haem in proplastids, chloroplasts and plant mitochondria
Authors: Porra R. J. , Lascelles June
Published: August 2015

1. Ferrochelatase was demonstrated in the chloroplasts and proplastids isolated from the primary leaves of beans (a dicotyledon) and oats (a monocotyledon). It was also detected in chloroplasts from etiolated bean seedlings made green by illumination before being harvested. The specific activities of the three types of bean organelles are similar, as are the specific activities of the oat proplastids and chloroplasts. 2. Chloroplasts from young spinach leaves also contain ferrochelatase; these chloroplasts were tested for their ability to form magnesium tetrapyrroles and found unable to catalyse the insertion of Mg2+ into mesoporphyrin IX. 3. Ferrochelatase was also detected in potato tuber mitochondria. 4. Ferrochelatase activity in these plant preparations is much less stable on storage than similar preparations from bacteria and animal tissues. 5. Temperature affects the activities of spinach chloroplast ferrochelatase and rat liver ferrochelatase differently. Activity of the chloroplast enzyme increases as the temperature rises from 20·6° to 26°, but becomes increasingly inactivated as the temperature rises further to 38°. The initial velocity of the mammalian enzyme, however, increases as the temperature rises from 25·8° to 65°, but the enzyme is inactivated after several minutes at 65°.
Specific dimerization of the light chains of human immunoglobulin
Authors: Stevenson G. T. , Straus D.
Published: August 2015

1. The light chains of human immunoglobulin were allowed to dimerize in vitro on removal of the dispersing agents acetic acid or urea. 2. On electrophoresis in polyacrylamide gel at pH8·8 the dimers yielded up to nine regularly spaced bands. This approximates to the number of electrophoretic components known to occur among the monomers. 3. Single electrophoretic components of the dimers were isolated from the gel, dissociated into monomers, and subjected as such to electrophoresis in urea-containing gels. Each gave two adjacent bands. 4. Similarly, after all the light chains as monomers had been subjected to electrophoresis in urea-containing gels, single electrophoretic components were isolated and allowed to dimerize. When examined now as dimers in the absence of urea, each component gave two adjacent bands. 5. These findings are explicable on the following basis. (a) The dimerization of the light chains is specific, at least inasmuch as it occurs between monomers of the same electrophoretic mobilities. (b) With the buffer constant, different light chains undergo different changes in net charge on being transferred from urea-containing to urea-free solution; in this way two different chains of the same initial charge can acquire a charge difference of 1. 6. Experiments with Bence-Jones proteins and other homogeneous light chains gave results substantiating the conclusions (a) and (b).
Correlated changes of some enzyme activities and cofactor and substrate contents of pea cotyledon tissue during germination
Authors: Brown A. P. , Wray J. L.
Published: August 2015

1. The activities of six enzymes (hexokinase, phosphoglucose isomerase, phosphofructokinase, aldolase, glucose 6-phosphate dehydrogenase and amylase) in extracts of pea cotyledons were determined. The activities during the first 10 days after germination showed individual and characteristic changes that indicate a specific control of both synthesis and destruction of enzymes. 2. Tissue contents of glucose, inorganic phosphate, glucose 6-phosphate, fructose 6-phosphate, ATP, ADP, AMP, NAD and NADP were also determined, and a correlation is reported between the substrate concentrations at day 1 and the subsequent enzymic activity. 3. The initial NAD+/NADH ratio value of 1 changed to about 3 by day 4; the NADP content was lower and changes in the oxidation state were less striking. The ratio of ATP to ADP and AMP remained virtually constant.
A comparison of the effects of NN′-dicyclohexylcarbodi-imide, oligomycin A and aurovertin on energy-linked reactions in mitochondria and submitochondrial particles
Authors: Roberton A. M. , Holloway Caroline T. , Knight I G , Beechey R B
Published: August 2015

1. The effects of dicyclohexylcarbodi-imide, oligomycin A and aurovertin on enzyme systems related to respiratory-chain phosphorylation were compared. Dicyclohexylcarbodi-imide and oligomycin A have very similar functional effects, giving 50% inhibition of ATP-utilizing and ATP-generating systems at concentrations below 0·8nmole/mg. of submitochondrial-particle protein. Aurovertin is a more potent inhibitor of ATP synthesis, giving 50% inhibition at 0·2nmole/mg. of protein. However, aurovertin is a less potent inhibitor of ATP-utilizing systems: the ATP-driven energy-linked nicotinamide nucleotide transhydrogenase is 50% inhibited at 3·0nmoles/mg. of protein and the ATP-driven reduction of NAD+ by succinate is 50% inhibited at 0·95nmole/mg. of protein. 2. With EDTA-particles (prepared by subjecting mitochondria to ultrasonic radiation at pH9 in the presence of 2mm-EDTA) the maximum stimulation of the ATP-driven partial reactions is effected by similar concentrations of oligomycin A and dicylcohexylcarbodi-imide, but the latter is less effective. The stimulatory effects of suboptimum concentrations of dicyclohexylcarbodi-imide and oligomycin A are additive. Aurovertin does not stimulate these reactions or interfere with the stimulation by the other inhibitors. 3. Dicyclohexylcarbodi-imide and oligomycin A stimulate the aerobic energy-linked nicotinamide nucleotide transhydrogenase of EDTA-particles, but the optimum concentration is higher than that required for the ATP-driven partial reactions. Aurovertin has no effect on this reaction. 4. The site of action of dicyclohexylcarbodi-imide is in CF0, the mitochondrial fraction that confers oligomycin sensitivity on F1 mitochondrial adenosine triphosphatase.