bluuebunny/crossref_metadata_2025 huggingface dataset:
Some enzymic activities and chemical properties of the mesosomes and cytoplasmic membranes of Bacillus licheniformis 6346
Authors: Reaveley D. A. , Rogers H. J.
Published: August 2015

1. The liberation of the vesicles of the mesosomes from protoplasts of Bacillus licheniformis strain 6346 is controlled by the Mg2+ concentration present during the removal of the walls by lysozyme. 2. The functioning of the cytoplasmic membranes is also critically controlled by the Mg2+ concentration. 3. The isolated mesosomes and the cytoplasmic membranes differ in their enzymic activities. Both succinate dehydrogenase and NADH oxidase activities are very low or absent from the mesosomes. 4. The cytoplasmic membranes can also be separated into materials of different density. 5. Distribution of the membranes between these fractions is controlled by the Mg2+ concentration and the growth conditions of the microorganisms.
The metabolism of hexachlorocyclohexanes and pentachlorocyclohexenes in flies and grass grubs
Authors: Clark A G , Murphy Sue , Smith J. N.
Published: August 2015

1. γ-Hexachlorocyclohexane, γ-pentachlorocyclohexene and δ-pentachlorocyclohexene were converted by houseflies and grass grubs into metabolites that had chromatographic properties identical with those of S-2,4-dichlorophenylglutathione. 2. The metabolism of γ-hexachlorocyclohexane and the pentachlorocyclohexene isomers was negligible in newly emerged blowflies, but increased over the next 10 days. 3. The metabolism of both γ-hexachlorocyclohexane and the pentachlorocyclohexene isomers was inhibited by simultaneous dosage with tetrabromophenolphthalein ethyl ester or Bromophenol Blue in both grass grubs and flies, but only the metabolism of pentachlorocyclohexenes in blowflies was stopped by simultaneous dosage with bis-(N-dimethylaminophenyl)methane. NN-Di-n-butyl-p-chlorobenzenesulphonamide had no effect on the metabolism of pentachlorocyclohexenes by blowflies. 4. The use of these inhibitors and colorimetric assays leads to the conclusion that a pentachlorocyclohexene is not a major intermediary metabolite of γ-hexachlorocyclohexane in these insects.
The binding of spermine to the ribosomes and ribosomal ribonucleic acid from Bacillus stearothermophilus
Authors: Stevens L.
Published: August 2015

1. The total intracellular concentrations of Na+, K+, Mg2+, spermine, spermidine and RNA were measured in Bacillus stearothermophilus. 2. The binding of spermine to ribosomes and to ribosomal RNA from B. stearothermophilus was studied under various conditions by using a gel-filtration technique. 3. The affinity of spermine for ribosomes and for ribosomal RNA decreased with increasing ionic strength of the medium in which they were suspended. 4. The extent of spermine binding did not change appreciably in the temperature range 4–60°. 5. Optimum binding occurred at about pH7·0. 6. The number of binding sites for spermine on either ribosomes or ribosomal RNA was 0·10–0·13/RNA phosphate group. 7. A high proportion of the intracellular spermine is likely to be bound to the ribosomes in vivo; spermine competes with Mg2+ on equal terms for sites on the ribosomes.
Mercapturic acid formation during the metabolism of arecoline and arecaidine in the rat
Authors: Boyland E. , Nery R.
Published: August 2015

1. In the rat, arecoline is converted into arecaidine and both compounds are converted into N-acetyl-S-(3-carboxy-1-methylpiperid-4-yl) -l-cysteine. 2. The structure of the metabolite was established by (a) synthesis, (b) conversion into N-acetyl-S-(3-methoxycarbonyl-1-methylpiperid-4-yl) -l-cysteine methyl ester, which was chromatographically identical with the synthetic material, and (c) n.m.r.-and i.r.-spectral analysis of the 3-methoxycarbonyl derivative. 3. In ethanolic solution, or in phosphate buffer at pH7·0, arecoline reacted with N-acetyl-l-cysteine to give N-acetyl-S-(3-methoxycarbonyl-1-methylpiperid-4-yl) -l-cysteine; under similar conditions, arecaidine reacted more slowly to give N-acetyl-S-(3-carboxy-1-methylpiperid-4-yl) -l-cysteine. 4. The reaction between arecoline and glutathione or N-acetyl-l-cysteine occurred maximally at neutral pH and decreased rapidly with increasing acidity. At neutral pH, the reactions were bimolecular and second-order when the reactants were in approximately equimolar concentrations and pseudo-unimolecular first-order when arecoline was in large excess. 5. Consideration of the pKa values and degrees of ionization of the reactants and the effect of pH on the stoicheiometry of reaction between arecoline and glutathione or N-acetyl-l-cysteine indicated that reaction between un-ionized species occurred more readily than nucleophilic addition (AdN) reactions involving charged intermediates.
Control of chloroplast formation in Euglena gracilis. Antagonism between carbon and nitrogen sources
Authors: Harris R. C. , Kirk J. T. O.
Published: August 2015

1. Cells of Euglena gracilis grown in the dark on high ratios of carbon source to nitrogen source (‘high-carbon cells’) are unable to form chlorophyll during a subsequent incubation in the light; cells grown in the dark on low ratios of carbon to nitrogen (‘low-carbon cells’) synthesize chlorophyll at a rapid rate during the subsequent incubation in the light. High-carbon cells will form chlorophyll rapidly if supplied with a nitrogen source during the incubation in the light: of the nitrogen sources tested, ammonium sulphate was the most effective at overcoming the block in chlorophyll synthesis. The nitrogen source does not have to be present during the actual incubation in the light: a 5hr. exposure of high-carbon cells to ammonium sulphate in the dark, followed by removal of the nitrogen source, is sufficient to bring about rapid chlorophyll synthesis during a subsequent incubation in the light. 2. The synthesis of chlorophyll by low-carbon cells exposed to the light is strongly repressed by the addition of ethanol or other utilizable carbon sources during the incubation in the light. Chlorophyll synthesis ceases altogether between 5 and 10hr. after the addition of the carbon source. Carotenoid synthesis is also inhibited, but to a smaller extent. The inhibitory effects of ethanol are prevented if ammonium sulphate is added at the same time. 3. High-carbon cells contain about four times as much carbohydrate per cell and about twice as much lipid per cell as low-carbon cells. The content per cell of total protein, soluble protein and DNA are about the same in both types of cell. The low-carbon cells sometimes, but not always, contain more RNA than the high-carbon cells. Analysis of cold-acid extracts indicates that the two kinds of cells contain about the same concentrations of pool amino acids, but that the low-carbon cells contain somewhat higher concentrations of peptides in the pool. Ion-exchange analysis of pool extracts shows a number of differences between high-carbon and low-carbon cells with respect to the concentrations of individual amino acids: in particular low-carbon cells contain higher concentrations of alanine. High-carbon cells have approximately twice as much protease activity as low-carbon cells. 4. The possible biochemical basis for the differing ability of high-carbon and low-carbon cells to form chloroplasts in the light is discussed.
Steroid metabolism in the cat. Biliary and urinary excretion of metabolites of [4−14C]cortisone
Authors: Taylor W.
Published: August 2015

1. [4−14C]Cortisone was administered to anaesthetized male cats as a single injection or as a 45–60min. infusion. 2. After the single dose a total of 69·6–89·6% of the radioactivity was excreted in bile, and 0·5–7·1% in urine. After infusion total recovery in bile was 73·4–92·1%, and 1·2–1·7% in urine. 3. When bile and urine samples were hydrolysed successively by β-glucuronidase, cold acid and hot acid, most of the radioactivity was converted into substances not extractable from neutral aqueous solution by ethyl acetate–ether. 4. In bile, metabolites hydrolysable by β-glucuronidase were found in only small proportions (3–4%) of the dose.
Pulse radiolysis and spectral studies of the interaction of cetylpyridinium chloride and Methylene Blue with connective-tissue glycosaminoglycans and related compounds
Authors: Davies J. V. , Dodgson K S , Moore J. S. , Phillips G. O.
Published: August 2015

1. Hydrated electrons produced by pulse radiolysis were used to study the interaction of polyanionic glycosaminoglycans and related compounds with the counterions Methylene Blue and cetylpyridinium chloride. 2. The effect of added salt (potassium chloride) on the interaction indicates that the relative binding affinities, with respect to the types of anionic site present, increases for both counterions in the order CO2−<CO2−+O·SO3−<CO2−+O·SO3− +N·SO3−<O·SO3−. The interactions of the polyanions with cetylpyridinium chloride are considerably stronger than those with Methylene Blue. 3. The effects of added salt on the metachromasia resulting from polyanion–Methylene Blue interaction were examined spectrophotometrically. 4. The collective results demonstrate a direct relationship between anionic site–dye binding and metachromasia, although a residual dye binding can be detected by pulse radiolysis when metachromasia is completely removed.
The interaction of various inhibitors and stimuli of insulin release studied with rabbit pancreas in vitro
Authors: Milner R. D. G. , Hales C. N.
Published: August 2015

The effect of various inhibitors on insulin release from pieces of rabbit pancreas incubated in vitro was studied. Insulin release was stimulated by glucose (3mg./ml.), leucine (5mm), tolbutamide (200μg./ml.), ouabain (10μm), a raised extracellular K+ concentration (60mm) and substitution of the Ca2+ content of the incubation medium by Ba2+ (2·5mm). (a) Mannoheptulose (6mg./ml.) inhibited glucose-stimulated insulin release only. (b) Anoxia abolished or inhibited insulin release stimulated by glucose, leucine, tolbutamide and K+, but had little or no effect on release stimulated by ouabain or Ba2+. (c) 2,4-Dinitrophenol (0·25mm) abolished or inhibited insulin release stimulated by glucose, ouabain or Ba2+. (d) Diazoxide (250μg./ml.) abolished or inhibited insulin release stimulated by glucose, leucine, tolbutamide, ouabain or Ba2+ (0·25 or 1mm). Diazoxide had no effect on insulin release stimulated by Ba2+ (2·5mm) and potentiated release stimulated by K+. (e) Adrenaline (1μm) abolished insulin release stimulated by glucose, leucine, tolbutamide, ouabain or Ba2+. K+-stimulated release was inhibited by adrenaline. (f) Tetrodotoxin (1μm) had no effect on insulin release stimulated by glucose, leucine, tolbutamide, ouabain, K+ or Ba2+. (g) Nupercaine (1mm) abolished insulin release stimulated by glucose or Ba2+.
Cleavage of bacterial flagellin with cyanogen bromide. Antigenic properties of the protein fragments
Authors: Parish C. R. , Wistar R. , Ada G. L.
Published: August 2015

1. Four polypeptide fragments, obtained by cyanogen bromide treatment of the protein flagellin from Salmonella adelaide, were tested for their antigenic activity by using them as inhibitors in three different assays: bacterial immobilization, haemagglutination of sensitized erythrocytes and quantitative micro precipitation. Immunodiffusion studies were also performed on the protein fragments. 2. Cleavage of the flagellin molecule in this way gave no detectable loss of antigenic determinants. Fragment A (mol.wt. 18000), the largest of the polypeptides, contained all the antigenic specificities present on flagellin that were recognized by the antisera used. In one test, fragment B (mol.wt. 12000) also contained antigenic activity to an extent not easily explainable by contamination with fragment A. Fragments C (mol.wt. 5500) and D (mol.wt. 4500) appeared to be antigenically inactive.
The metabolism of aromatic compounds by Rhodopseudomonas palustris. A new, reductive, method of aromatic ring metabolism
Authors: Dutton P L , Evans W C
Published: August 2015

1. Rhodopseudomonas palustris grows both aerobically and photosynthetically on aromatic acids. p-Hydroxybenzoate and protocatechuate are able to support aerobic growth; these compounds are metabolized by the protocatechuate 4,5-oxygenase pathway. 2. The photoassimilation of benzoate and hydroxybenzoates and the effects of air and darkness on the photoassimilation of benzoate are described. 3. Evidence in conflict with the pathway previously proposed for the photometabolism of benzoate is discussed. 4. The photometabolism of benzoate is accomplished by a novel reductive pathway involving its reduction to cyclohex-1-ene-1-carboxylate, followed by hydration to 2-hydroxycyclohexanecarboxylate and after dehydrogenation to 2-oxocyclohexanecarboxylate further hydration results in ring-fission and the production of pimelate. 5. Attempts were made to prepare cell-free extracts capable of dissimilating benzoate.
Crystalline 3-phosphoglycerate kinase from skeletal muscle
Authors: Scopes R. K.
Published: August 2015

1. A procedure for preparing crystalline 3-phosphoglycerate kinase from rabbit or pig skeletal muscle is presented. 2. The preparation phosphorylates up to 975μmoles of 3-phosphoglycerate/min./mg. at 30° and is not contaminated with myokinase. 3. The enzyme has an estimated molecular weight of 36500±1000, and contains three residues each of tyrosine and tryptophan. 4. The preparation is suitable for use in the enzymic procedures for determining ATP, phosphocreatine and 3-phosphoglycerate.
The mechanism of biosynthesis and direction of chain extension of a polyl-(N-acetylglucosamine 1-phosphate) from the walls of Staphylococcus lactis N.C.T.C. 2102
Authors: Brooks D. , Baddiley J
Published: August 2015

1. The synthesis of a polymer of N-acetylglucosamine 1-phosphate, occurring in the walls of Staphylococcus lactis N.C.T.C. 2102, was examined by using cell-free enzyme preparations. The enzyme system was particulate, and probably represents fragmented cytoplasmic membrane. 2. Uridine diphosphate N-acetylglucosamine was the only substrate required for polymer synthesis and labelled substrate was used to show that N-acetylglucosamine 1-phosphate is transferred as an intact unit from substrate to polymer. 3. The properties of the enzyme system were studied. A high concentration of Mg2+ or Mn2+ was required for optimum activity, and the pH optimum was about 8·5. 4. End-group analysis during synthesis in vitro showed that newly formed chains contain up to about 15 repeating units. Pulse-labelling indicated that chain extension occurs by transfer from the nucleotide to the ‘sugar-end’ of the chain, i.e. to the end that is not attached to peptidoglycan in the wall.
The hydrolysis of monolayers of phosphatidyl-[Me−14C]choline by phospholipase D
Authors: Quarles R. H. , Dawson R. M. C.
Published: August 2015

1. The hydrolysis of monolayers of phosphatidyl[Me−14C]choline at the air/water interface by phospholipase D (phosphatidylcholine phosphatidohydrolase) was investigated by a surface-radioactivity technique by using a flow counter. 2. Phosphatidylcholine of high specific radioactivity was prepared biosynthetically in good yield from [Me−14C]choline by using Saccharomyces cerevisiae. 3. At initial monolayer pressures between 12 and 25 dynes/cm. the hydrolysis occurred in two stages, an initial slow hydrolysis followed by a rapid hydrolysis. Below 3dynes/cm. and above 28dynes/cm. no enzymic hydrolysis of pure phosphatidylcholine monolayers could be detected. 4. The rapid hydrolysis was proportional to the enzyme concentration in the subphase, its pH optimum was 6·6, and 0·2mm-Ca2+ was required for maximal activity. 5. Hydrolysis of the film was accompanied by a pronounced fall in the surface pressure even though the phosphatidic acid formed did not leave the film. When the pressure fell to low values the hydrolysis ceased even if the film was only partially hydrolysed. 6. Above monolayer pressures of 28dynes/cm. enzymic hydrolysis could be initiated by inclusion of phosphatidic acid (and less effectively stearyl hydrogen sulphate) in the film, although the rates were not appreciably higher than those observed at 25dynes/cm. with a pure phosphatidylcholine film. 7. The initiation of the hydrolysis by phosphatidic acid was facilitated by the inclusion of high Ca2+ concentrations and certain carboxylic acid buffer anions in the subphase, although these did not activate by themselves. 8. The initiation of the hydrolysis at high pressures could not be related to any change in the surface potential brought about by the addition of the long-chain anions to the film, nor could it be ascribed to a surface dilution effect. 9. The results are discussed in relation to previous studies on the hydrolysis of phosphatidylcholine particles by the enzyme and also similar investigations on phosphatidylcholine monolayers with other phospholipases.
Studies on the microsomal sodium-plus-potassium ion-stimulated adenosine triphosphatase system in rat ventral prostate
Authors: Ahmed K. , Williams-Ashman H G
Published: August 2015

A Mg2++Na++K+-stimulated adenosine triphosphatase (ATPase) preparation was isolated from rat ventral prostate by flotation of microsomal membranes in high-density sucrose solutions. The reaction medium for optimum Na++K+-stimulated ATPase activity was found to be: Na+, 115mm; K+, 7–10mm; Mg2+, 3mm; ATP, 3mm; tris buffer, pH7·4 at 38°, 20mm. The average ΔPi (Mg2++Na++K+ minus Mg2++Na+) was 9μmoles/mg. of protein/hr., representing a 30% increase over the Mg2++Na+-stimulated ATPase activity. At high concentrations, K+ was inhibitory to the enzyme activity. Half-maximal inhibition of Na++K+-stimulated ATPase activity was elicited by ouabain at 0·1mm. The preparation exhibited phosphatase activity towards ribonucleoside triphosphates other than ATP. However, stimulation of Pi release by Na++K+ was observed only with ATP as substrate. The apparent Km for ATP for Na++K+-stimulated activity was about 0·3×10−3m. Ca2+ inhibited only the Na++K+-stimulated ATPase activity. Mg2+ could be replaced by Ca2+ but then no Na++K+ stimulation of ATPase activity was noticed. The addition of testosterone or dihydrotestosterone (17β-hydroxy-5α-androstan-3-one) in vitro at 0·1–10μm under a variety of experimental conditions did not significantly increase the Na++K+-stimulated ATPase activity. The enzyme preparations from prostates of orchidectomized rats, however, exhibited a drastic decrease in the specific activity of Na++K+-stimulated ATPase; these changes were prevented in the orchidectomized rats by injection of testosterone propionate.
The inhibition by fluorocitrate of rat liver mitochondrial and extramitochondrial aconitate hydratase
Authors: Guarriera-Bobyleva Valentina , Buffa P.
Published: August 2015

1. The effects of synthetic fluorocitrate were studied on: (a) the oxidation of citrate and cis-aconitate by rat liver mitochondria; (b) the activity of the aconitate hydratase found in the liver cell sap; (c) the activity of the aconitate hydratase solubilized from liver mitochondria. 2. Fluorocitrate was found to be a potent inhibitor of oxidation of citrate but only a weak inhibitor of oxidation of cis-aconitate: 6·7μm-fluorocitrate (containing 4% of the inhibitory isomer) caused 94% inhibition of the oxidation of citrate (2mm) whereas 1·0mm-fluorocitrate was necessary to provoke the same inhibition when cis-aconitate (2mm) was the substrate. The degree of inhibition varied in relation to the respiratory state of mitochondria when fluorocitrate was added. The inhibition could be partially reversed by cis-aconitate. 3. The aconitate hydratase extracted from the mitochondria was much less inhibited by fluorocitrate than was the mitochondria-bound enzyme, and the aconitate hydratase found in the cell sap was even less sensitive. 0·3mm-Fluorocitrate was required to cause 50% inhibition of the reaction citrate→cis-aconitate, catalysed by the aconitate hydratase extracted from the mitochondria, and 1·2m-fluorocitrate for the extramitochondrial enzyme. For both enzymes the reaction citrate→cis-aconitate was 2–3 times more sensitive to fluorocitrate than was the reaction isocitrate→cis-aconitate. The inhibition was of the competitive type for both reactions.
Ketone-body utilization by adult and suckling rat brain in vivo
Authors: Hawkins R. A. , Williamson D. H. , Krebs H. A.
Published: August 2015

1. Ketone-body utilization in fed and starved adult and suckling rats has been investigated by measuring arterio-venous differences across the brain. Venous blood was collected from the confluence of sinuses and arterial blood from the femoral artery in adult rats and by cardiac puncture in suckling rats. 2. During starvation the arterio-venous difference of ketone bodies increased in proportion to their concentrations in the blood and reached a value of 0.16mm at 48h. At a given concentration of the respective ketone bodies the arterio-venous differences of acetoacetate were about twice those of 3-hydroxybutyrate. 3. Fed rats in which the concentrations of ketone bodies were raised by intravenous infusion of sodium acetoacetate had the same arterio-venous differences as starved rats at corresponding ketone-body concentrations. Thus the ability of the rat brain to utilize ketone bodies is independent of the nutritional state. 4. The concentrations of glucose, acetoacetate and 3-hydroxybutyrate were much lower in the brain than in the arterial blood. The measured (blood concentration)/(brain concentration) ratio was 4.4 for glucose, 4.5 for acetoacetate and 8.1 for 3-hydroxybutyrate in 48h-starved rats. 5. The mean arterio-venous difference of glucose across the brain was 0.51mm in fed rats and 0.43mm in 96h-starved rats. 6. Conversion of glucose into lactate rose from negligible values in the fed state to 0.2mm after 48h starvation and decreased to zero after 96h starvation. 7. In 16–22-day-old suckling rats the arterio-venous differences of ketone bodies across the brain were also proportional to the ketone-body concentration, but they were about 3–4 times greater than in adult rats at the same blood ketone-body concentration. 8. Arterio-venous differences of glucose were about the same in adult and suckling rats. 9. The brain of fed suckling rats formed more lactate from glucose than fed adult rats. 10. The results indicate that ketone bodies are major metabolic fuels of the brain of the suckling rat under normal conditions.
The biosynthesis of galactofuranosyl residues in galactocarolose
Authors: Trejo A. Garcia , Haddock J. W. , Chittenden G. J. F. , Baddiley J.
Published: August 2015

1. Cell-free extracts of Penicillium charlesii G. Smith were used in a study of the biosynthesis of the galactofuranose polymer, galactocarolose. 2. UDP-glucose and UDP-galactopyranose were precursors of galactocarolose and it was shown that the galactofuranose residues in the polymer were formed from glucose without fission of the hexose carbon chain. 3. A new nucleotide, UDP-α-d-galactofuranose, was formed by the system and was a major product when polymer synthesis was inhibited by F- or Zn2+; the nucleotide was isolated and its structure determined. 4. UDP-α-d-galactofuranose was efficiently utilized for polymer synthesis and shown to be formed from the pyranose nucleotides. 5. A route for the biosynthesis of galactocarolose, involving a novel ring contraction of the hexose residue while still attached to the nucleotide, is proposed.
Reactions of d-glyceraldehyde 3-phosphate dehydrogenase facilitated by oxidized nicotinamide–adenine dinucleotide
Authors: Trentham D. R.
Published: August 2015

Transient kinetic methods have been used to study the influence of NAD+ on the rate of elementary processes of the reversible oxidative phosphorylation of d-glyceraldehyde 3-phosphate catalysed by d-glyceraldehyde 3-phosphate dehydrogenase. In the pH range 5–8 NAD+ is bound to the enzyme during the following elementary processes of the mechanism: phosphorolysis of the acyl-enzyme, its formation from 1,3-diphosphoglycerate and the enzyme and the formation and breakdown of the glyceraldehyde 3-phosphate–enzyme complex. The rates of these four elementary processes only equal or exceed the turnover rate of the enzyme when NAD+ is bound and are as much as 104 times the rates in the absence of NAD+. Autocatalysis of the reductive dephosphorylation of 1,3-diphosphoglycerate occurs when glyceraldehyde 3-phosphate release is rate determining because NAD+ is a reaction product. An important feature of the enzyme mechanism is that the negative-free-energy change of a chemical reaction, acyl-enzyme formation, is linked in a simple way to the positive-free-energy change of a dissociation reaction, NAD+ release.
Chemical composition of an oestrogen-induced calcium-binding glycolipophosphoprotein in Xenopus laevis
Authors: Ansari A. Q. , Dolphin P. J. , Lazier C. B. , Munday K. A. , Akhtar M.
Published: August 2015

1. Oestrogen treatment has previously been shown to induce the formation of large amounts of a serum protein, vitellogenin (xenoprotein), in Xenopus laevis. Vitellogenin was purified from serum by dimethylformamide precipitation and was shown to be homogeneous by a variety of electrophoretic techniques. 2. The molecular weight of vitellogenin was estimated by gel filtration to be about 6×105. The chemical constituents of vitellogenin were determined and lead to the characterization of this protein as a serum calcium-binding glycolipophosphoprotein. 3. The extractable lipid accounted for 12% of vitellogenin. Gas–liquid-chromatographic analysis of the saponified lipid moiety showed the presence of palmitic acid, palmitoleic acid, stearic acid, oleic acid and linoleic acid in the molecular proportions 6.8: 1.5: 1.0: 3.6: 1.4. 4. The carbohydrate moiety consisted of 0.4g of hexose, 0.77g of hexosamine and 0.18g of sialic acid/100g of vitellogenin. 5. The calcium and phosphorus contents were 0.85 and 1.65g/100g of vitellogenin respectively. 6. Serum from oestrogen-treated animals injected with 45CaCl2 contained 9.7 times the radioactivity present in serum from untreated 45CaCl2-injected animals. Of the radioactivity due to 45CaCl2 in the serum of oestrogen-treated animals 72% was non-diffusible on dialysis. Of this activity 65.4% was associated with the vitellogenin band on cellulose acetate electrophoresis.
Ribosomal ribonucleic acid synthesis in Bacillus subtilis
Authors: Avery R. J. , Midgley J. E. M.
Published: August 2015

The mode of biosynthesis of the 16S and 23S ribosomal ribonucleic acids (rRNA) was studied in Bacillus subtilis 168thy-. Three criteria were used to define the characteristics of the rRNA species: (i) the time required at 37°C to synthesize 16S and 23S rRNA chains de novo in growing cultures; (ii) the degree of reactivity of the 3′-terminal groups of the rRNA molecules with periodate and [carbonyl-14C]isonicotinic acid hydrazide; and (iii) the reactivity of the 5′-terminal regions of the rRNA molecules with the bacterial exonuclease purified by Riley (1969). The 16S and 23S chains of B. subtilis were synthesized at rates of 22±2 and 21±2 nucleotides added/s. The periodate–[14C]isonicotinic acid hydrazide and the exonuclease techniques for estimating apparent chain lengths of RNA indicated that the chain length of the 23S rRNA was 1.8 times that of the 16S fraction. The apparent chain lengths of each rRNA species were: 16S rRNA, 1650±50 nucleotide residues; 23S rRNA, 3050±90 nucleotide residues. It appears that, the 16S and 23S rRNA molecules in B. subtilis are synthesized in the expected manner, by simple polymerization of the final products on independent cistrons.
The diglyceride kinase of rat cerebral cortex
Authors: Lapetina E. G. , Hawthorne J. N.
Published: August 2015

1. Formation of phosphatidic acid by diglyceride kinase (EC 2.7.1.-) in the presence of ATP and Mg2+ was shown in a homogenate and subcellular fractions of rat cerebral cortex. 2. The kinase was activated by Mg2+. Ca2+ activated to a smaller extent but was inhibitory in the presence of optimum concentration of Mg2+. Activity was greatly increased in the presence of added 1,2-diglyceride. 3. Sodium deoxycholate markedly stimulated the reaction, but other detergents (Cutscum and Triton X-100) did not. 4. Diglyceride kinase was concentrated in the supernatant and microsomal fractions from rat cerebral cortex. The distribution of the kinase in the particulate fractions resembled that of acetylcholinesterase and 5′-nucleotidase. 5. The rate of phosphatidic acid synthesis by the diglyceride kinase route was much greater than reported rates for acylation of 3-glycerophosphate and was also very rapid in comparison with the rates of other steps in the synthesis of phosphoinositides. 6. Acetylcholine had no stimulatory effect on diglyceride kinase of isolated intact nerve-ending particles or of nerve-ending membranes obtained after osmotic shock.
The rate-limiting step of protein synthesis in vivo and in vitro and the distribution of growing peptides between the puromycinlabile and puromycin-non-labile sites on polyribosomes
Authors: Earl D. C. N. , Hindley Susan T.
Published: August 2015

1. At 3 min after an intravenous injection of radioactive amino acids into the rat, the bulk of radioactivity associated with liver polyribosomes can be interpreted as growing peptides. 2. In an attempt to identify the rate-limiting step of protein synthesis in vivo and in vitro, use was made of the action of puromycin at 0°C, in releasing growing peptides only from the donor site, to study the distribution of growing peptides between the donor and acceptor sites. 3. Evidence is presented that all growing peptides in a population of liver polyribosomes labelled in vivo are similarly distributed between the donor and acceptor sites, and that the proportion released by puromycin is not an artifact of methodology. 4. The proportion released by puromycin is about 50% for both liver and muscle polyribosomes labelled in vivo, suggesting that neither the availability nor binding of aminoacyl-tRNA nor peptide bond synthesis nor translocation can limit the rate of protein synthesis in vivo. Attempts to alter this by starvation, hypophysectomy, growth hormone, alloxan, insulin and partial hepatectomy were unsuccessful. 5. Growing peptides on liver polyribosomes labelled in a cell-free system in vitro or by incubating hemidiaphragms in vitro were largely in the donor site, suggesting that either the availability or binding of aminoacyl-tRNA, or peptide bond synthesis, must be rate limiting in vitro and that the rate-limiting step differs from that in vivo. 6. Neither in vivo nor in the hemidiaphragm system in vitro was a correlation found between the proportion of growing peptides in the donor site and changes in the rate of incorporation of radioactivity into protein. This could indicate that the intracellular concentration of amino acids or aminoacyl-tRNA limits the rate of protein synthesis and that the increased incorporation results from a rise to a higher but still suboptimum concentration.
Effect of nitrogenase compounds from mutant and wild-type strains of Azotobacter on the dilution effect of nitrogenase
Authors: Sorger G. J.
Published: August 2015

Nitrogenase activity exhibits a dilution effect. Evidence is presented that the reason for the dilution effect is that one of the component proteins of nitrogenase is limiting in preparations of this enzyme. The limiting component appears to be the non-haem-iron-containing protein (also called fraction II, iron protein, azoferredoxin), which is equivalent to the enhancement factor for nitrogenase activity present in extracts of nitrogenaseless mutant 22R1. A mathematical function of specific activity is described that is useful in describing nitrogenase. It takes into account the dilution effect and the exponential nature of the relationship between nitrogenase activity and enzyme protein concentration.
Protein aggregation in C-phycocyanin. Studies at very low concentrations with the photoelectric scanner of the ultracentrifuge
Authors: MacColl R. , Lee J. J. , Berns D. S.
Published: August 2015

Solutions of C-phycocyanin of very low concentrations were examined by sedimentation-velocity studies in the Spinco model E ultracentrifuge equipped with a photoelectric scanning system and a monochromator. At sufficiently low concentrations complete disaggregation from the hexamer to the monomer was observed. The equilibrium constant of monomer to hexamer was estimated to be approx. 1030. For studies of aggregation over the complete range of concentration, C-phycocyanins from Phormidium luridum and Lyngbya sp. were used. Sedimentation-velocity studies at high concentration with schlieren optics are reported for C-phycocyanins from Anabaena variabilis and Lyngbya sp. The pH-dependence of aggregation and the temperature-dependence of trimer–hexamer equilibrium for phycocyanins from these algae were found to be similar to those of other C-phycocyanins. The principal feature of the pH-dependence is the dominance of hexamers at the isoelectric point. Increasing temperature increased the amount of hexamer and decreased the amount of trimer.
The metabolism of retinyl methyl ether in the rat in vivo
Authors: Narindrasorasak S. , Pimpa P. , Lakshmanan M. R.
Published: August 2015

1. Retinyl methyl ether was converted into vitamin A in vitamin A-deficient rats regardless of whether administered by oral, intraperitoneal, intramuscular or subcutaneous route; intramuscular administration seemed to be the best for conversion as well as storage. 2. Significantly, unchanged retinyl methyl ether was also found in the liver after oral administration but not after administration by other routes. 3. Oral administration of 1mg of retinyl methyl ether led to a progressive increase in liver vitamin A with time reaching a value of 16% of administered dose after 24h. No retinyl methyl ether was detectable in liver at any time-interval in this experiment. 4. Conversely, oral administration of 4mg of retinyl methyl ether/day for 4 days led to the accumulation of 25% of the dose as unchanged retinyl methyl ether in the liver 1 day after the last dose; however, it was gradually but completely converted into vitamin A over a period of 18 days. 5. The significance of these findings with special reference to the fundamental metabolism of vitamin A, the site of conversion of retinyl methyl ether into vitamin A, the relative efficiency of various routes of administration and its biological activity are discussed.
The effect of pH on cathepsin activities in mouse liver heterolysosomes
Authors: Mego John L.
Published: August 2015

1. Mouse liver heterolysosomes containing 125I-labelled albumin were incubated at 35°C in 0.25m-sucrose–0.05m-mercaptoethanol, and various concentrations of buffers at pH4, 5, 7 or 8, and the degradation of intraparticulate protein was measured. 2. Buffers at pH4, 7 or 8 inhibited proteolytic activity and the inhibitions were greater with increasing concentrations of buffers. Tris–acetate buffer, pH8, was a more effective inhibitor. Tris–acetate buffer, pH5, neither inhibited nor stimulated proteolytic activity at concentrations up to 0.1m. 3. Inhibition by pH8 buffers could be accounted for by increased breakage rates of heterolysosomes. 4. Preincubation of heterolysosomes in 0.2m-sodium bicarbonate inhibited proteolytic activity when the particles were washed free of bicarbonate, but the inhibition was reversed if these particles were incubated in media containing pH5 buffer. The inhibition was shown not to be due to an increased breakage of heterolysosomes. 5. It was concluded that the pH within heterolysosomes is about 5, and it is possible to alter reversibly the pH within these particles. The possibility for the existence of an intralysosomal buffering system which maintains the pH at about 5 in heterolysosomes is discussed. This buffering system may be related to the presence of large quantities of acidic lipoproteins in lysosomes observed by other investigators.
The disulphide bonds of erabutoxin a, a neurotoxic protein of a sea-snake (Laticauda semifasciata) venom
Authors: Endo Y. , Sato S. , Ishii S. , Tamiya N.
Published: August 2015

Erabutoxin a was partially hydrolysed with enzymes and sulphuric acid and the resulting peptides were separated from each other by column chromatography and paper electrophoresis. From the results of amino acid analyses of the sulphur-containing peptides and their oxidized components, all four disulphide bridges in the toxin molecule were located. The disulphide bonds were found between half-cystine residues at positions 3 and 24, 17 and 41, 43 and 54, and 55 and 60 from the N-terminus.
Development of adenylate kinase isoenzymes in rat liver
Authors: Filler R. , Criss W. E.
Published: August 2015

Total adenylate kinase activity was determined in developing rat liver. The activity was 18 units/g wet weight of tissue in foetal liver; this increased to 41 units/g immediately after birth and continued increasing until adult activities of 150 units/g were reached after two weeks. The adenylate kinase activity was separated into four isoenzymes. Only isoenzymes II and III were observed in foetal rat liver. Isoenzyme II activity was 2 units/g in the foetal liver and increased to 25 units/g in adult liver. Adenylate kinase III activity was 20 units/g in the foetal liver and increased to 118 units/g in adult liver. The possible role that adenylate kinase might have in regulating the energy flow in the developing liver cell is discussed.
The distribution of some hydrolases in glomeruli and tubular fragments prepared from rat kidney by zonal centrifugation
Authors: Taylor D. G. , Price R. G. , Robinson D.
Published: August 2015

1. A collagenase digest of rat kidney cortex was separated into four bands by zonal centrifugation. 2. Two of these bands were shown by light-microscopy to contain glomeruli and tubular fragments, which were free from each other and well separated from other renal material. 3. Protein, N-acetyl-β-glucosaminidase, 5′-nucleotidase, l-leucine β-naphthylamidase, leucine aminopeptidase, acid phosphatase and alkaline phosphatase were assayed across the gradient. 4. The greater proportion of these enzyme activities was recovered in the tubular fragments and acid phosphatase was the only enzyme detected in significant amounts in the glomeruli. 5. Tubular fragments and glomeruli were sedimented and multiple forms of β-naphthylamidase, N-acetyl-β-glucosaminidase, acid phosphatase and alkaline phosphatase were investigated by starch-gel electrophoresis.
Synthesis of pyruvate carboxylase from its apoenzyme and (+)-biotin in Bacillus stearothermophilus. Mechanism and control of the reaction
Authors: Sundaram T. K. , Cazzulo J. J. , Kornberg H. L.
Published: August 2015

1. Acetyl-CoA acts as a positive allosteric effector in the formation of active pyruvate carboxylase from its apoenzyme, ATP and (+)-biotin which is catalysed by holoenzyme synthetase; this effect is counteracted by l-aspartate. 2. The Hill coefficients (apparent n values) were approximately 2 for acetyl-CoA and 4 for l-aspartate; the n value for each effector remained constant when the concentration of the other effector was varied. 3. Active pyruvate carboxylase was formed also when the apoenzyme was incubated with holoenzyme synthetase and synthetic biotinyl-5′-AMP; acetyl-CoA and l-aspartate affected this process as they did the overall reaction from (+)-biotin and ATP. 4. When hydroxylamine replaced the apoenzyme, holoenzyme synthetase catalysed the formation of biotinylhydroxamate from (+)-biotin and ATP. This reaction was not affected by the allosteric effectors. 5. The apoenzyme was protected against thermal denaturation by acetyl-CoA and, to a lesser degree, by l-aspartate. The holoenzyme synthetase was not markedly protected by these effectors. 6. It is concluded that the allosteric effectors act on the apoenzyme and not the synthetase.
A survey of the interaction of calcium ions with mitochondria from different tissues and species
Authors: Carafoli Ernesto , Lehninger Albert L.
Published: August 2015

A survey was made of the capacity of mitochondria isolated from a number of different tissues and species to accumulate Ca2+ from the suspending medium during electron transport. The species examined included the rat, mouse, rabbit, hamster, guinea pig, cow, chicken, turtle, blowfly, yeast and Neurospora crassa. The tissues examined included vertebrate liver, kidney, brain, heart, spleen, thyroid and adrenal cortex, and the flight muscle of the blowfly. The mitochondria from all vertebrate tissues examined showed: (a) stimulation of State 4 respiration by added Ca2+ (Ca2+/~ activation ratio about 2.0), accompanied by accumulation of Ca2+ and ejection of H+, with a H+/Ca2+ ratio about 1.0; (b) a requirement of phosphate for accumulation of large amounts of Ca2+; (c) respiration-independent high-affinity binding sites for Ca2+; (d) endogenous Ca2+, which is largely released by uncoupling agents. However, mitochondria from yeast and blowfly flight muscle are unable to accumulate Ca2+ in a respiration-dependent process and possess no high-affinity Ca2+-binding sites. These findings support the view that the high-affinity sites represent the ligand-binding sites of a specific Ca2+ ‘permease’ or transport system in the membrane. The relatively high affinity for Ca2+, which equals or exceeds the affinity for ADP, and the generally uniform characteristics of Ca2+ transport in all the vertebrate mitochondria tested strongly suggest that respiration-linked Ca2+ accumulation plays a general and fundamental role in vertebrate cell physiology.
Stoicheiometrical proton and potassium ion movements accompanying the absorption of amino acids by the yeast Saccharomyces carlsbergensis
Authors: Eddy A. A. , Nowacki J. A.
Published: August 2015

1. Proton uptake into the yeast Saccharomyces carlsbergensis, was studied at pH4.5–5.5 in the presence of both antimycin and 2-deoxyglucose to inhibit energy metabolism. Previous work had shown that the cells then absorbed about 20nmol of glycine or l-phenylalanine against a considerable amino acid concentration gradient. The addition of the amino acid immediately stimulated the rate of uptake of protons two- to three-fold. About 2 extra equivalents of H+ accompanied a given amount of the amino acids into the yeast preparations exposed to the metabolic inhibitors for 2–4min and about 1.2 equivalents after 20min exposure. 2. Analogous observations were made during serial additions of glycine, l-phenylalanine, l-leucine and l-lysine to preparations lacking the metabolic inhibitors and deficient in substrates needed for energy metabolism. In fresh cellular preparations the influx of glycine was then closely coupled to a stimulated flow of 2.1 equiv. of H+ into the yeast. A similar number of K+ ions left the cells. About 30% of the extra protons was subsequently ejected from the yeast. Deoxyglucose and antimycin together inhibited the ejection of protons. When the yeast had been fed with glucose energy metabolism was stimulated and almost as many protons as were absorbed with the amino acid were apparently ejected again. 3. Yeast preparations containing Na+, instead of K+, as the principal cation absorbed about 1 extra equivalent of H+ after the addition of phenylalanine, glycine or leucine. This response was not observed in the presence of both deoxyglucose and antimycin. 4. The observations show that H+ and, in certain circumstances, K+ are co-substrates in the transport of the amino acids into the yeast. An analogy is drawn with the roles of Na+ and K+ as co-substrates in certain mammalian systems. The results lead to various models relating the physical flow of the co-substrate ions on the amino acid carrier to the transduction of chemical energy in an associated ion pump forming part of the mechanism for transporting amino acids into the yeast.
Histone kinase and cell division
Authors: Pawse A. R. , Ord Margery G. , Stocken L. A.
Published: August 2015

1. The activity of the soluble phosphokinase for histone F1 increases in regenerating rat liver during the first period of DNA synthesis after partial hepatectomy. The increase probably represents new enzyme synthesis. 2. A dose of 500rd of γ-irradiation given early in G1 decreases the amount of histone F1 phosphokinase found 22h after partial hepatectomy by 60–70%. 3. The enzyme preparations also contained a histone F1 phosphatase; the presence together of the kinase and phosphatase caused a disproportion between net 31P uptake and 32P incorporation into histone F1. 4. All four subclasses of histone F1 could accept phosphate from ATP. 5. Crude enzyme preparations transferred more 31P into histone F1 with an initially low phosphate content than into one with a high phosphate content; conversely, more 32P was transferred into the latter.
Studies on the origin of choline in the brain of the rat
Authors: Ansell G. B. , Spanner Sheila
Published: August 2015

1. Labelled precursors of choline, namely ethanolamine, dimethylaminoethanol and methionine and also labelled choline itself were injected intraperitoneally into the adult female rat and the incorporation into lipids and water-soluble fractions was traced in liver, blood and brain. 2. No significant free choline was detected and no labelling of the phosphorylcholine of blood. There was, however, considerable labelling of the phosphorylcholine of brain and liver. 3. After intracerebral injection, [1,2-14C]dimethylaminoethanol was rapidly phosphorylated and converted into phosphatidyldimethylaminoethanol, presumably by the cytidine pathway. 4. In view of the pattern of labelling and the amount of phosphatidylcholine in the tissues examined, it seems highly likely that choline is transported to the brain by the blood in a lipid-bound form.
Cathepsin D. Characteristics of immunoinhibition and the confirmation of a role in cartilage breakdown
Authors: Dingle J. T. , Barrett A. J. , Weston P. D.
Published: August 2015

1. Antisera were raised against lysosomal cathepsin D of man, chicken and rabbit. 2. The antisera were found to be specific and potent inhibitors of cathepsin D activity. 3. The immunological nature of the inhibition was established. 4. The inhibitory effect was studied by varying pH, antiserum/enzyme ratio, time of incubation, concentration of components and order of mixing, and by using purified antibody and univalent antibody fragments. 5. The specificities of the antisera were examined with respect to other enzymes, isoenzymes of cathepsin D and cathepsin D from different organs. 6. The antisera prevented the action of cathepsin D on isolated proteoglycans and on cartilage. 7. The antisera produced up to 90% inhibition of the autolysis of cartilage from chicks and rabbits, indicating that cathepsin D is the enzyme mainly responsible for the breakdown of proteoglycans in this system.
The catabolism of phosphatidylethanolamine by the rumen protozoon Entodinium caudatum and its conversion into the N-(1-carboxyethyl) derivative
Authors: Coleman G. S. , Kemp P. , Dawson R. M. C.
Published: August 2015

1. The N-(2-hydroxyethyl)alanine esterified to phosphatidic acid in anaerobic ciliate rumen protozoa has the l configuration. 2. Labelling experiments with Entodinium caudatum cultures using [32P]Pi [2-14C]ethanolamine and 32P- and 14C-labelled phosphatidylethanolamine show that phosphatidylethanolamine is the direct lipid precursor of the N-(2-hydroxyethyl)alanine-containing phospholipid. 3. Labelling experiments with [14C]starch, [14C]lactate and [14C]pyruvate with E. caudatum cultures indicate that a three-carbon glycolytic intermediate is probably the precursor of the N-(1-carboxyethyl) grouping which substitutes on the amino group of phosphatidylethanolamine. 4. [32P]phosphatidylethanolamine is catabolized by E. caudatum forming initially glycerylphosphorylethanolamine and subsequently glycerophosphate and Pi. A little phosphorylethanolamine formed may possibly arise from bacterial enzymes ingested by the protozoa.
The number of catalytic sites in acetylcholinesterase
Authors: Leuzinger W.
Published: August 2015

By using two methods of titration, the number of active sites in acetylcholinesterase was determined. Either stepwise inhibition of the enzyme by an irreversible inhibitor, namely di-isopropyl phosphorofluoridate, or direct measurement of the concentration of active sites by titration with o-nitrophenyl dimethylcarbamate yielded an equivalent weight of approx. 130000 for an active site in acetylcholinesterase. This indicates two sites per molecule, since the native enzyme has a molecular weight of 260000.