bluuebunny/crossref_metadata_2025 huggingface dataset:
Ribonucleic acid synthesis by spermatozoa from the rat and hamster
Authors: MacLaughlin Julia , Terner Charles
Published: August 2015

Spermatozoa from the cauda of the epididymis of the hamster and rat were incubated with [5-3H]uridine and glucose. By using a procedure avoiding bacterial and other cellular contamination, sonic extracts were prepared and digested with deoxyribonuclease and Pronase. Radioactive RNA of high molecular weight was isolated by two methods: (a) gel filtration on Sephadex G-75 columns and (b) polyacrylamide-gel electrophoresis in which it migrated in the region of 28S and 23S RNA markers. The macromolecules were alkali-labile and hydrolysed by ribonuclease. From 3H radioactivity and E260 of the isolated RNA the rate of incorporation of uridine into RNA of spermatozoa was calculated to be 0.1–0.5nmol/h per mg of RNA.
Fluorocitrate inhibition of aconitate hydratase and the tricarboxylate carrier of rat liver mitochondria
Authors: Brand M. D. , Evans Susan M. , Mendes-Mourão J. , Chappell J. B.
Published: August 2015

1. The effect of biologically synthesized and purified fluorocitrate on the metabolism of tricarboxylate anions by isolated rat liver mitochondria was investigated, in relation to the claim by Eanes et al. (1972) that this fluoro compound inhibits the tricarboxylate carrier at concentrations at which it has little effect on the aconitate hydratase activity. 2. That the inhibitory action of fluorocitrate is at the level of the aconitate hydratase and not at the level of the tricarboxylate carrier is indicated by the following findings. Although the oxidation of citrate and cis-aconitate, but not that of isocitrate, was inhibited by fluorocitrate, the exchange of internal citrate for external citrate or l-malate was not. Had the tricarboxylate carrier been affected, these latter exchange reactions would have been inhibited. 3. By using aconitate hydratase solubilized from mitochondria it was found that with citrate as substrate the inhibition by fluorocitrate was partially competitive (Ki=3.4×10−8m), whereas with cis-aconitate as substrate the inhibition was partially non-competitive (Ki=3.0×10−8m).
The biosynthesis of glucagon in perfused rat pancreas
Authors: O'Connor Kevin J. , Gay Adrian , Lazarus Norman R.
Published: August 2015

The biosynthesis of glucagon was studied by using the recirculated, isolated perfused rat pancreas. [3H]Tryptophan was initially incorporated into acid–ethanol-extractable protein, which on gel filtration was eluted with a molecular weight of about 9000 and contained a small amount of glucagon immunoreactivity. With longer incubation [3H]tryptophan incorporation into a second peak was obtained in an identical position with that of the majority of rat glucagon immunoreactivity. This peak of labelled protein exhibited migration characteristics on polyacrylamide-gel electrophoresis identical with those of rat glucagon and was identified as newly synthesized glucagon by demonstration of specific binding and dissociation behaviour with glucagon antibodies. The incorporation of [3H]tryptophan into acid–ethanol-extractable protein was inhibited by cycloheximide. High concentrations of glucose increased [3H]tryptophan incorporation into high-molecular-weight protein but decreased incorporation into proteins smaller than cytochrome c. The pattern of [3H]leucine incorporation into protein was similar to that of [3H]tryptophan.
Concentration of adenosine 3′:5′-cyclic monophosphate in mouse pancreatic islets measured by a protein-binding radioassay
Authors: Cooper R. H. , Ashcroft S. J. H. , Randle P. J.
Published: August 2015

A protein-binding radioassay for cyclic AMP was modified to detect less than 0.025pmol of the nucleotide. The method was applied to the measurement of cyclic AMP in small numbers of mouse pancreatic islets (as little as 25μg of tissue) by use of barium acetate–H2SO4 for deproteinization. The concentration of cyclic AMP in mouse islets incubated in media containing 3.3 or 20mm-glucose was 0.016pmol/10 islets (approx. 1μm in intracellular water). Glucose concentration (3.3 or 20mm) had no detectable effect on islet concentrations of cyclic AMP with periods of incubation or perifusion ranging from 0.5 to 60min, although insulin release rate was rapidly increased by 20mm-glucose. Caffeine (5mm) or 3-isobutyl-1-methylxanthine (1mm), which are known inhibitors of islet cyclic AMP phosphodiesterase, produced marked and rapid increases in islet cyclic AMP concentration at 3.3 or 20mm-glucose, but only enhanced the insulin release rate at the higher glucose concentration. The role of cyclic AMP in insulin release induced by glucose is discussed.
Determination and significance of l-tyrosine O-sulphate and its deaminated metabolites in normal human and mouse urine
Authors: Hext Paul M. , Thomas Susan , Rose Frederick A. , Dodgson Kenneth S.
Published: August 2015

Methods were developed for the determination of the O-sulphate esters of l-tyrosine, p-hydroxyphenylpyruvate, p-hydroxyphenylacetate and p-hydroxybenzaldehyde in human urine. The amounts of these esters normally present in human female urine were determined. The quantities and specific radioactivities of l-tyrosine O-sulphate and p-hydroxyphenylpyruvate O-sulphate in mouse urine after labelled tyrosine had been fed were determined and were consistent with the hypothesis that the sole source of p-hydroxyphenylpyruvate O-sulphate was l-tyrosine O-sulphate. It is suggested that the turnover of known polypeptides containing l-tyrosine O-sulphate residues can account for only a portion of the quantities of the ester and its metabolite(s) that are present in normal female human urine.
Regulation of nitrate reductase of Neurospora at the level of transcription and translation
Authors: Sorger G. J. , Davies J.
Published: August 2015

The presence of nitrate is required for the induced synthesis of NADPH–nitrate reductase and its related partial activity Benzyl Viologen–nitrate reductase in a wild-type strain of Neurospora. In nit-3, a mutant lacking complete NADPH–nitrate reductase activity but retaining the partial activity Benzyl Viologen–nitrate reductase, the presence of nitrate ions is not required for the de-repressed synthesis of the latter enzyme. The accumulation of the capacity to synthesize nitrate reductase, and the related Benzyl Viologen–nitrate reductase, in the absence of protein synthesis does not require nitrate in the normal strain or in strain nit-3. Ammonia antagonizes the accumulation of this capacity in both strains. Nitrate is required for the synthesis of nitrate reductase and related activities from presumedly preformed mRNA in the wild-type strain. Nitrate is not required for the comparable function in strain nit-3. Ammonia appears to stop the synthesis of nitrate reductase and related activities from presumedly preformed mRNA in the wild-type strain and in strain nit-3. The effects of nitrate, or ammonia and of no nitrogen source on the induced synthesis of nitrate reductase cannot be explained on the basis of the effects of the different nitrogen sources on general synthesis of RNA or of protein.
The electron-transport system of mitochondria from the slime mould Physarum polycephalum
Authors: Barnes Roger , Colleran Emer M. , Jones O. T. G.
Published: August 2015

A method is described for the preparation of mitochondria from the slime mould Physarum polycephalum; the mitochondria were not coupled. P. polycephalum mitochondria oxidized added NADH via a rotenone-insensitive pathway, but the oxidation of malate plus glutamate was rotenone sensitive; both of these substrates reduced much less cytochrome b than did succinate, in both aerobic and anaerobic steady states. Spectroscopy at 77°K separated three absorption maxima in the α-band region, at 560nm, 553nm and one at 547nm due to cytochrome c. The absorption at 553nm was increased in the aerobic steady state by the addition of 2-heptyl-4-hydroxyquinoline N-oxide, suggesting that it was due to a b-type cytochrome. All three absorption maxima appeared in the aerobic steady state after the addition of a range of substrates. The respiratory activity with different substrates and the response to inhibitors of respiration were similar to those previously described for fungus mitochondria (Weiss et al., 1970; Erickson & Ashworth, 1969). When grown under conditions of haem limitation the mitochondria contained a lower concentration of cytochromes than normal.
Polyprenyl pyrophosphate–p-hydroxybenzoate polyprenyltransferase activity in mitochondria of broad-bean seeds and yeast (Short Communication)
Authors: Thomas Graham , Threlfall David R.
Published: August 2015

Cell-free homogenates prepared from broad-bean seeds and yeast cells are capable of synthesizing 4-carboxy-2-polyprenylphenols from p-hydroxybenzoate and either isopentenyl pyrophosphate or protein-bound polyprenyl pyrophosphates (produced by incubating a Micrococcus lysodeikticus extract with isopentenyl pyrophosphate). The mitochondria contained all the polyprenyl pyrophosphate–p-hydroxybenzoate polyprenyltransferase activity; however, unlike the homogenates they could not synthesize a side chain from isopentenyl pyrophosphate and had to be provided with protein-bound polyprenyl pyrophosphates.
A model for the regulation of δ-aminolaevulinate synthetase induction in rat liver
Authors: Padmanaban Govindarajan , Rao Manchanahalli R. Satyanarayana , Malathi Krishnamachari
Published: August 2015

A reciprocal relationship exists between the cytochrome P-450 content and δ-aminolaevulinate synthetase activity in adult rats. In young rats the basal δ-aminolaevulinate synthetase activity is higher and the cytochrome P-450 content is lower compared with the adult rat liver. Administration of allylisopropylacetamide neither induces the enzyme nor causes degradation of cytochrome P-450 in the young rat liver, unlike adult rat liver. Allylisopropylacetamide fails to induce δ-aminolaevulinate synthetase in adrenalectomized–ovariectomized animals or intact animals pretreated with successive doses of the drug, in the absence of cortisol. The cortisol-mediated induction of the enzyme is sensitive to actinomycin D. Allylisopropylacetamide administration degrades microsomal haem but not nuclear haem. Haem does not counteract the decrease in cytochrome P-450 content caused by allylisopropylacetamide administration, but there is evidence for the formation of drug-resistant protein-bound haem in liver microsomal material under these conditions. Phenobarbital induces δ-aminolaevulinate synthetase under conditions when there is no breakdown of cytochrome P-450. On the basis of these results and those already published, a model is proposed for the regulation of δ-aminolaevulinate synthetase induction in rat liver.
The structure of a β-(1→6)-d-glucan from yeast cell walls
Authors: Manners David J. , Masson Alan J. , Patterson James C. , Björndal Håkan , Lindberg Bengt
Published: August 2015

By selective enzymolysis, or chemical fractionation, a minor polysaccharide component has been isolated from yeast (Saccharomyces cerevisiae) glucan. This minor component has a degree of polymerization of about 130–140, a highly branched structure, and a high proportion of β-(1→6)-glucosidic linkages. The molecules also contain a smaller proportion of β-(1→3)-glucosidic linkages that serve mainly as interchain linkages, but some may also be inter-residue linkages.
Effect of high- and low-protein diets on the regulation of rat liver tyrosine aminotransferase and tryptophan oxygenase activities by l-tyrosine and l-tryptophan
Authors: Mukhtar Hasan , Sahib Maharaj K. , Murti Coimbatore R. Krishna
Published: August 2015

Induction of rat liver tyrosine aminotransferase by l-tyrosine and tryptophan oxygenase by l-tryptophan was studied in groups of rats fed on diets containing 18 or 5% protein. The basal activity of hepatic tyrosine aminotransferase of rats receiving 5% protein gradually increased with the age of the animals but that of rats receiving 18% protein did not. l-Tyrosine induced hepatic tyrosine aminotransferase in rats receiving 18% protein when tested at ages from 4 to 20 weeks. When induction by l-tyrosine was carried out in rats receiving the 5% protein diet, significant induction of tyrosine aminotransferase occurred only in 4- or 6-week-old rats. Induction by l-tryptophan of tryptophan oxygenase in liver or the basal activity of this enzyme in liver did not differ between the groups fed on 5 and 18% protein. On changing the diet from 0 to 18% protein, the above-mentioned effects on the induction of hepatic tyrosine aminotransferase were reversed.
The interaction between ribonuclease A and surfactants
Authors: Jones Malcolm N. , Skinner Henry A. , Tipping Edward , Wilkinson Alan
Published: August 2015

1. U.v. difference spectra show that the anionic surfactant sodium n-dodecyl sulphate unfolds ribonuclease A at pH7.3 and 10.3, but that the cationic surfactant n-dodecyltrimethylammonium bromide does not affect the conformation of the enzyme. 2. Equilibrium-dialysis experiments show that sodium n-dodecyl sulphate binds to ribonuclease A, but no binding of n-dodecyltrimethylammonium bromide could be detected at pH7.3. 3. The enzymic activity of ribonuclease A is unaffected by n-dodecyltrimethylammonium bromide up to a concentration of 0.03m at 25°C. 4. Ultracentrifuge studies support the conclusion that n-dodecyltrimethylammonium bromide does not interact significantly with ribonuclease A. 5. The enthalpy change as measured by microcalorimetry on binding of sodium n-dodecyl sulphate to ribonuclease A is consistent with an exothermic enthalpy of binding occurring simultaneously with an endothermic enthalpy of chain unfolding.
Evidence for the binding of the carcinogen 3-methylcholanthrene to both the purine and the pyrimidine bases of hamster fibroblast deoxyribonucleic acid (Short Communication)
Authors: Jones Peter A. , Gevers Wieland , Hawtrey Arthur O.
Published: August 2015

The binding of [3H]3-methylcholanthrene to the DNA of hamster fibroblasts was studied by using chemical methods for DNA degradation. DNA depurinated by mild acid hydrolysis released approximately half of the radioactivity at the same rate as the purine bases, but the resulting apurinic acid still contained radioactive carcinogen.
Superoxide dismutase: a contaminant of bovine catalase (Short Communication)
Authors: Halliwell Barry
Published: August 2015

Some commercial samples of bovine catalase contain superoxide dismutase activity. Therefore the inhibition of a reaction on the addition of a catalase preparation need not necessarily mean that H2O2 is responsible for the reaction.
Collagen cross-linking. Isolation of cross-linked peptides from collagen of chicken bone
Authors: Eyre D. R. , Glimcher M. J.
Published: August 2015

Cross-linked peptides were isolated from chicken bone collagen that had been digested with CNBr or with bacterial collagenase. Analyses of 3H radioactivity in disc electrophoretic profiles of the CNBr peptides from bone collagens that had been treated with NaB3H indicated that a major site of intermolecular cross-linking in chicken bone collagen is located between the carboxy-terminal region of an α1 chain and a small CNBr peptide, probably situated near the amino-terminus of an α1 or α2 chain in an adjacent collagen molecule. A small amount of this cross-linked CNBr peptide was isolated from a CNBr digest of chicken bone collagen by column chromatography. Amino acid analysis showed that the CNBr peptide, α1CB6B, the carboxy-terminal peptide of the α1 chain, was the major CNBr peptide in the preparation, and the reduced cross-linking components were identified as hydroxylysinohydroxynorleucine (HylOHNle), with a smaller amount of hydroxylysinonorleucine (HylNle). However, the composition and the low recovery of the cross-linking amino acids suggested that the preparation was a mixture of CNBr peptides α1CB6B and α1CB6B cross-linked to a small CNBr peptide whose identity could not be determined. A small cross-linked peptide was isolated from chicken bone collagen that had been reduced with NaB3H4 and digested with bacterial collagenase. This peptide was the major cross-linked peptide in the digest and contained a stoicheiometric amount of the reduced cross-linking compounds. A peptide which had the same amino acid composition, but contained the cross-linking compounds in their reducible forms, was isolated from a collagenase digest of chicken bone collagen that had not been treated with NaBH4. The absence of the reduced cross-links from this peptide indicates that, at least for the cross-linking site from which the peptide derives, natural reduction is not a significant pathway for biosynthesis of stable cross-links. However, most of the reducible cross-linking component in the peptide appeared to stabilize in the bone collagen by rearrangement from aldimine to ketoamine form.
Palmitoyl-coenzyme A synthetase. Isolation of an enzyme-bound intermediate
Authors: Bar–Tana J. , Rose G. , Shapiro B.
Published: August 2015

An enzyme-bound intermediate of the overall reaction catalysed by rat liver microsomal long-chain fatty acyl-CoA synthetase is described. It was found to contain equimolar amounts of adenylate and fatty acid moieties bound to protein, and was stabilized by ATP. The intermediate reacted with CoA to give palmitoyl-CoA.
The isolation and partial characterization of ribonuclease A from Bison bison
Authors: Stewart Gordon R. , Stevenson Kenneth J.
Published: August 2015

1. Bison ribonuclease was isolated from pancreas glands of Bison bison by acid extraction, (NH4)2SO4 fractionation, affinity chromatography on Sepharose–5′-(4-aminophenylphosphoryl)uridine 2′,3′-phosphate and ion-exchange chromatography on Bio-Rex-70. 2. The selectivity of the affinity column towards bison ribonuclease in heterogeneous protein solutions was greatly improved by employing piperazine buffers at pH5.3, which decreased non-specific interactions of other proteins. Rapid desorption from the affinity column was obtained with sodium phosphate buffer (pH3). 3. Bison ribonuclease has a total amino acid content very similar to ox ribonuclease. Inactivation of bison ribonuclease with iodoacetic acid leads to the formation of 0.62 residues of π-carboxymethylhistidine and 0.36 residues of τ-carboxymethylhistidine. The amino acid composition of peptides isolated from diagonal peptide ‘maps’ and also of peptides isolated after pH1.6 and 2.4 two-dimensional high-voltage electrophoresis of a digest of bison ribonuclease labelled with pyridoxal 5-phosphate indicates that there is complete homology between ox and bison ribonucleases. 4. The Schiff-base attachment site of pyridoxal 5-phosphate was identified as lysine-41 by NaBH4 reduction followed by peptide isolation.
Qualitative determination of N-terminal amino acids of peptides and proteins with cobalt(III) chelates
Authors: Bentley Keith W. , Creaser Ernest H.
Published: August 2015

1. A method of N-terminal peptide-bond hydrolysis with the cis-β-hydroxyaquo(triethylenetetramine)cobalt(III) ion, i.e. β-[Co(trien)(OH)(OH2)]2+, is reported. The method has been demonstrated with 22 small peptides and ten proteins. 2. The procedure is rapid (an N-terminal amino acid determination can be made easily in one day), it involves no acid hydrolysis step and thus no destruction of labile amino acids, and it involves the use of easily prepared inexpensive reagents. 3. The released N-terminal amino acids can be identified as their cobalt(III) derivatives, or directly as the amino acid or as their dansylated derivatives. 4. The method is to treat 1 μmol of peptide or protein with β-[Co(trien)(OH)(OH2)]2+ reagent at pH8.0, 45°C for 3h. Addition of 0.5m-phosphate buffer, pH10.5 at 45°C for 10min cleaves the N-terminal bidentate amino acid–cobalt complex, which can be identified directly. For greater sensitivity with 10nmol of peptide) the free amino acid is prepared from the complex by treatment (with NaCN (0.1m, 40°C, 30min), or H2S or NaBH4 (25°C, 5min), dried, dansylated and the dansyl-amino acid identified by high-voltage electrophoresis. The method is unaffected by the presence of 4–8m-urea, but will not cleave blocked N-terminal acids.
Nitrogenase of Klebsiella pneumoniae. Interaction of the component proteins studied by ultracentrifugation
Authors: Eady Robert R.
Published: August 2015

Sedimentation-velocity analyses of mixtures of the component proteins of nitrogenase of Klebsiella pneumoniae at a 1:1 molar ratio, showed a single peak of sedimentation coefficient (12.4S) considerably greater than that obtained for the larger (Fe+Mo-containing) protein centrifuged alone (10.4S). When the ratio exceeded 1:1 (the smaller Fe-containing protein in excess) an additional peak corresponding in sedimentation coefficient (about 4.5S) to free Fe-containing protein appeared. When proteins, which had been inactivated by exposure to air were used, no interaction occurred. Na2S2O4 at 2mm both reversed and prevented interaction between the two proteins; sedimentation coefficients corresponded to those of the proteins when centrifuged alone. These results demonstrate the formation of a complex between the nitrogenase proteins, and, together with data of activity titration curves, are consistent with the formulation of the nitrogenase complex of K. pneumoniae as (Fe-containing protein)–(Fe+Mo-containing protein).
Purification and properties of an enzyme duet, phosphoribosylaminoimidazole carboxylase and phosphoribosylaminoimidazolesuccinocarboxamide synthetase, involved in the biosynthesis of purine nucleotides de novo
Authors: Patey Carole Anne Hilton , Shaw Gordon
Published: August 2015

Purification and some properties of an adjacent pair of enzymes in the biosynthetic pathway to AMP, phosphoribosylaminoimidazole carboxylase (EC 4.1.1.21) and phosphoribosylaminoimidazolesuccinocarboxamide synthetase (EC 6.3.2.6), are described. The enzymes form a duet, which may be regarded as a single molecule with a dual function, or as two very similar substances.
The effect of surfactant structure on ribonuclease A–surfactant interactions
Authors: Blinkhorn Cecilia , Jones Malcolm N.
Published: August 2015

The enzymic activity of ribonuclease A was measured in the presence of several surfactants at pH7.2. Cationic surfactants with trimethylammonium and pyridinium head groups do not deactivate or denature the enzyme, whereas n-dodecylamine hydrochloride, like the anionic surfactant sodium n-dodecyl sulphate, deactivates and denatures ribonuclease A.
Inhibition of 3-phosphoglycerate dehydrogenase from Pisum sativum by purine nucleotides
Authors: Slaughter J. Colin
Published: August 2015

The inhibition of 3-phosphoglycerate dehydrogenase from etiolated pea epicotyls by purine nucleoside di- and tri-phosphates is linear, competitive with regard to NADH, and the nucleotides are mutually exclusive in their binding. Free ATP and ADP are more effective inhibitors than are the respective magnesium complexes.
Oxaloacetate decarboxylases of rat liver
Authors: Dean B. , Bartley W.
Published: August 2015

1. Two oxaloacetate decarboxylases of rat liver are described; one is mitochondrial with no metal ion requirement and activity in the alkaline pH region; the other is cytoplasmic with Mn2+or Mg2+requirement and activity around pH5. 2. A method for the partial purification of the mitochondrial enzyme is described. 3. The apparent Km of the mitochondrial enzyme is 0.23mm. 4. Inhibition of the mitochondrial enzyme by substrate, CoA, acetyl-CoA, citrate and phosphoenolpyruvate is described.
Some components of the cholinergic system in the prawn Palaemonetes varians (Leach)
Authors: Veldsema-Currie Robin D.
Published: August 2015

1. An enzyme similar to mammalian acetylcholinesterase is found in high activity in the nervous tissue of Palaemonetes varians, i.e. eyes plus stalks, brain, suboesophageal ganglion and ventral cord. Acetylcholinesterase is also found associated with the abdominal muscles. Multiple enzyme forms are found in extracts of nervous tissues and muscles by electrophoresis and isoelectric focusing. 2. Cholinesterase is present in high activity in the stomatogastric system of P. varians. Three electrophoretically separable forms are found, having isoelectric points at pH4.2, 4.5 and 5.4. 3. Approx. 50% of the total acetylcholinesterase activity, approx. 80% of the choline acetyltransferase activity and 100% of the acetylcholine equivalents are found associated with the nervous tissue. Among the tissues examined, eyes plus stalks were the richest source of both choline acetyltransferase and acetylcholine equivalents. Suboesophageal ganglion and brain also contained large amounts of these components. 4. The distribution of these components could support the function of acetylcholine as a central and/or sensory transmitter in P. varians.
Characterization of the proteins of guinea-pig hair and hair-follicle tissue
Authors: Steinert Peter M. , Rogers George E.
Published: August 2015

This work forms a part of a study of the mechanism and control of protein synthesis in the hair follicle and concerns the characterization of the proteins of hair-follicle tissue and for comparative reasons those of the hair itself. 1. Five different groups of reduced carboxymethylated proteins were delineated from both tissues; these were: group 1A proteins, which appeared to be aggregates of the group 2 proteins; group 1B proteins, soluble at pH4.4, which were thought to originate from the medulla and inner-rootsheath layers; group 2 proteins, which were defined as the main low-sulphur keratin proteins insoluble at pH4.4; group 3 proteins, the precise origin of which is not known; and the group 4 proteins, which were defined as the main high-sulphur keratin proteins soluble at pH4.4. 2. With the single exception of the group 1B proteins, the types and properties of all hair and hair-follicle proteins were identical as far as could be determined by use of such criteria as multiplicity of components, molecular charge, molecular weight and amino acid composition. 3. Two significant quantitative differences were noted: in follicle extracts there were more group 2 proteins but less group 3 and group 4 proteins than in hair extracts; and secondly, in the follicle group 4 proteins, there were more proteins of lowest molecular weight and S-carboxymethylcysteine content, but fewer proteins of the highest molecular weight and S-carboxymethylcysteine conent than in the hair group 4 proteins. 4. These quantitative differences are discussed in terms of the mechanism of synthesis of the keratin proteins. 5. Follicle group 1B proteins are postulated to have arisen from the trichohyalin droplets of the developing medulla and inner-root-sheath layers of the follicle and may be precursors of the proteins of the mature medulla and inner root sheath.
Use of affinity chromatography for the quantitative study of acceptor–ligand interactions: the lactose synthetase system (Short Communication)
Authors: Andrews P. , Kitchen B. J. , Winzor D. J.
Published: August 2015

From the effects of N-acetyl-d-glucosamine and d-glucose on the elution of the A protein of human lactose synthetase from a column of Sepharose–α-lactalbumin, values of 200m−1and 0.57m−1are deduced for the association constants describing the interaction between the enzyme and the respective monosaccharides.
The amino acid sequence around the reactive cysteine residue in human plasma Factor XIII (Short Communication)
Authors: Holbrook J. John , Cooke Rodney D. , Kingston I. Barry
Published: August 2015

1. Activated human plasma factor XIII was 65% inhibited by iodo[14C]acetate and incorporated 0.6 mol of label into the α subunit, which eventually was allowed to form a precipitate. 2. All the label was recovered as S-carboxymethylcysteine in a tetra-peptide of sequence Gly-Gln-Cys-Trp.
Transport of methionine and proline by rat liver slices and the effect of certain hormones
Authors: Crawhall J. C. , Purkiss P.
Published: August 2015

1. Transport characteristics of l-methionine and l-proline in rat liver slices in vitro were studied. 2. Intracellular concentration gradients for methionine were obtained. 3. Methionine uptake was inhibited by iodoacetate, dinitrophenol, Na+-free media and also by glycine, lysine, cysteine and dithiothreitol but not by α-aminoisobutyrate. 4. The rate of methionine metabolism in the slice was slow. 5. Puromycin inhibited methionine incorporation into protein, but not methionine uptake. 6. Methionine inhibited the transport of α-aminoisobutyrate but not of cystine. 7. Efflux and exchange diffusion of methionine was studied. 8. Amino acid transport in rat liver slices was not affected by thyroidectomy. 9. Addition of insulin, glucagon, adrenaline or cortisol did not affect the transport of methionine. 10. Addition of 6-N,2′-O-dibutyryladenosine 3′:5′-cyclic monophosphate increased methionine transport after a 120min incubation period in some experiments. 11. Studies of l-proline transport were invalidated because of the rapid evolution of CO2 from the substrate.
The effect of Triton WR-1339 on the subcellular distribution of Trypan Blue and 125I-labelled albumin in rat liver
Authors: Davies Malcolm
Published: August 2015

1. The density-gradient distribution patterns of acid phosphatase, Trypan Blue and denatured 125I-labelled albumin were studied by discontinuous sucrose- and isopycnic sucrose-density-gradient centrifugation on combined heavy and light mitochondrial (M+L) fractions of liver isolated from normal rats and from rats injected with Triton WR-1339. 2. The results obtained from the subfractionation of the M+L pellet of normal animals indicate that the equilibrium density of Trypan Blue and acid-insoluble radioactivity is the same as that for acid phosphatase, which suggests they are bound by a common membrane to form a distinct subcellular population of lysosomal nature. 3. In contrast, the analysis of the isopycnic gradients obtained on subfractionation of M+L pellets of liver isolated from rats treated with Triton WR-1339 show that the acid-insoluble radioactivity has an equilibrium density around 1.21, whereas the acid hydrolases, including cathepsin D, show the characteristic shift to an equilibrium density of around 1.12. Trypan Blue is distributed along the gradient with distinct peaks at densities 1.22 and 1.12. 4. Similar equilibrium-density distribution patterns were obtained with M+L pellets isolated from rats pretreated with Triton WR-1339 but not injected with Trypan Blue. 5. Treatment of the rats with Triton WR-1339 does not affect albumin digestion of isolated intact lysosomes despite the fact that most of the cathepsin D and the albumin ingested by phagocytosis are located in different vacuoles. 6. It is concluded from these experiments that in the liver of animals treated with Triton WR-1339 125I-labelled albumin is located within heterophagosomes which do not fuse with heterolysosomes containing the non-ionic detergent Triton WR-1339. The inability of these two lysosomal populations to fuse is not due to Trypan Blue.
Oscillations of enzyme activities during the cell-cycle of a glucose-repressed fission-yeast Schizosaccharomyces pombe 972h
Authors: Poole R. K. , Lloyd D.
Published: August 2015

1. Increased specific activities of cytochrome c oxidase, catalase, succinate dehydrogenase, succinate–cytochrome c oxidoreductase, NADH–cytochrome c oxidoreductase and malate dehydrogenase were observed during glucose de-repression of Schizosaccharomyces pombe. 2. The cell-cycle of this organism was analysed by three different methods: (a) harvesting of cells at intervals from a synchronous culture, (b) separation of cells by rate-zonal centrifugation into different size classes and (c) separation of cells by isopycnic-zonal centrifugation into different density classes. 3. Measurement of enzyme activities during the cell-cycle showed that all the enzymes assayed [cytochrome c oxidase, catalase, acid p-nitrophenylphosphatase, NADH-dehydrogenase, NADH–cytochrome c oxidoreductase, NADPH–cytochrome c oxidoreductase, succinate dehydrogenase, malate dehydrogenase, isocitrate dehydrogenase (NADP) and fumarate hydratase] show periodic expression as ‘peaks’. 4. Cytochrome c oxidase shows a single maximum at 0.67 of a cycle, whereas succinate dehydrogenase exhibits two maxima separated by 0.5 of a cell-cycle. 5. All other enzymes assayed showed two distinct maxima per cell-cycle; for catalase, malate dehydrogenase and NADPH–cytochrome c oxidoreductase there is the possibility of multiple fluctuations. 6. The single maximum of cytochrome c oxidase appears at a similar time in the cycle to one maximum of each of the other enzymes studied, except for NADH dehydrogenase. 7. These results are discussed with reference to previous observations on the expression of enzyme activities during the cell-cycle of yeasts.
The formation of choline O-sulphate by Pseudomonas C12B and other Pseudomonas species
Authors: Fitzgerald John W.
Published: August 2015

Pseudomonas C12B and other Pseudomonas species released larger amounts of a 35S-labelled metabolite into the medium when cultured on growth-limiting concentrations of Na2SO4 as opposed to growth in SO42--sufficient media. The metabolite was found at all stages of the culture cycle of Pseudomonas C12B and maximum quantities occurred in stationary-phase culture supernatants. The metabolite was not detected when the bacterium was cultured on growth-limiting concentrations of potassium phosphate. The amount of the metabolite present in the medium greatly exceeded that which could be extracted from intact cells and, except for choline chloride, it was independent of the carbon source used for growth. If choline chloride was present in high concentration, then larger amounts of the metabolite were found in the culture medium. The metabolite was not detected extracellularly or intracellularly when the bacterium was grown in SO42--deficient media containing 5mm-l-cysteine. The same metabolite was also synthesized in vitro only when Pseudomonas C12B extracts were incubated with choline chloride, ATP, MgCl2 and Na235SO4. The metabolite-forming system was not subject to repression by Na2SO4 and was completely inhibited by 0.5mm-l-cysteine and activated by Na2SO4 (up to 1.0mm). The metabolite was identified as choline O-sulphate by electrophoresis, chromatography and isotope-dilution analysis. Another 35S-labelled metabolite was also detected in culture supernatants, but was not identified.
Clearing-factor lipase in muscle and adipose tissue of pigs
Authors: Enser M.
Published: August 2015

1. Clearing-factor lipase was assayed in acetone–ether-dried powders of heart and adipose tissue of pigs. The enzyme activity in heart was higher than that in adipose tissue. The activity in the outer layer of subcutaneous fat was greater than that in the inner subcutaneous fat and the perirenal fat, which had similar activities. 2. Starvation for 48h, but not for 24h, decreased the activity of the heart enzyme. 3. Starvation for 24h caused a rapid decrease in the activity in all three adipose tissues, but even after 72h of starvation the activity was still highest in the outer subcutaneous fat. 4. Plasma fatty acid, glucose and insulin concentrations were determined in fed and starved pigs. Starvation decreased the plasma insulin concentration and increased the non-esterified fatty acid concentration.
Thymidine metabolism in regenerating rat liver one to two hours after partial hepatectomy
Authors: Ord Margery G. , Stocken Lloyd A.
Published: August 2015

1. When [3H]thymidine was injected intravenously into rats in amounts up to 40mg/kg body wt. and the3H radioactivity in the livers measured at 30min, saturation kinetics for thymidine uptake were not found. If the animals were examined 3 min after intravenous injection, saturation could be attained in normal rats with 12mg of thymidine/kg and in partially hepatectomized rats with 4mg/kg. At concentrations of thymidine close to saturation, no differences were found in rate or amount of uptake/g of liver between normal and partially hepatectomized rats 1–2h after operation. 2. Perfusion techniques were used to compare thymidine uptakes in the two sets of rats at concentrations up to 40μm-thymidine. Uptakes with tracer amounts of thymidine after 30min were identical in vivo and in the perfusion studies and were twice as great in livers from partially hepatectomized rats with concentrations up to 40μm-thymidine. 3. At 1.5h after operation there was nearly twice as much β-aminoisobutyrate present per g of liver from partially hepatectomized as compared with normal rats.
Quantitative studies on ferredoxin in greening bean leaves
Authors: Haslett B. G. , Cammack R. , Whatley F. R.
Published: August 2015

Two methods of measuring small amounts of the iron–sulphur protein ferredoxin are described. One involves measurements of the signal at g=1.96 produced by reduced ferredoxin in an e.p.r. (electron-paramagnetic-resonance) spectrometer; the other depends on the rate of ferredoxin-dependent electron transport in a chloroplast bioassay measured in an O2 electrode. These methods of measurement were used to examine the development of ferredoxin during the greening of etiolated bean leaves. Ferredoxin is present in low concentrations in the leaves and cotyledons of 14-day-old etiolated beans (Phaseolus vulgaris L. var. Canadian Wonder), and develops in a linear manner with time when the leaves are illuminated. This synthesis appears to be independent of chlorophyll synthesis during the early stages of greening. However, the chlorophyll/ferredoxin ratio reaches a final value of approx. 360 irrespective of the light intensity, indicating the existence of a control mechanism operative in deciding the stoicheiometry of these components in the mature chloroplast. The ferredoxin synthesis appears to be light-dependent, and red light is the most effective in its promotion. The effect of red illumination is not reversed by far-red light, indicating the absence of a phytochrome control of ferredoxin synthesis. From experiments using specific inhibitors of chloroplast protein synthesis, it is concluded that ferredoxin is synthesized on cytoplasmic ribosomes.
Metabolic adaptations during lactogenesis. Fatty acid and lactose synthesis in cow mammary tissue
Authors: Mellenberger R. W. , Bauman D. E. , Nelson D. R.
Published: August 2015

1. Mammary-tissue biopsies were obtained from multiparous cows at 30 and 7 days pre partum and 7 and 40 days post partum. Investigations of the effect of lactogenesis on fatty acid and lactose synthesis involved measurements of biosynthetic capacity (tissue-slice incubations in vitro) and activities of relevant enzymes. 2. Fatty acid synthesis from acetate increased over 20-fold from 30 days pre partum to 40 days post partum. Changes in the lipogenic capacity of mammary-tissue slices more closely paralleled increases in the activities of acetyl-CoA carboxylase (EC 6.4.1.2) and acetyl-CoA synthetase (EC 6.2.1.1) than of other enzymes involved in acetate incorporation into fatty acids or in NADPH generation. 3. Lactose biosynthesis by mammary-tissue slices, lactose synthetase activity (EC 2.4.1.22) and α-lactalbumin concentration were all negligible at 30 days pre partum but increased 2.5–4-fold between 7 days pre partum and 40 days post partum. Phosphoglucomutase (EC 2.7.5.1), UDP-glucose pyrophosphorylase (EC 2.7.7.9) and UDP-glucose 4-epimerase (EC 5.1.3.2) had substantial activities at 30 days pre partum and increased less dramatically during lactogenesis. 4. Results are consistent with acetyl-CoA carboxylase and perhaps acetyl-CoA synthetase representing the regulatory enzyme(s) in fatty acid synthesis, with lactose synthetase (α-lactalbumin) serving a similar function in lactose biosynthesis.
Biosynthesis of respiratory-tract mucins. Incorporation of radioactive precursors into glycoproteins by canine tracheal explants in vitro
Authors: Ellis Daniel B. , Stahl Glenn H.
Published: August 2015

1. Canine tracheal explants, cultured in medium 199, actively incorporated radioactive precursors into secreted macromolecules in vitro. 2. Puromycin, 6-diazo-5-oxo-l-norleucine and ouabain markedly inhibited the incorporation of these precursors. 3. Exogenous glucosamine at concentrations above 20mm caused a greater than 50% inhibition of the incorporation of l-[G-3H]fucose and l-[U-14C]serine. 4. Carbohydrate content of the purified secretions was approximately 50% and consisted principally of galactose, N-acetylglucosamine, N-acetylgalactosamine, fucose and sialic acids. 5. Chromatography on DEAE-cellulose and Bio-Gel A-150m and equilibrium density-gradient centrifugation in a CsCl gradient confirmed the presence of mucous glycoproteins. 6. Electrophoresis on 1% agarose gels gave profiles that were identical with canine respiratory mucus obtained in vivo. 7. These results support the utility of the explant system for studies of respiratory secretions.
Studies on selenium incorporation into, and electron-transfer function of, liver microsomal fractions from normal and vitamin E-deficient rats given phenobarbitone
Authors: Caygill Christine P. J. , Diplock A. T. , Jeffery Elizabeth H.
Published: August 2015

1. The incorporation of75Se from Na275SeO3 into the liver endoplasmic reticulum of rats given phenobarbitone was investigated by using a zonal centrifuge technique. 2. It was found that, in rats deprived of vitamin E, or of vitamin E and selenium, phenobarbitone was without effect on the incorporation of75Se or on its conversion to75Se2-. When vitamin E was given at the same time as the phenobarbitone and75Se, there was a large increase in the amount of75Se and75Se2-found in the smooth reticulum. It is concluded that there may be a specific vitamin E-dependent role for selenium and selenide in the smooth endoplasmic reticulum, and it is suggested, in the light of these and other observations, that the selenide may form a part of the active centre of a non-haem iron-containing protein ‘X‘, that may function in microsomal electron transport. 3. Measurements of the contents of cytochromes P-450 and b5 in liver microsomal fractions of rats given vitamin E-deficient, and vitamin E- and selenium-deficient diets, showed that haemoprotein biosynthesis is unimpaired in these rats and phenobarbitone treatment resulted in the expected increase in the haemoproteins. 4. When the reduction of cytochrome P-450 by NADH and NADPH was measured, no difference was found between normal and deficient animals. 5. These results are discussed in relation to current knowledge of microsomal electron transfer.
Biliary excretion of some diquaternary ammonium cations in the rat, guinea pig and rabbit
Authors: Hughes Robin D. , Millburn Peter , Williams R. Tecwyn
Published: August 2015

1. The extent of the excretion in the bile and urine of the14C-labelled dications, diquat, paraquat, morfamquat, decamethonium and dimethyltubocurarine in bile-duct-cannulated rats, guinea pigs and rabbits was examined. 2. These compounds were excreted unchanged in bile and urine, except diquat, which was metabolized to a significant extent (18% of the dose) in the rabbit only. 3. The extent of the biliary excretion of diquat (mol wt. of ion 184), paraquat (186), decamethonium (258) and morfamquat (469) was less than 10% of the dose in the three species, whereas that of dimethlytubocurarine (653) was greater than 10% in the rat and rabbit but not in the guinea pig. 4. These results together with data from the literature suggest that the molecular weight at which the excretion of dications in the bile exceeds 10% of the dose is in the region of 500–600, which differs from the values for monocations (Hughes et al., 1973) and anions (Millburn et al., 1967; Hirom et al., 1972).
Studies on cystathionase activity in rat liver and brain during development. Effects of hormones and amino acids in vivo
Authors: Heinonen Kirsti
Published: August 2015

High activity of cystathionase was present in rat liver but only low amounts of activity in rat brain during development. Triamcinolone had no effect on liver cystathionase activity in foetuses but increased the enzyme activity significantly in postnatal rats. l-Thyroxine decreased liver cystathionase activity significantly in newborn rats; administration of pyridoxal 5′-phosphate did not prevent this effect. l-Methionine significantly increased liver cystathionase activity in newborn rats.
Biosynthesis of carnitine and 4-N-trimethylaminobutyrate from 6-N-trimethyl-lysine
Authors: Cox Richard A. , Hoppel Charles L.
Published: August 2015

The conversion of 6-N-[Me-14C]trimethyl-lysine into carnitine and 4-N-trimethylaminobutyrate (butyrobetaine) was demonstrated in rats kept on a lysine-deficient diet. After the rats were given [14C]trimethyl-lysine for 4 days, a total of 17% of the injected label was recovered as carnitine from carcass and urine extracts. Another 8% of the trimethyl-lysine label was converted into 4-N-trimethylaminobutyrate, most of which was recovered from the urine. The conversion of trimethyl-lysine into the above two metabolites supports the pathway of carnitine biosynthesis as lysine+methionine → 6-N-trimethyl-lysine → 4-N-trimethylaminobutyrate → carnitine. In addition, three other metabolites representing 2% of the injected dose were recovered. Only an insignificant portion of the label was recovered as free trimethyl-lysine from the carcass, whereas 22% of the injected label was recovered in the urine. A relatively low specific radioactivity in carnitine was found when 5-N-[Me-14C]trimethylaminopentanoate and 6-N-[Me-14C]trimethylaminohexanoate were administered to rats in amounts similar to the [14C]trimethyl-lysine, suggesting that they were not free intermediates.
Sexual differences in galactose metabolism: galactosyl ceramide galactosidase and other galactosidases in mouse kidney
Authors: Lin Yuh-Nan , Radin Norman S.
Published: August 2015

Male and female mice were compared at two ages, 15 and 50 days, with respect to the activities of three galactosidases in kidney. No sex difference in enzyme activity was seen in the young mice, but appreciable differences were found in the older animals. The male kidneys had about one-third higher specific activities of cerebroside β-galactosidase and nitrophenyl β-galactosidase, but there was no difference with nitrophenyl α-galactosidase. A listing and discussion of other galactose-metabolizing enzymes influenced by sex differences is presented.
Distribution of sulphate and iduronic acid residues in heparin and heparan sulphate
Authors: HÖÖk Magnus , Lindahl Ulf , Iverius Per-Henrik
Published: August 2015

1. A method was developed for determination of the uronic acid composition of heparin-like glycosaminoglycans. Polymers or oligosaccharides are degraded to monosaccharides by a combination of acid hydrolysis and deamination with HNO2. The resulting uronic acid monosaccharides (accounting for about 70% of the uronic acid contents of the starting materials) are isolated and converted into the corresponding aldono-1,4-lactones, which are separated by g.l.c. The calculated ratios of glucuronic acid/iduronic acid are reproducible within 5%. 2. Samples of heparin from pig intestinal mucosa (molar ratio of sulphate/disaccharide unit, 2.40) and heparan sulphate from human aorta (sulphate/disaccharide ratio, 0.46) were subjected to uronic acid analysis. l-Iduronic acid constituted 77% and 19% respectively of the total uronic acid contents. 3. The correlation between the contents of sulphate and iduronic acid indicated by this finding also applied to the fractionated deamination products of the two polymers. The sulphated fragments varied in size from disaccharide to octasaccharide (or larger) and showed sulphate/disaccharide molar ratios in the range of 0.05–2.0. The proportion of iduronic acid increased with increasing ester sulphate contents of the oligosaccharides. 4. Previous studies on the biosynthesis of heparin in a cell-free system have shown that l-iduronic acid residues are formed by C-5 epimerization of d-glucuronic acid units at the polymer level; the process requires concomitant sulphation of the polymer. The results obtained in the present structural study conform to these findings, and suggest further that similar mechanisms may operate in the biosynthesis of heparan sulphate. The epimerization reaction appears to be linked to the sulphation of hydroxyl groups but does not seem to require sulphation of the target uronic acid residues. The significance of sulphamino groups in relation to the formation of iduronic acid is unknown.