bluuebunny/crossref_metadata_2025 huggingface dataset:
Coproporphyrinogenase in tobacco (Nicotiana tabacum L.)
Authors: Hsu W. P. , Miller G. W.
Published: August 2015

1. Coproporphyrinogenase was extracted and purified from tobacco (Nicotiana tabacum L.). Enzyme activity was mainly located in mitochondria rather than in chloroplasts. The enzyme was purified by differential centrifugation, ammonium sulphate fractionation, calcium phosphate gel adsorption and dialysis. A 69-fold final purification was obtained. 2. An apparent Km value of 3.6×10−5m was found, the value being largely dependent on the amount of coproporphyrin III recovered after reduction with sodium amalgam to coproporphyrinogen III. Protoporphyrin formation was linear up to 3h and decreased with further incubation. The enzyme activity increased with the concentration of enzyme protein up to 30μg/ml of solution. 3. Enzyme activity was greatly enhanced by increasing Fe2+ concentrations up to 0.5mm, beyond which inhibition occurred. Co2+ and Mn2+ were also found to activate at low concentrations (0.1mm) and inhibit at higher concentrations (5mm). Fe3+ and Cu2+, both at 0.1mm, and o-phenanthroline and EDTA, each at 1mm, were found to be inhibitory.
Topography of nucleic acid helices in solutions. The interaction specificities of optically active amino acid derivatives
Authors: Gabbay Edmond J. , Kleinman Roberta W.
Published: August 2015

The synthesis and interactions of the d- and l-enantiomers of the amino acid amide derivatives [Formula: see text] (I) and lysyl dipeptides [Formula: see text] (II) with poly rI·poly rC, poly rA·poly rU and calf thymus DNA is reported. The following results were found. (1) The degree of stabilization of the helices as measured by the Tm (‘melting’ temperature) of the helix–coil transition was dependent on the nature of the amino acid. (2) For the poly rI·poly rC helix, the l-enantiomers of salts (I) and (II) stabilized more than the d-enantiomers. The same was true for calf thymus DNA in the presence of salts (II) and for poly rA·poly rU in the presence of salts (II) and the proline derivatives of salts (I). (3) As R increased in size and became more apolar, the amount of stabilization of the poly rI·poly rC helix in the presence of salts (I) decreased. On the other hand, the amount of stabilization increased with more polar substituents. An attempt was then made to determine whether the difference in stabilization of the double-stranded helices at the Tm in the presence of the l- and d-enantiomers of salts (I) is due to the interaction with the helix, the random coil or both. A new method was developed for determining the binding of small ions to polyions that involves a competition between an insoluble polystyrene ion-exchange resin and the soluble polyion for the counterion. Dissociation constants are obtained for the complexes of single- and double-stranded helices with the salts (I). The results are illuminating and indicate that with certain helices, i.e. poly rA·poly rU, the interactions of salts (I) with the single strands may not be ignored. It is concluded that the high optical specificity found in Nature, i.e. d-ribose in nucleic acids and l-amino acids in proteins, cannot be attributed solely to monomer–polymer interactions described by Gabbay (1968).
Some effects of testosterone on the rat ventral prostate
Authors: Parsons I. C. , Mangan F. R. , Neal G. E.
Published: August 2015

1. Labelled testosterone, injected directly into the ventral prostate of castrated rats became associated, in part, with a cytoplasmic high-molecular-weight fraction, fraction ‘A’. 2. The label present in fraction ‘A’ was found to be mainly associated with dihydrotestosterone. 3. Unlike fraction ‘A’ from testosterone-pelleted castrated rats, fraction ‘A’ obtained from untreated castrated rats, 48h or more after castration, was strongly inhibitory towards Escherichia coli RNA polymerase in vitro. 4. The inhibition of RNA polymerase by fraction ‘A’ from castrated rats was not changed by the addition of testosterone or dihydrotestosterone in vitro, but pre-heating it to 80°C resulted in a loss of its inhibitory capacity. 5. Fraction ‘A’ from castrated rats contained ribonuclease activity. The elution profile of ribonuclease activity from Sephadex columns indicated that this activity was responsible for the inhibitory effect on the RNA polymerase assays. 6. It is concluded that, unlike the inhibitor present in the uterus of ovariectomized rats (Talwar, Segal, Evans & Davidson, 1964), no direct connexion exists between the steroid-binding capacity of prostatic fraction ‘A’ and its effect on E. coli RNA polymerase activity in vitro.
Pathways leading to and from serine during growth of Pseudomonas AM1 in C1 compounds or succinate
Authors: Heptinstall J. , Quayle J. R.
Published: August 2015

1. The following enzymes of the phosphorylated pathway of serine biosynthesis have been found in methanol- and succinate-grown Pseudomonas AM1: phosphoglycerate dehydrogenase, phosphoserine-α-oxoglutarate aminotransferase and phosphoserine phosphohydrolase. Their specific activities were similar in the organism grown on either substrate. 2. A procedure for preparation of auxotrophic mutants of Pseudomonas AM1 is described involving N-methyl-N′-nitro-N-nitrosoguanidine as mutagen and a penicillin enrichment step. 3. A mutant, M-15A, has been isolated that is unable to grow on methanol and that lacks phenazine methosulphate-linked methanol dehydrogenase. The mutant is able to grow on methylamine, showing that the amine is not oxidized by way of methanol. 4. Loss of methanol dehydrogenase activity in mutant M-15A led to loss of phenazine methosulphate-linked formaldehyde dehydrogenase activity showing that the same enzyme is probably responsible for both activities. 5. A mutant, 20B-L, has been isolated that cannot grow on any C1 compound tested but can grow on succinate. 6. Mutant 20B-L lacks hydroxypyruvate reductase, and revertants that regained the ability to grow on methanol, methylamine and formate contained hydroxypyruvate reductase activity at specific activities similar to that of the wild-type organism. This shows that hydroxypyruvate reductase is necessary for growth on methanol, methylamine and formate but not for growth on succinate. 7. The results suggest that during growth of Pseudomonas AM1 on C1 compounds, serine is converted into 3-phosphoglycerate by a non-phosphorylated pathway, whereas during growth on succinate, phosphoglycerate is converted into serine by a phosphorylated pathway.
Activation of δ-aminolaevulate synthetase in extracts of Rhodopseudomonas spheroides
Authors: Marriott J. , Neuberger A. , Tait G. H.
Published: August 2015

1. The spontaneous activation of δ-aminolaevulate synthetase in extracts from Rhodopseudomonas spheroides grown semi-anaerobically in the light requires oxygen and does not take place anaerobically in the dark. 2. Activation is completely prevented by azide or cyanide and is partially inhibited by chlorpromazine. These compounds inhibit markedly the succinoxidase activity of extracts. 3. NADH delays activation, but when it has been oxidized by the extract activation begins at the normal rate and complete activation occurs. By contrast both the rate and the extent of activation are markedly decreased by even small amounts of carboxylic acids. 4. The inhibitory effects of succinate and citrate on activation can be prevented by malonate and fluorocitrate respectively. 5. These results suggest that for activation to occur some endogenous compound has to be oxidized via the electron transport chain. 6. Activation occurs under anaerobic conditions in the light, probably by photo-oxidation.
Free and protein-bound glutathione in normal and cataractous human lenses
Authors: Harding J. J.
Published: August 2015

Protein-bound glutathione was identified and measured in normal and cataractous human lenses. In a major group of cataracto us lenses the bound glutathione concentration was higher than normal. Study of normal lenses showed that their glutathione content is age-dependent, decreasing steadily from about 3.5μmol/g of lens at age 20 years to about 1.8μmol/g of lens at age 65 years. Cataract brings further decreases.
N-Iodoacetyl-d-glucosamine, an inhibitor of growth and glycoside uptake in Escherichia coli
Authors: Kent P. W. , Ackers J. P. , White R. J.
Published: August 2015

1. Synthesis of N-iodoacetyl-d-glucosamine and its N-iodo[1,2-14C2]acetyl form has been achieved from the tetra-O-acetyl amino sugar and iodoacetic acid in the presence of dicyclohexylcarbodi-imide followed by catalytic deacetylation. 2. N-Iodoacetylglucosamine (up to 0.1mm) linearly inhibits uptake (up to 1min) of methyl α-d-glucoside by Escherichia coli ML308 and K12. Uptake of methyl β-d-thiogalactoside and glycerol is also inhibited. 3. Growth of the organism (strain ML308) on glucose, succinate and glycerol is strongly inhibited by the iodoacetyl compound. The inhibition is relieved by N-acetylglucosamine. 4. The inhibitor has multiple effects, some of which are considered to be intracellular. 5. A separate transport pathway exists for N-acetylglucosamine by means of which the iodoacetyl analogue may enter the cell.
Liquid-scintillation-spectrometer data processing by desk-top computer
Authors: Williams M. A. , Cope G. H. , Jackson J. L. , Hill P.
Published: August 2015

A desk-top computing system has been programmed to store accurately the quench curves necessary for the calculation of total disintegrations/minute (d.p.m.) of samples containing either one or two radioactive isotopes. In producing d.p.m. values background counts are subtracted, and in binary-labelled samples the counts attributable to each radioactive isotope are separated. The programme also relates d.p.m. to the weight, volume and density of the sample. Each variable is easily recalled and adjustments can be made for different batches of samples without reprogramming. Equally easily changes of radioactive isotope, quenching agent, scintillator or window setting can be accommodated. Quadratic equations are used to express the quench curves. Counting efficiencies obtained when the coefficients in the quadratic equations are derived from three carefully chosen points on a quench curve are compared with those obtained when the coefficients are derived by the method of least squares. The results of both mathematical approximations are compared with the efficiencies read by the eye from graphs.
Purification and properties of two extracellular β-lactamases from Bacillus cereus 569/H
Authors: Kuwabara S.
Published: August 2015

1. When Bacillus cereus 569/H was grown in a casamino acid (casein-hydrolysate) medium containing zinc sulphate rapid production of extracellular β-lactamase II preceded that of β-lactamase I. 2. β-Lactamase I was separated from β-lactamase II by fractional precipitation with ammonium sulphate. 3. β-Lactamase I was purified by a process involving chromatography on Celite and DEAE-cellulose and β-lactamase II by chromatography on DEAE-cellulose after denaturation of β-lactamase I by heat. Both enzymes were obtained in crystalline form. 4. β-Lactamase II prepared in this way appeared to have a higher molecular weight than β-lactamase I and required Zn2+ as a cofactor for both cephalosporinase and penicillinase activities.
The structure of anticapsin, a new biologically active metabolite of Streptomyces griseoplanus
Authors: Neuss N. , Molloy B. B. , Shah R. , DeLaHiguera N.
Published: August 2015

1. Physical and analytical data obtained on crystalline anticapsin indicated the empirical formula C9H13NO4. Spectral data (u.v., i.r. and proton magnetic resonance) and formation of l-tyrosine on hydrolysis revealed the functionalities and carbon skeleton of the new epoxy keto amino acid. 2. The optical properties of anticapsin (optical rotatory dispersion and circular dichroism) permitted assignment of absolute configuration to the new metabolite. 3. Treatment of anticapsin with hot methanolic hydrochloric acid followed by acetylation gave C18H19NO5, the α-alkoxycyclohexenone derivative. Analysis of the nuclear-magnetic-resonance and mass spectra of the latter allowed its structure to be determined and confirmed the assigned structure of anticapsin.
Sporulation in Bacillus subtilis. Correlation of biochemical events with morphological changes in asporogeneous mutants
Authors: Waites W. M. , Kay D. , Dawes I. W. , Wood D. A. , Warren S. C. , Mandelstam J.
Published: August 2015

A comparison was made of morphological changes and successive, mainly biochemical, marker events for sporulation in 14 asporogenous mutants. The morphological and biochemical sequences are linked so that arrested development in one is accompanied by corresponding effects in the other. Thus mutants that fail to produce both protease and antibiotic do not progress beyond stage 0, formation of alkaline phosphatase appears to be associated with the transition from stage II to stage III and glucose dehydrogenase with that from stage III to stage IV. Stage II mutants may produce `pygmy' cells or other bizarre cell-division forms. The biochemical sequence is dependent in the sense that if the occurrence of any one event is blocked that of all the succeeding events is also blocked. This has implications for biochemical models that have been proposed to explain the temporal sequence observed in spore development.
Purification and amino acid sequence of melanocyte-stimulating hormone from the dogfish Squalus acanthias
Authors: Lowry P. J. , Chadwick A.
Published: August 2015

A melanocyte-stimulating hormone (MSH) was isolated by gel filtration and ion-exchange chromatography from extracts of the pituitary glands of dogfish. Sequence studies were carried out on the hormone and its enzymically and chemically cleaved fragments. The sequence of the hormone, Ser-Met-Glu-His-Phe-Arg-Trp-Gly-Lys-Pro-Met, shows that ten of its 11 residues are the same as ten of the 13 residues of mammalian α-MSH. About half of its molecules have the carboxyl group at the C-terminus free and about half are amidated; about a fifth have an extra tyrosine residue on the N-terminus, thereby making 11 residues the same as in mammalian α-MSH. Unlike the mammalian hormone, however, none of it was found to be N-acetylated.
Ultrastructural studies on mitochondrial swelling
Authors: Butler W. H. , Judah J. D.
Published: August 2015

Mitochondrial swelling induced by valinomycin, calcium chloride and Pi was studied after potassium permanganate fixation in suspension. Valinomycin induces a rapid K+ influx, increase of the matrix space and out-folding of the cristae, with good preservation of the matrix material. This swelling is reversible but the cristae do not completely re-form and have a blebbed appearance. On repeated swelling and contraction cycles there is a gradual loss of matrix material. Calcium chloride and Pi produce a slow swelling of the matrix space. Shrinkage induced by ATP was partial and not associated with return to the original structure.
Selective inhibition of glucose oxidation by triethyltin in rat brain in vivo
Authors: Cremer J. E.
Published: August 2015

Results are reported of a comparative study in vivo of the metabolism of [2-14C]-glucose and [1-14C]acetate in brains of rats intoxicated with triethyltin sulphate. The incorporation of 14C from glucose into glutamate, glutamine, γ-aminobutyrate and aspartate was greatly decreased. The incorporation of 14C from acetate into these amino acids was unaffected. The experimental data indicated that the main action of triethyltin was to decrease the rate at which pyruvate formed from glucose is oxidized. Glycolysis was not inhibited. Changes in glucose metabolism in the brain are shown not to be directly due to hypothermia. Some of the advantages of measuring the labelling of intermediates at very short time intervals after the injection of the labelled glucose are demonstrated.
Haem a, cytochrome c and total protein turnover in mitochondria from rat heart and liver
Authors: Aschenbrenner V. , Druyan R. , Albin R. , Rabinowitz M.
Published: August 2015

Haem a and cytochrome c were isotopically labelled in mitochondria from rat heart and liver after injection of δ-amino[2,3-3H2]laevulate, a specific haem precursor. [guanido-14C]Arginine or l-[4,5-3H2]leucine were used to label mitochondrial proteins. Half-lives were measured from biological decay in vivo and were similar (5.5–6.2 days) for haem a, cytochrome c and [14C]arginine-labelled proteins. Labelling of hepatic mitochondrial proteins with [3H2]leucine resulted in a prolonged apparent half-life.
Ribosomes from Xenopus laevis ovaries and the polyuridylic acid-directed biosynthesis of polyphenylalanine
Authors: Cox R. A. , Ford P. J. , Pratt Hester
Published: August 2015

Single ribosomes account for 90% or more of the ribosome fraction of Xenopus laevis ovaries. These ribosomes have low endogenous activity in a cell-free protein-synthesizing system but synthesize polyphenylalanine in the presence of poly U under conditions comparable with those reported for mammalian or bacterial systems. These observations agree with the view that the ribosome is deficient in mRNA.
Disappearance of radioactivity from the various ribonucleic acid pools and acid-soluble fractions of mouse liver and kidney after a single injection of labelled orotic acid. The effect of castration
Authors: Avdalovic Nebojša
Published: August 2015

1. An attempt was made to study the rate of synthesis as well as the distribution of RNA in the various cellular fractions in the livers and kidneys of normal and castrated mice. 2. The tissue was fractionated by the procedure of Blobel & Potter (1967). By using this method it was not possible to find any pronounced difference in the relative proportions of RNA in isolated subcellular fractions when kidneys from normal and castrated mice were compared. On the other hand there was an indication of a shift toward the bound ribosomes in livers from normal mice in comparison with livers from castrated mice. 3. Disappearance of the radioactivity followed the pattern of the first-order reaction. Comparing the half-lives of RNA in liver and kidneys it was found that in the latter in both groups of animals half-lives were shorter no matter which cellular fraction was studied. 4. The half-lives for total homogenate RNA, total ribosomal RNA and low-molecular-weight RNA from kidneys of castrated mice were approximately 20–25% longer than the half-lives for the corresponding fractions from normal mouse kidneys. 5. An explanation is put forward for the anomalous finding that RNA from the castrated-mouse kidneys has a higher specific radioactivity than that isolated from normal mice.
The use of zonal centrifugation to study membrane formation during the life cycle of mammalian cells. Synthesis of ‘marker’ enzymes and other components of cellular organelles
Authors: Warmsley A. M. H. , Phillips Beverly , Pasternak C. A.
Published: August 2015

1. The activity of enzymes characteristic of microsomes (NADPH–cytochrome c reductase and uridine diphosphatase) and of inner mitochondrial membranes (cytochrome c oxidase and succinate–cytochrome c reductase) increases during the cell cycle of P815Y neoplastic mast cells in concert with total protein. The activity of glutamate dehydrogenase, an enzyme of the mitochondrial matrix, increases in a somewhat different manner. 2. The specific activity of mitochondrial structures involved in energy-coupling measured with a fluorescent probe remains constant during the cell cycle. 3. Mitochondrial and microsomal protein increases during the cycle at the same time as total protein; nuclear protein increases rather more sharply. 4. The rate of incorporation of labelled choline or inositol into nuclear, mitochondrial or microsomal phospholipid during the cell cycle follows the rate of incorporation into total phospholipid. 5. It is concluded that the major components of cellular membranes are synthesized, like total protein or phospholipid, throughout most of the intermitotic period.
Disulphide bridges of the heavy chain of human immunoglobulin G2
Authors: Milstein C. , Frangione B.
Published: August 2015

Amino acid sequences around the disulphide bridges of the heavy chain of an immunoglobulin of the γ2 subclass have been studied. The protein was digested with pepsin and the digest fractionated by Sephadex. Screening of the eluate by one-dimensional electrophoresis of oxidized and unoxidized samples was used as an assay and pools of fractions were prepared. Identification by diagonal electrophoresis of several inter- and intra-chain disulphide bridges was done on the pooled fractions. The inter-heavy-chain bridged peptide included four cystine residues. Comparison with proteins of other human subclasses indicated that the intrachain bridges identified are the bridges of the invariable section of γ2 heavy chains. The amino acid sequence of one cysteic acid peptide that may have been derived from the variable part of the molecule was determined. Partial reduction followed by carboxymethylation with radioactive iodoacetate of two proteins of the γ2 class showed a number of labelled peptides that could be identified as being related to the inter-chain bonded cystine residues.
Studies on the porphobilinogen deaminase–uroporphyrinogen cosynthetase system of cultured soya-bean cells
Authors: Llambías Elena B. C. , Batlle Alcira M. Del C.
Published: August 2015

1. Porphobilinogenase was isolated and purified from soya-bean callus tissue; its components, porphobilinogen deaminase and uroporphyrinogen isomerase, were separated and purified. 2. The purified porphobilinogenase was resolved into two bands on starch-gel electrophoresis. The molecular weights of porphobilinogenase, deaminase and isomerase fractions were determined by the gel-filtration method. Porphobilinogenase activity was affected by the presence of air; uroporphyrinogens were only formed under anaerobic conditions, although substrate consumption was the same in the absence of oxygen as in its presence. 3. pH-dependence of both porphobilinogenase and deaminase was the same and a sharp optimum at pH 7.2 was obtained. Isomerase was heat-labile, but the presence of ammonium ions or porphobilinogen afforded some protection against inactivation. The action of several compounds added to the system was studied. Cysteine, thioglycollate, ammonium ions and hydroxylamine inhibited porphobilinogenase; certain concentrations of sodium and magnesium salts enhanced activity; some dicarboxylic acids and 2-methoxy-5-nitrotropone inhibited the deaminase. 4. δ-Aminolaevulate and ethionine in the culture media stimulated porphyrin synthesis and increased porphobilinogenase activity, whereas iron deficiency resulted in porphyrin accumulation. 5. The development of chlorophyll and porphobilinogenase on illumination of dark-grown callus was followed. 6. A hypothetical scheme is suggested for the enzymic synthesis of uroporphyrinogens from porphobilinogen.
Nucleic acid enzymology of extremely halophilic bacteria. Halobacterium cutirubrum polynucleotide phosphorylase
Authors: Peterkin Pearl I. , Fitt P. S.
Published: August 2015

1. Polynucleotide phosphorylase was purified 200-fold from Halobacterium cutirubrum. 2. It is membrane-associated and can be solubilized by sonication. 3. The purified enzyme requires a high ionic strength for both stability and activity. 4. It is Mn2+-dependent, has all three typical polynucleotide phosphorylase activities and is specific for nucleoside diphosphates. 5. The enzyme is of low molecular weight.
Nucleic acid enzymology of extremely halophilic bacteria. Halobacterium cutirubrum deoxyribonucleic acid-dependent ribonucleic acid polymerase
Authors: Louis B. Gregory , Fitt P. S.
Published: August 2015

1. DNA-dependent RNA polymerase was purified 150-fold from crude extracts of the extreme halophile Halobacterium cutirubrum. 2. The enzyme requires the presence of native DNA and all four nucleoside triphosphates to incorporate 14C-labelled nucleoside triphosphate into an acid-insoluble ribonuclease-sensitive product. 3. It has an absolute requirement for both Mn2+ and Mg2+. 4. The polymerase requires a high salt concentration for stability, but is markedly inhibited by univalent cations. 5. Its molecular weight is very low compared with that of Escherichia coli RNA polymerase.
The activities of phosphorylase, hexokinase, phosphofructokinase, lactate dehydrogenase and the glycerol 3-phosphate dehydrogenase in muscles from vertebrates and invertebrates
Authors: Crabtree B. , Newsholme E. A.
Published: August 2015

1. The maximum activities of hexokinase, phosphorylase and phosphofructokinase have been measured in extracts from a variety of muscles and they have been used to estimate the maximum rates of operation of glycolysis in muscle. These estimated rates of glycolysis are compared with those calculated for the intact muscle from such information as oxygen uptake, glycogen degradation and lactate formation. Reasonable agreement between these determinations is observed, and this suggests that such enzyme activity measurements may provide a useful method for comparative investigations into quantitative aspects of maximum glycolytic flux in muscle. 2. The enzyme activities from insect flight muscle confirm and extend much of the earlier work and indicate the type of fuel that can support insect flight. The maximum activity of hexokinase in some insect flight muscles is about tenfold higher than that in vertebrate muscles. The activity of phosphorylase is greater, in general, in vertebrate muscle (particularly white muscle) than in insect flight muscle. This is probably related to the role of glycogen breakdown in vertebrate muscle (particularly white muscle) for the provision of ATP from anaerobic glycolysis and not from complete oxidation of the glucose residues. The activity of hexokinase was found to be higher in red than in white vertebrate muscle, thus confirming and extending earlier reports. 3. The maximum activity of the mitochondrial glycerophosphate dehydrogenase was always much lower than that of the cytoplasmic enzyme, indicating that the former enzyme is rate-limiting for the glycerol 3-phosphate cycle. From the maximum activity of the mitochondrial enzyme it can be calculated that the operation of this cycle would account for the reoxidation of all the glycolytically produced NADH in insect flight muscle but it could account for only a small amount in vertebrate muscle. Other mechanisms for this NADH reoxidation in vertebrate muscle are discussed briefly.
Oxidation of the capsular polysaccharide of pneumoccal type IX by periodate
Authors: Das A , Higginbotham J. D. , Heidelberger M
Published: August 2015

1. The pneumococcal type IX polysaccharide (polysaccharide S IX) has been oxidized by sodium metaperiodate and reduced by sodium borohydride. Of the constituent monosaccharides, N-acetylglucosamine and N-acetylmannosamine remain unaltered, whereas 40% of the glucose and 90% of the glucuronic acid are oxidized. 2. The effect of oxidation and subsequent reduction on the precipitation of polysaccharide S IX in anti-(pneumococcal) sera is described and interpreted in structural terms. 3. Oligosaccharides produced by oxidation, reduction and hydrolysis with dilute acid have been isolated and partially characterized. 4. The results in this paper and the preceding one (Higginbotham et al., 1972) are used to postulate a possible structure for polysaccharide S IX.
Choline kinase in Cuscuta reflexa
Authors: Setty Pravina N. , Krishnan P. S.
Published: August 2015

1. Choline kinase is a mitochondrial enzyme in Cuscuta reflexa. It can be solubilized from the particles by treatment with 350mm-sodium chloride, or by freezing and thawing. 2. Choline kinase of C. reflexa was purified by starting from the crude mitochondrial fraction. A 33–52% recovery of the enzyme, on the basis of the activity in the original homogenate, in 1200–2250-fold enrichment, was effected. 3. The purified preparation of choline kinase had a sigmoid saturation curve with respect to choline, with a Hill number of 2.3, and was inhibited by ADP (competitive in nature and allosteric in binding, with a Hill number of 2.7) and by phosphorylcholine (non-competitive and non-allosteric). The kinetic characteristics of the enzyme were consistent with the K type allosteric model of Monod et al. (1965). 4. The enzyme was desensitized, with respect to choline regulation, by prolonged storage in the cold, was activated significantly on warming before assay and was inactivated by high concentrations of sodium chloride. 5. The significance of allostery in choline kinase in relation to the intracellular regulation of phospholipid synthesis is discussed.
Molecular characteristics of chicken liver arginase
Authors: Grazi E. , Magri E.
Published: August 2015

A purification procedure for the preparation of chicken liver arginase in a homogeneous form is presented. The enzyme hydrolyses both arginine and argininic acid. Kinetic analysis reveals that the enzyme binds arginine when the amino group is protonated or unprotonated; however, the unprotonated form seems to be hydrolysed more rapidly. The enzyme exchanges with the medium approx. 1.6Mn2+ions per molecule.
Escherichia coli alkaline phosphatase. Relaxation spectra of ligand binding
Authors: Halford S. E.
Published: August 2015

The temperature-jump technique was used to study the binding equilibrium between the Escherichia coli alkaline phosphatase dimer and 2-hydroxy-5-nitrobenzyl phosphonate in 0.1m-tris buffer, pH8.0. Three partially discrete relaxations were observed, two of which could be related to the bimolecular associations of ligand with different conformations of the enzyme and the third to the interconversion of these states. Relaxation spectra were also used to analyse the changes in the mechanism of ligand binding to alkaline phosphatase caused by increase in ionic strength. The relaxation spectrum observed after the addition of Pi to the equilibrium mixture of phosphonate and enzyme was also studied. Difference spectroscopy indicated that both of these ligands were bound to the alkaline phosphatase dimer at the same time. These results are related to the catalytic mechanism of this enzyme, with particular reference to the role of two identical subunits in a dimeric enzyme that exhibits only one active site functioning in catalysis at any given time.
The regulation of rat liver xanthine oxidase. Involvement of thiol groups in the conversion of the enzyme activity from dehydrogenase (type D) into oxidase (type O) and purification of the enzyme
Authors: Corte E D , Stirpe F.
Published: August 2015

1. The ‘xanthine oxidase’ activity of rat liver supernatant, most of which behaves as an NAD+-dependent dehydrogenase (type D) can be rapidly converted into an oxidase (type O) by thiol reagents such as tetraethylthiuram disulphide, copper sulphate, 5,5′-dithiobis-(2-nitrobenzoic acid), N-ethylmaleimide and p-hydroxymercuribenzoate. Treatment with copper sulphate, if prolonged, leads to almost complete inactivation of the enzyme. The effect of these reagents is prevented by dithioerythritol, and in all cases but that of N-ethylmaleimide is reversed by the same thiol. 2. Dithioerythritol prevents and reverses the conversion of xanthine oxidase from type D into type O brought about by storage of rat liver supernatant at -20°C, preincubation under anaerobic conditions, treatment with carbon or with diethyl ether, and reverses, but does not prevent, the conversion obtained by preincubation of the whole liver homogenate. 3. Conversion of the enzyme from type D into type O is effected by preincubation of rat liver supernatant with the sedimentable fraction from rat liver but not from chick or pigeon liver. The xanthine dehydrogenase activity of chick liver supernatant is not changed into an oxidase by preincubation with the sedimentable fraction from rat liver. 4. The enzyme activity of rat liver supernatant is converted from type D into type O during purification of the enzyme: the purified enzyme can be reconverted into type D by dithioerythritol. 5. The enzyme appears as an oxidase in the supernatant of rat heart, intestine, spleen, pancreas, lung and kidney. The enzyme of all organs but intestine can be converted into a dehydrogenase by dithioerythritol.
The formation of sulphur-containing amino acids in germinating seeds of rape (Brassica napus L.)
Authors: Ngo T. T. , Shargool P. D.
Published: August 2015

Sodium [35S]sulphide was fed to batches of germinating rapeseed, in some instances with the addition of unlabelled cysteine. Both the total radioactivity and specific radioactivity of the free sulphur-containing amino acids were examined. Cysteine and homocysteine were rapidly labelled; label subsequently appeared in cystathionine and methionine. The results obtained indicated that both the sulphydration and trans-sulphuration pathways were operating. This conclusion was reinforced by the results of experiments in which batches of rapeseed were incubated with l-[14C]homoserine. These showed the formation of labelled homocysteine, cystathione and methionine. It was thought the trans-sulphuration pathway was making the greater contribution to the biosynthesis of methionine in germinating rapeseed.
Isolation of human synovial-fluid hyaluronate by density-gradient ultracentrifugation and evaluation of its protein content
Authors: Scher Irwin , Hamerman David
Published: August 2015

1. A compound of hyaluronate and protein, called hyaluronate–protein was isolated from pooled human synovial fluids by caesium chloride density-gradient ultracentrifugation. 2. The isolated hyaluronate–protein was labelled with [125I]iodide and the following studies were done. (a) Ultracentrifugation in caesium chloride showed that the protein moiety (125I counts) and hyaluronate (hexuronate) sedimented together in the middle of the gradient. (b) The labelled hyaluronate–protein was treated with trypsin, and ultracentrifugation showed that peptide fragments (125I counts) were dispersed throughout the gradient, indicating proteolytic digestion. Hyaluronate sedimented in the middle of the gradient. (c) The labelled hyaluronate–protein was digested with streptococcal hyaluronidase, and ultracentrifugation showed that hyaluronate fragments were dispersed throughout the gradient, indicating digestion of the polysaccharide. The protein moiety, without attached hyaluronate, now sedimented at the top of the gradient. (d) Ultracentrifugation of labelled hyaluronate–protein in 4m-guanidinium chloride showed that protein and hyaluronate sedimented together. 3. These studies confirm that hyaluronate is combined with a small quantity of protein in normal human synovial fluid. A mild method for the rapid isolation of hyaluronate–protein in good yield is described.
Methylation of ribonucleic acid by the carcinogens dimethyl sulphate, N-methyl-N-nitrosourea and N-methyl-N′-nitro-N-nitrosoguanidine. Comparisons of chemical analyses at the nucleoside and base levels
Authors: Lawley P. D. , Shah S. A.
Published: August 2015

1. The following methods for hydrolysis of methyl-14C-labelled RNA, and for chromatographic isolation and determination of the products, were investigated: enzymic digestion to nucleosides at pH6 or 8; alkaline hydrolysis and conversion into nucleosides; hydrolysis by acid to pyrimidine nucleotides and purine bases, or completely to bases; chromatography on Dowex 50 (NH4+form) at pH6 or 8.9, or on Dowex 50 (H+form), or on Sephadex G-10. 2. The suitability of the various methods for determination of methylation products was assessed. The principal product, 7-methylguanosine, was unstable under the conditions used for determinations of nucleosides. 3- and 7-Methyladenine and 3- and 7-methylguanine are best determined as bases; 1-methyladenine and 3-methylcytosine can be isolated as either nucleosides or bases; O6-methylguanine is unstable under the acid hydrolysis conditions used and can be determined as the nucleoside; 3-methyluracil was detected, but may be derived from methylation of the ionized form of uracil. 3. Differences between the patterns of methylation of RNA and homopolyribonucleotides by the N-methyl-N-nitroso compounds and dimethyl sulphate were found: the nitroso compounds were able to methylate O-6 of guanine, were relatively more reactive at N-7 of adenine and probably at N-3 of guanine, but less reactive at N-1 of adenine, N-3 of cytosine and probably at N-3 of uridine. They probably reacted more with the ribose–phosphate chain, but no products from this were identified. 4. The possible influences of these differences on biological action of the methylating agents is discussed. Nitroso compounds may differ principally in their ability to induce miscoding in the Watson–Crick sense by reaction at O-6 of guanine. Both types of agent may induce miscoding to a lesser extent through methylation at N-3 of guanine; both can methylate N atoms, presumably preventing Watson–Crick hydrogen-bonding. N-Methyl-N-nitrosourea can degrade RNA, possibly through phosphotriester formation, but this mechanism is not proven.
Self-renaturing fractions in the separated strands of mouse satellite deoxyribonucleic acid
Authors: Sutton W. D. , Walker P. M. B.
Published: August 2015

Pyrimidine- and purine-rich strands of Mus musculus satellite DNA prepared by alkaline CsCl-gradient centrifugation can self-renature to a variable extent to give partial duplexes with high thermal stability. These duplexes were purified by treatment with nuclease S1 followed by hydroxylapatite chromatography, and have been shown by pyrimidine-tract analysis to be very similar in sequence to total reassociated satellite DNA. It is believed that the self-renaturing fractions result from variable contamination of each strand with fragments of the other, rather than from molecular inversions. The predominantly single-stranded properties of these fractions may be partly due to the ability of mouse satellite DNA strands to reassociate in non-stoicheiometric proportions.
The amino acid sequence of cytochrome c from Helix aspersa Müller (garden snail)
Authors: Brown R. H. , Richardson M. , Boulter D. , Ramshaw J. A. M. , Jefferies R. P. S.
Published: August 2015

The amino acid sequence of a snail cytochrome c has been determined. The molecule consists of a single polypeptide chain of 104 residues, and is homologous with other mitochondrial cytochromes c. Unlike the cytochromes c from vertebrates, there is no acetyl blocking group at the N-terminus. A change in an otherwise invariant position has been observed in position 87. Comparison with amino acid sequences of cytochromes c from other sources indicates that the point of divergence of the molluscs and the vertebrates in evolutionary time was 720 million years ago. Experimental details are given in a supplementary paper that has been deposited as Supplementary Publication SUP 50009 at the National Lending Library for Science and Technology, Boston Spa, Yorks. LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1972), 126, 5.
Effects of starvation on intermediary metabolism in the lactating cow. A comparison with metabolic changes occurring during bovine ketosis
Authors: Baird G. D. , Heitzman R. J. , Hibbitt K. G.
Published: August 2015

1. The purpose of this study was to determine the nature of the metabolic changes associated with carbohydrate and fat metabolism that occurred in the blood and liver of lactating dairy cows during starvation for 6 days. 2. During starvation, the blood concentrations of the free fatty acids and ketone bodies increased, whereas that of citrate decreased. After an initial increase, the blood concentration of glucose subsequently declined as starvation progressed. Starvation caused a significant decrease in the plasma concentration of serine and a significant increase in that of leucine. 3. After 6 days of starvation the hepatic concentrations of oxaloacetate, citrate, phosphoenolpyruvate, 2-phosphoglycerate, 3-phosphoglycerate, glucose, glycogen, ATP and NAD+ had all decreased, as had the hepatic activities of phosphopyruvate carboxylase (EC 4.1.1.32) and pyruvate kinase (EC 2.7.1.40). 4. The above metabolic changes are similar to those previously found to occur in cows suffering from spontaneous ketosis (Baird et al., 1968; Baird & Heitzman, 1971). 5. Milk yield decreased progressively during starvation. 6. There were marked differences in the ability of individual animals to resist the onset of severe starvation ketosis.
A study of stabilization of gluconeogenic activity in rat liver slices by calcium and manganese ions
Authors: Roobol Anne , Alleyne G. A. O.
Published: August 2015

1. The effect of some bivalent cations on gluconeogenesis by the rat liver-slice preparation has been investigated. 2. Ca2+and Mn2+stimulated glucose production from a range of substrates but not from glycerol. Mg2+had no effect on the rate of glucose production. 3. Ca2+were required to maintain phosphoenolpyruvate carboxylase activity in the slice preparation. 4. Ca2+and Mn2+, but not Mg2+, retarded the release of lysosomal enzymes from the slice into the incubation medium. 5. It is proposed that Ca2+and Mn2+stimulate glucose production by stabilizing the lysosome system in the liver-slice preparation. 6. The value of the liver-slice preparation as a means of measuring hepatic gluconeogenesis is discussed.
Thermal inactivation and reactivation of an enzyme in vivo. Pantothenate hydrolase of Pseudomonas fluorescens
Authors: Airas R. Kalervo
Published: August 2015

Thermal inactivation and reactivation of pantothenate hydrolase were studied in whole cells of Pseudomonas fluorescens. The enzyme is susceptible to thermal inactivation in whole cells at 37–40°C, and is reactivated when the temperature is lowered again. Chloramphenicol does not prevent reactivation. The activation energy of enzyme inactivation in vivo is about 540kJ/mol. This activation energy is 220kJ/mol in vitro, but it is increased to 550–630kJ/mol by several metabolites, such as succinate, glyoxylate and oxalate. Generally, good carbon sources, causing rapid growth, protect the enzyme from thermal inactivation in vivo, and enable reactivation to occur at a fast rate. The enzyme is also inactivated below 35°C, showing an activation energy of about 35kJ/mol. Good carbon sources prevent this inactivation as well, and cause slight reactivation. Glycine, although not utilized for growth, protects the enzyme well from this inactivation but not from inactivation at 37–40°C, and prevents reactivation totally. From the activation energies of inactivation and the effects of the various carbon sources, it appears possible that changes in the concentrations of intracellular metabolites may be responsible for the changes in inactivation and reactivation.
The identification of a lipid-mobilizing factor from sheep midbrain
Authors: Shah P. P. , Tee A. R. , English P. D. , Redman B. T. , Bunyan J.
Published: August 2015

The mass spectrum of a lipolytic substance from purified sheep midbrain extract indicated that it was isoprenaline, not N-n-propylnoradrenaline, noradrenaline or adrenaline. The identification of the lipid-mobilizing factor as isoprenaline was confirmed qualitatively by t.l.c. and g.l.c. and quantitatively by determination of its lipolytic activity and u.v. absorption and by fluorimetric determination of its catecholamine content.
Histone acetylation and hormone action. Early effects of oestradiol-17β on histone acetylation in rat uterus
Authors: Libby P. R.
Published: August 2015

The effect of oestradiol treatment on the acetylation of histones of the immature rat uterus has been studied. A 10μg dose of oestradiol causes a 70% increase at 5min and a 140% increase at 10min after administration in the labelling of the histone fraction F2+F3. No effect of oestradiol is seen on the labelling of histones F1 or acidic non-histone chromatin proteins. The oestradiol stimulation is seen in animals pretreated with either cycloheximide or actinomycin D. The stimulation of labelling caused by oestradiol is completely abolished by pretreatment of the animals with the anti-oestrogen, nafoxidine. The stimulation is given by lower doses of oestradiol, by stilboestrol and oestriol, but is not given by testosterone. These results suggest that stimulation of histone acetylation in the uterus is the earliest known effect of the hormone on its target tissue.
The activities of lipases and carnitine palmitoyl-transferase in muscles from vertebrates and invertebrates
Authors: Crabtree B. , Newsholme E. A.
Published: August 2015

1. The activities of tri-, di- and mono-glyceride lipase and carnitine palmitoyltransferase were measured in homogenates of a variety of muscles. These activities were used to estimate the rate of utilization of glycerides and fatty acids by muscle. In muscles whose estimated rates of fat utilization can be compared with rates calculated for the intact muscle from such information as O2 uptake, there is reasonable agreement between the estimated and calculated rates. 2. In all muscles investigated the maximum rates of hydrolysis of glycerides increase in the order triglyceride, diglyceride, monoglyceride. The activity of diglyceride lipase is highest in the flight muscles of insects such as the locust, waterbug and some moths and is lowest in the flight muscles of flies, bees and the wasp. These results are consistent with the utilization of diglyceride as a fuel for some insect flight muscles. 3. In many muscles from both vertebrates and invertebrates the activity of glycerol kinase is similar to that of lipase. It is concluded that in these muscles the metabolic role of glycerol kinase is the removal of glycerol produced during lipolysis. However, in some insect flight muscles the activity of glycerol kinase is much greater than that of lipase, which suggests a different role for glycerol kinase in these muscles.
Microbial metabolism of the pyridine ring. Formation of pyridinediols (dihydroxypyridines) as intermediates in the degradation of pyridine compounds by micro-organisms
Authors: Houghton C. , Cain R. B.
Published: August 2015

1. Several species of micro-organisms that were capable of utilizing pyridine compounds as carbon and energy source were isolated from soil and sewage. Compounds degraded included pyridine and the three isomeric hydroxypyridines. 2. Suitable modifications of the cultural conditions led to the accumulation of pyridinediols (dihydroxypyridines), which were isolated and characterized. 3. Three species of Achromobacter produced pyridine-2,5-diol from 2- or 3-hydroxypyridine whereas an uncommon Agrobacterium sp. (N.C.I.B. 10413) produced pyridine-3,4-diol from 4-hydroxypyridine. 4. On the basis of chemical isolation, induction of the necessary enzymes in washed suspensions and the substrate specificity exhibited by the isolated bacteria, the initial transformations proposed are: 2-hydroxypyridine → pyridine-2,5-diol; 3-hydroxypyridine → pyridine-2,5-diol and 4-hydroxypyridine → pyridine-3,4-diol. 5. A selected pyridine-utilizer, Nocardia Z1, did not produce any detectable hydroxy derivative from pyridine, but carried out a slow oxidation of 3-hydroxypyridine to pyridine-2,3-diol and pyridine-3,4-diol. These diols were not further metabolized. 6. Addition of the isomeric hydroxypyridines to a model hydroxylating system resulted in the formation of those diols predicted by theory.
The effect of hypophysectomy and subsequent replacement therapy with sheep prolactin or bovine growth hormone on the lactose synthetase activity of rabbit mammary gland
Authors: Jones E. A. , Cowie A. T.
Published: August 2015

1. The effects of hypophysectomy and replacement therapy with sheep prolactin and bovine growth hormone on the lactose synthetase activity of the mammary glands of lactating rabbits were studied. 2. There was an approximately fourfold decline in the lactose synthetase activity of homogenates calculated on a DNA basis within 6–7 days of hypophysectomy. Prolactin reversed this decline but growth hormone had no effect. 3. Changes in the properties of a particulate fraction isolated from the glands indicated that a decline in the effective concentration of α-lactalbumin was one factor contributing to the decreased lactose synthetase activity after hypophysectomy. 4. As the changes in lactose output produced by hypophysectomy and prolactin therapy are much greater than the changes in total lactose synthetase activity it is concluded that the activity of this enzyme is not the main factor controlling lactose output under these conditions.