bluuebunny/crossref_metadata_2025 huggingface dataset:
The effect of scurvy on hexosamine-containing substances in healing wounds in guinea pigs
Authors: Bates C. J. , Levene C. I. , Kodicek E.
Published: August 2015

1. Granulation tissue from healing tendonectomy wounds in guinea pigs was analysed and the effects of inanition and ascorbic acid deficiency on this tissue were investigated. 2. Inanition produced no significant effect on either the glucosamine or the galactosamine content of the tissue. Ascorbic acid deficiency decreased the galactosamine content without affecting the glucosamine content. 3. Fractionation of papain-digested granulation tissue gave three major fractions, which behaved respectively as glycopeptide, hyaluronic acid and a sulphated glycosaminoglycan mixture. At least half of the sulphated glycosaminoglycan mixture behaved as dermatan sulphate. 4. Inanition produced no consistent effect on the fractions examined. In ascorbic acid deficiency, a decrease in the sulphated glycosaminoglycan fraction was observed, which accounted for the decreased galactosamine content of the tissue. This was accompanied by a decrease in hyaluronic acid and a slight increase in the glycopeptide fraction.
The effect of genetic complexity on the time-course of ribonucleic acid–deoxyribonucleic acid hybridization
Authors: Bishop J O
Published: August 2015

1. The rate of RNA–DNA hybridization was studied under conditions of RNA excess, with RNA synthesized in vitro. The initial rate of the reaction was proportional to the initial RNA concentration. Throughout the observed course of the reaction there was a linear relationship between the reciprocal of the amount of RNA hybridized/μg. of DNA and the reciprocal of time. The slope of the reciprocal plot was inversely proportional to the initial RNA concentration. 2. A comparison was made of the hybridization of DNA from Escherichia coli and from bacteriophages T4 and λ with homologous RNA. The initial rate of hybridization was inversely proportional to the genetic complexity of the hybridizing system. The slope of the reciprocal-time plot was directly proportional to genetic complexity. These results are interpreted to indicate that the rate of hybridization reflects the mean concentration of the various unique RNA species in a preparation.
The effects of calcium and magnesium ions on the adenosine triphosphatase and inosine triphosphatase activities of myosin A
Authors: Sugden E. A. , Nihei T.
Published: August 2015

1. The effects of Ca2+ and Mg2+ on the enzymic activity of myosin were studied with myosin preparations treated by the ion-exchange resin Chelex-100. A reaction mixture containing 0·05m-potassium chloride was chosen in which the effects of univalent ions such as K+, Na+ and Cl− do not change significantly with small variations in their concentrations. 2. The relationship between the rate of hydrolysis of ATP or ITP and the concentration of Ca2+ suggests that a relatively weak binding of Ca2+ either to myosin or to the substrate nucleotide is responsible for the activation of the enzymic activity. According to the experiments with an ultrafiltration technique, the binding of Ca2+ to myosin proceeds in at least two steps, the first occurring at one site on every 500000 atomic mass units of myosin with an apparent association constant, Kapp., 1·3×106m−1, and the second seeming to be so weak that its binding parameters cannot be determined by the method used. The first type of Ca2+ binding is not observable with N-ethylmaleimide-modified myosin, yet this modified myosin shows activation by Ca2+ of its adenosine triphosphatase and inosine triphosphatase. 3. The inhibition by Mg2+ can be related to a binding reaction of Mg2+ with myosin having Kapp. ∼106m−1. Mg2+ replaces the Ca2+ bound tightly to myosin. The Kapp. for Mg2+–myosin binding calculated by assuming a competition between Ca2+ and Mg2+ for the same site is 2·1×105−3·0×105m−1. When myosin is modified with a thiol reagent (p-mercuribenzoate) at a certain ratio to myosin, the inhibition by Mg2+ becomes unobservable. 4. The behaviour of the hydrolytic activity of myosin on ATP or ITP in the presence of both Ca2+ and Mg2+ is consistent with the explanation that the inhibition by Mg2+ is due to the tight binding of Mg2+ to myosin, whereas the activation by Ca2+ is caused either by a weak binding of Ca2+ to myosin or by CaATP2− or by both.
Studies on protein–polysaccharides from pig laryngeal cartilage. Heterogeneity, fractionation and characterization
Authors: Tsiganos C. P. , Muir Helen
Published: August 2015

1. Protein–polysaccharides from pig laryngeal cartilage extracted by two procedures described in the preceding paper (Tsiganos & Muir, 1969) were shown to consist of macromolecules of various sizes as assessed by gel filtration in 4% and 6% agarose. 2. A larger proportion of the smaller molecules was present in the preparation obtained by brief extraction in iso-osmotic sodium acetate (procedure I) than in that obtained by more prolonged extraction in 10% (w/v) calcium chloride (procedure II). 3. Two fractions were separated by gel filtration in 6% agarose and by electrophoresis in compressed glass fibre. These fractions differed in chemical composition and in antigenic determinants. The gel-retarded fraction R and that of higher electrophoretic mobility possessed the same single antigen, whereas the gel-excluded fraction E and the slower electrophoretic fraction contained all the antigens of the starting material including that of fraction R. 4. Five N-terminal amino acid residues were identified in preparation I and fraction E, only two of which were present in fraction R. 5. The relative proportions of gel-excluded and gel-retarded fractions did not change when solutions of high ionic strength, urea or guanidine hydrochloride were used for elution. 6. The differences in chemical and amino acid composition between fractions R and E showed that the latter was not a simple aggregate of the former. Fraction E contained more basic and aromatic amino acids, and some methionine and cystine; the last two were absent from fraction R. Hydroxyproline was not detected in either fraction. 7. The number of glycosidic linkages in both fractions was estimated by alkaline β-elimination. Appreciable amounts of threonine as well as serine were destroyed in both fractions. An average chain length for chondroitin sulphate was calculated from the galactosamine content of both fractions and the amounts of hydroxy amino acid destroyed. Average chain lengths were also calculated from the xylose and galactosamine content of each fraction. Each independent method gave a value of approximately 28 disaccharide units for the chain length in both fractions and hence their difference in size could not be explained by differences in the length of carbohydrate chains. 8. All fractions contained glucosamine, which was attributed to keratan sulphate. Content of both protein and keratan sulphate increased with the size of the macromolecules. 9. It is suggested, from these results, that chondroitin sulphate–protein complexes normally exist as a heterogeneous population of macromolecules in cartilage, and that keratan sulphate is involved in the formation of larger molecules.
The binding of cyanide to ferroperoxidase
Authors: Phelps Charles , Antonini Eraldo , Brunori Maurizio
Published: August 2015

1. The equilibrium and kinetics of cyanide binding to ferroperoxidase were investigated. At pH9.1 the equilibrium and kinetic measurements agree closely and disclose a single process with an affinity constant of 1.1×103m@!-1 and combination and dissociation velocity constants of 29m-1·s-1 and 2.5×10-2s-1 respectively. 2. At pH values below 8 the affinity constant falls until at pH6.0 the ferroperoxidase·cyanide complex is no longer formed. This is shown to be associated with the formation of ferriperoxidase·cyanide complex in the mixture even in the presence of excess of sodium dithionite. 3. Rapid-pH-jump experiments show a fast pseudo-first-order interconversion between ferroperoxidase·cyanide complex at pH9.1 and ferriperoxidase·cyanide complex at pH6.0. 4. The kinetics of binding of cyanide to dithionite-reduced peroxidase at pH6.0 are complicated and radically different from those observed at pH9.1. 5. Above pH8 the change of affinity constant with pH is consistent with the undissociated species, HCN, being bound by the ferroperoxidase. The enthalpy for this process measured both by equilibrium and kinetic methods is about -8kcal/mol. 6. The binding of cyanide to reconstituted peroxidases, proto, meso and deutero, was investigated. 7. The results are discussed in relation to known data on cyanide binding to other haemoproteins.
Isoelectric focusing of proteins in the native and denatured states. Anomalous behaviour of plasma albumin
Authors: Salaman M. R. , Williamson A. R.
Published: August 2015

1. An analytical technique of isoelectric focusing in thin layers of polyacrylamide gel has been used to determine the isoelectric point, pI, of several proteins in the presence and in the absence of concentrated urea. 2. The presence of urea did not greatly affect pI except for bovine plasma albumin, where an increase of approx. 1pH unit was found. 3. Evidence is presented that this change in the pI of bovine plasma albumin is due to the normalization of certain ionizable groups on unfolding of the protein in urea. 4. Evidence is also presented that prolonged exposure of bovine plasma albumin to urea results in intramolecular disulphide interchange and that, on removal of urea, the new patterns of disulphide bonding stabilize abnormal conformations with pI values intermediate between those of the native and denatured states. 5. The studies demonstrate heterogeneity in bovine plasma albumin based on primary-sequence differences. 6. Isoelectric focusing of proteins in urea appears to be useful in the study of various aspects of protein structure.
Some features of the bacterial membrane studied with the aid of a new fractionation technique
Authors: Daniels M. J.
Published: August 2015

Membranes of Bacillus megaterium and other bacteria were bound to crystals of cadmium lauroylsarcosinate and were resolved into nine lipoprotein fractions by elution with potassium chloride and sodium deoxycholate solutions. The fractions differed widely in protein/lipid ratio. Some major protein species were probably common to all the fractions, but the phospholipid composition showed some variation. Electron-microscopic examination after negative staining revealed that material in certain fractions was in the form of particles of diameter about 12nm, and other fractions consisted of amorphous aggregates. The composition of the membrane in terms of the nine fractions was influenced markedly by the conditions of culture of the bacteria. Radioactive-labelling experiments suggested that some fractions served as precursors of others, and one fraction appeared to contain the attachment points of the DNA.
Amino acid sequences around the cysteine residues of rabbit muscle triose phosphate isomerase
Authors: Miller Janet C. , Waley S. G.
Published: August 2015

1. The nature of the subunits in rabbit muscle triose phosphate isomerase has been investigated. 2. Amino acid analyses show that there are five cysteine residues and two methionine residues/subunit. 3. The amino acid sequences around the cysteine residues have been determined; these account for about 75 residues. 4. Cleavage at the methionine residues with cyanogen bromide gave three fragments. 5. These results show that the subunits correspond to polypeptide chains, containing about 230 amino acid residues. The chains in triose phosphate isomerase seem to be shorter than those of other glycolytic enzymes.
The inhibition of pepsin-catalysed reactions by structural and stereochemical product analogues
Authors: Kitson T. M. , Knowles J. R.
Published: August 2015

1. The inhibition of pepsin-catalysed hydrolysis of N-acetyl-l-phenylalanyl-l-phenylalanylglycine by products and product analogues was studied. 2. Inhibitors of the l-configuration give rise to linear non-competitive inhibition, whereas those of the d-configuration show linear competitive behaviour. 3. Non-competitive inhibition by the product N-acetyl-l-phenylalanine indicates an ordered release of products, which supports a common mechanism (involving an ‘amino-enzyme’) for pepsin-catalysed transpeptidation and hydrolysis reactions. 4. The differences in the types of inhibition caused by product analogues of the l- and d-series emphasize the stereospecificity of the binding of these inhibitors to free enzyme and to the putative amino-enzyme intermediate. 5. The results suggest that it is the anion of the acyl product that is released first in the hydrolytic reaction (see Kitson & Knowles, 1971).
The pathway of pepsin-catalysed transpeptidation. Evidence for the reactive species being the anion of the acceptor molecule
Authors: Kitson T. M. , Knowles J. R.
Published: August 2015

1. The inhibition of pepsin-catalysed hydrolysis of N-acetyl-l-phenylalanyl-l-phenylalanylglycine by the acyl product and product analogues was studied at pH4.3. 2. The acyl product, N-acetyl-l-phenylalanine, gives rise to linear competitive inhibition at pH4.3, whereas at pH2.1 it shows linear non-competitive behaviour. 3. The extent of transpeptidation to N-acetyl-l-[3H]phenylalanine during the pepsin-catalysed hydrolysis of N-acetyl-l-phenylalanyl-l-phenylalanyl-glycine is significant at pH4.7, but is undetectable at pH1.3. 4. Both the inhibition and transpeptidation experiments are consistent with the anion of the acceptor molecule being the substrate in pepsin-catalysed transpeptidation. This conclusion supports the formulation of pepsin-catalysed reactions put forward by Knowles et al. (1970).
Carbon and sulphur utilization during growth of Pseudomonas fluorescens on potassium d-glucose 6-O-sulphate as the sole sulphur source
Authors: Fitzgerald J. W. , Dodgson K. S.
Published: August 2015

Pseudomonas fluorescens N.C.I.B. 8248, cultured on potassium d-glucose 6[35S]-O-sulphate as the sole sulphur source, liberated the 6-O-sulphate ester of d-gluconate into the culture medium. Extracts of bacteria grown under this cultural condition oxidized d-glucose 6-O-sulphate to yield the gluconate ester. Results suggest the involvement of a glucose dehydrogenase-like enzyme. The gluconate ester was apparently not oxidized further to any significant extent; however, it served as substrate for a desulphating enzyme found in extracts. Growth on d-glucose 6-O-sulphate as the sole source of sulphur was not associated with the appearance of a true glycosulphatase. Collectively, these results suggest that d-gluconate 6-O-sulphate, rather than the glucose ester, supplied the necessary sulphur for growth. Oxidative activities toward d-glucose 6-O-sulphate, d-glucose, d-gluconate 6-O-sulphate and d-gluconate found in extracts of P. fluorescens adapted to grow on d-glucose 6-O-sulphate as the sole source of carbon and sulphur are presented for comparative purposes.
Dihydroxyacetone phosphate. Its structure and reactivity with α-glycerophosphate dehydrogenase, aldolase and triose phosphate isomerase and some possible metabolic implications
Authors: Reynolds S. J. , Yates D. W. , Pogson C. I.
Published: August 2015

1. Dihydroxyacetone phosphate exists in neutral aqueous solution at 20°C as a mixture of keto, gem-diol and enolic forms in the ratio 55:44:1. 2. The three forms are freely interconvertible and rate constants for these reactions have been determined. 3. Keto-dihydroxyacetone phosphate is the primary reactive species in the reactions catalysed by α-glycerophosphate dehydrogenase, aldolase and triose phosphate isomerase. 4. The proportion of keto form to gem-diol forms of dihydroxyacetone phosphate is temperature-dependent. At 37°C, 83% is keto-dihydroxyacetone phosphate. 5. The enzymological and metabolic consequences of these results are discussed.
Membranes of chromaffin granules. Isolation and partial characterization of two proteins
Authors: Hörtnagl Heide , Winkler H. , Schöpf J. A. L. , Hohenwallner W.
Published: August 2015

Membranes of chromaffin granules were isolated from the adrenal glands of four different species. The solubilized membrane proteins could be resolved into several bands by polyacrylamide-gel electrophoresis (alkaline and acid gel systems). Two major protein components appeared to be common to the chromaffin granule membranes of ox, horse, pig and man. The various membrane proteins of bovine chromaffin granules were separated by filtration on Sephadex G-200 in the presence of sodium dodecyl sulphate. Two major membrane proteins (A and B) were obtained in purified form. Treatment of protein A with 2-mercaptoethanol before electrophoresis resulted in two more rapidly migrating subunits, whereas protein B was unaffected by mercaptoethanol treatment. The amino acid compositions of the two purified proteins were determined. They are very similar to that of the total membrane proteins but significantly different from that of the chromogranins, the soluble proteins of chromaffin granules.
An anaerobic reaction between lipoxygenase, linoleic acid and its hydroperoxides
Authors: Garssen G. J. , Vliegenthart J. F. G. , Boldingh J.
Published: August 2015

In an anaerobic system soya-bean lipoxygenase together with linoleic acid induces a structural rearrangement of 13-hydroperoxyoctadeca-cis-9-trans-11-dienoic acid leading to the formation of 13-oxotrideca-cis(trans)-9-trans-11-dienoic acid and n-pentane as well as 13-oxo-octadeca-9,11-dienoic acid. It is proposed that the 13-peroxyoctadeca-cis-9-trans-11-dienoic acid radical formed through hydrogen radical abstraction by the linoleic acid radical is the key intermediate for these reactions.
Thymidine kinase of rat liver. Activation by phospholipase C and subcellular distribution in the liver
Authors: Stirpe F. , La Placa M.
Published: August 2015

1. The thymidine kinase activity of rat liver is greatly enhanced on addition of phospholipase C to the assay mixture. 2. Most of the thymidine kinase activity of the liver is recovered in the mitochondrial and in the impure ‘nuclear’ fractions. No activity was detected in purified nuclei prepared in high-density sucrose. 3. A substantial thymidine kinase activity could be detected, with the aid of phospholipase C, in all rat tissues examined.
Studies on the structure-bound sedimentability of some rat liver lysosome hydrolases
Authors: Baccino F. M. , Rita G. A. , Zuretti Maria Franca
Published: August 2015

1. Lysosome-rich fractions from rat liver were subjected to several disruptive procedures: osmotic lysis or freezing and thawing in different media, shearing forces in a high-speed blender, treatment with Triton X-100. 2. The soluble and particulate phases were then separated by high-speed centrifugation and assayed for their content of acid phosphatase, β-galactosidase, β-N-acetylglucosaminidase, acid proteinase, acid ribonuclease, acid deoxyribonuclease and protein. 3. The degree of elution of these hydrolases appeared to depend on both the enzyme species and the treatment. The resulting patterns of solubilization were rather complex, so that a clear-cut discrimination between soluble and structure-bound enzymes could not always be traced. 4. Although only β-galactosidase was readily solubilizable after all treatments, acid proteinase could also be extensively eluted from the sedimentable material in the presence of EDTA and acid phosphatase was fully extracted by Triton X-100. On the other hand, considerable proportions of the other activities could not be solubilized by any of the procedures used. 5. In other experiments, the adsorbability of hydrolases on subcellular structures was investigated by measuring the partition between sedimentable particles and soluble fraction of solubilized enzymes added to ‘intact’ liver homogenates. 6. Large proportions of acid proteinase, ribonuclease and deoxyribonuclease, and almost all of β-N-acetylglucosaminidase, were found to be adsorbed on the particulate material.
Kinetic analysis of insulin action on amino acid uptake by isolated chick embryo heart cells
Authors: Guidotti G. G. , Borghetti A. F. , Lüneburg Britta , Gazzola G. C.
Published: August 2015

1. Isolated chick embryo heart cells were used to investigate the mode of action of insulin on the transport of three naturally occurring amino acids: l-proline, l-serine and glycine. Initial velocities of uptake were measured over a period of 5min with an 80-fold range of amino acid concentration. Corrections for amino acid diffusion, incorporation into protein and conversion into carbon dioxide were introduced. 2. The uptake processes approximated Michaelis–Menten kinetics within definite ranges of amino acid concentrations. A single transport system for proline and at least two transport systems for serine and glycine were detected. 3. The kinetic effects of insulin on transport systems for the amino acids tested were consistent with an acceleration of the maximal velocity of the process, without substantial changes in substrate concentration for half-maximal transport velocity. 4. These hormonal effects were not essentially altered by the corrections for amino acid incorporation into protein and conversion into carbon dioxide.
Phosphatidylcholine–cholesterol acyltransferase in the ultracentrifugal residual protein fraction of rat plasma
Authors: Sugano Michihiro
Published: August 2015

1. The phosphatidylcholine–cholesterol acyltransferase of rat plasma was dissociated from the plasma lipoproteins by ultracentrifugation and shown to be present in the plasma residual-protein fraction of density >1.210. 2. The general properties of the acyltransferase were substantially unchanged in the residual fraction as judged from the effects of differences in the substrates and of overnight starvation on the formation of different cholesterol esters. 3. The enzyme from rats starved overnight, by comparison with the enzyme from fed rats, preferentially formed cholesteryl arachidonate at the expense of cholesteryl linoleate. 4. The results suggest that ultracentrifugal separation of the plasma residual fraction may be used as an initial step for the purification of the acyltransferase.
Properties of fractionated chondroitin sulphate from ox nasal septa
Authors: Wasteson Åke
Published: August 2015

1. Chondroitin sulphate was isolated from bovine nasal septa by precipitation with cetylpyridinium chloride after digestion of the tissue with papain. 2. The material was divided into two portions, one of which was partially degraded with testicular hyaluronidase. 3. Untreated and hyaluronidase-digested material were fractionated into a total of eleven subfractions by gel chromatography on Sephadex G-200 and Sephadex G-100 respectively. 4. Chemical analyses indicated that the composition of all the fractions was similar to that of chondroitin sulphate. However, electrophoresis revealed a charge-inhomogeneity in the low-molecular-weight fractions obtained after hyaluronidase digestion. 5. The physicochemical properties of the subfractions were investigated by sedimentation-velocity, diffusion and sedimentation-equilibrium studies, osmometry, viscometry and gel chromatography. The individual fractions were essentially monodisperse and showed molecular weights ranging from 2400 to 36000. 6. The relationship between the intrinsic viscosity and the molecular weight was [η]=5.0×10−6×M1.14, indicating that the chondroitin sulphate molecules assume a shape intermediate between that of a random coil and a stiff rod. 7. The relationship between the sedimentation constant and the molecular weight (>104) was s020,w=2.3×10−2×M0.44.
Enzymic formation of p-hydroxybenzoate from p-hydroxycinnamate
Authors: Ranganathan S. , Ramasarma T.
Published: August 2015

An enzyme that converts p-hydroxycinnamate into p-hydroxybenzoate was found in rat liver. It is localized in the mitochondria and requires ATP. The activity is lost when the mitochondria are stored frozen overnight. Addition of magnesium chloride, cytochrome c, GSH, coenzyme A or potassium cyanide did not have any effect on the activity. When the rats were fed with α-p-chlorophenoxyisobutyrate, the rate of formation of p-hydroxybenzoate increased twofold. The reaction has some similar properties to fatty acid oxidation, but appears to be different in many respects.
Isolation and properties of a ‘malic’ enzyme from cauliflower bud mitochondria
Authors: Macrae A. R.
Published: August 2015

1. A ‘malic’ enzyme [l-malate–NAD oxidoreductase (decarboxylating), EC 1.1.1.39] has been isolated from cauliflower bud mitochondria and partially purified. 2. The enzyme is specific for l-malate and has an absolute requirement for either Mn2+, Co2+ or Mg2+. 3. The enzyme shows activity with both NAD+ and NADP+, but NAD+ is the preferred cofactor. 4. No appreciable oxaloacetate decarboxylase activity is present in the enzyme preparations even at low pH values. 5. The enzyme is inhibited by NADH and by oxaloacetate and stimulated by SO42− and by low concentrations of CoA. 6. The regulatory properties of the enzyme support the proposed role of the enzyme in the utilization of tricarboxylic acid-cycle acids for energy production when glycolysis is suppressed.
Metabolism of 4-chloro-2-methylphenoxyacetate by a soil pseudomonad. Preliminary evidence for the metabolic pathway
Authors: Gaunt J. K. , Evans W. C.
Published: August 2015

1. A pseudomonad capable of utilizing the herbicide 4-chloro-2-methylphenoxyacetate as a sole carbon source was isolated from soil and cultured in liquid medium. 2. Analysis of induction patterns of 4-chloro-2-methylphenoxyacetate-grown cells suggests that 5-chloro-o-cresol and 5-chloro-3-methylcatechol are early intermediates in the oxidation of 4-chloro-2-methylphenoxyacetate. Cells were not adapted to oxidize 4-chloro-6-hydroxy-2-methylphenoxyacetate. 3. In culture, 4-chloro-2-methylphenoxyacetate rapidly disappeared and the chlorine in the molecule was quantitatively released as Cl− ion. 4. A lactone (γ-carboxymethylene-α-methyl-Δαβ-butenolide) was isolated from cultures and established as an intermediate. 5. The following metabolic pathway is suggested: 4-chloro-2-methylphenoxyacetate → 5-chloro-o-cresol → 5-chloro-3-methylcatechol → cis–cis-γ-chloro-α-methylmuconate → γ-carboxymethylene-α-methyl-Δαβ-butenolide → γ-hydroxy-α-methylmuconate. 6. The tentative identification of 5-chloro-o-cresol, a γ-chloro-α-methylmuconate and γ-hydroxy-α-methylmuconate in culture extracts supports this scheme. However, the catechol was never observed to accumulate in cultures. 7. The detection of 4-chloro-6-hydroxy-2-methylphenoxyacetate, 2-methyl-phenoxyacetate, a dehalogenated cresol and oxalate in culture extracts is discussed in relation to the proposed metabolic pathway.
Synthesis of pyruvate carboxylase from its apoenzyme and (+)-biotin in Bacillus stearothermophilus. Purification and properties of the apoenzyme and the holoenzyme synthetase
Authors: Cazzulo J. J. , Sundaram T. K. , Dilks Susan N. , Kornberg H. L.
Published: August 2015

1. Methods are described for the assay and purification of pyruvate apocarboxylase and pyruvate holocarboxylase synthetase from biotin-deficient Bacillus stearothermophilus. 2. Pyruvate apocarboxylase was obtained 200-fold purified and in a nearly homogeneous state; it closely resembled the holoenzyme of the thermophile in fractionation properties, electrophoretic mobility and molecular weight (estimated to be 350000 by gel filtration). 3. Pyruvate holocarboxylase synthetase, purified more than 50-fold, was estimated to have a molecular weight of approx. 40000. 4. The conversion of the purified apoenzyme into the holoenzyme required the presence of the synthetase, ATP (Km3.3×10−7m), (+)-biotin (Km7.5×10−8m) and Mg2+; it differed from the conversions effected by systems forming other carboxylases in mesophilic organisms in also requiring the presence of acetyl-CoA.
Electrophoretic and immunological charactorization of rat neurophysin
Authors: Norström A. , Sjöstrand J. , Livett B. G. , Uttenthal L. O. , Hope D. B.
Published: August 2015

1. The electrophoretic properties of rat posterior pituitary proteins have been compared on starch gel with those of bovine and porcine neurophysins. 2. [35S]-Cysteine was injected into the supraoptic nucleus of male rats and 16–24h later the distribution of labelled neural-lobe protein in starch and polyacrylamide gels was determined. In both systems a single major protein component was found to contain more than 80% of the total recovered radioactivity. Between 5 and 10% of the radioactivity was found in a minor component in polyacrylamide gel. 3. In agar, microimmuno-diffusion and -electrophoresis of the rat neural-lobe proteins gave a single arc with neurophysin antiserum, and after starch-gel electrophoresis this arc was shown to be due to the major labelled component. 4. The molecular weights of the rat neural-lobe proteins were estimated by polyacrylamide-gel electrophoresis in the presence of sodium dodecyl sulphate. The molecular weight of the major labelled component was found to be 12000. 5. It is concluded that the rat neurophysin consists of one major and possibly one minor component.
The metabolism of phosphatidylinositol in the thyroid gland of the pig
Authors: Jungalwala F. B. , Freinkel N. , Dawson R. M. C.
Published: August 2015

1. The metabolism of phosphatidylinositol in pig thyroid has been investigated as a basis for understanding the specific stimulation of the synthesis of this phospholipid in the gland by thyrotropin. 2. The gland contained an active Ca2+-dependent phosphatidylinositol-splitting enzyme with an optimum pH of 5.3–5.5. 3. The major water-soluble product (65%) formed by this catabolic enzyme was not phosphorylinositol but a related compound, which may be a cyclic phosphorylinositol. Both this and phosphorylinositol (35%) were released simultaneously from the phosphatidylinositol substrate. 4. The phosphatidylinositol-splitting enzyme was found almost exclusively in the supernatant fraction obtained by homogenization of the gland. It was not present in the acid-phosphatase-containing particulate fraction. 5. The incorporation of [2-3H1]inositol into phosphatidylinositol in the presence of either CDP-diglyceride or CTP+ATP was most active in the microsomal fraction. 6. When thyroidal microsomes were labelled with [3H]inositol and 32P, and then incubated with unlabelled inositol, there was a dramatic loss of 3H labelling from the phosphatidylinositol, which was not accompanied by an equivalent loss of 32P from the phosphate moiety. This turnover of the inositol moiety required nucleotide coenzymes. It is postulated that the phosphatidylinositol is split into inositol and a phosphorus-containing lipid precursor of the phospholipid that remains on the microsomal membrane and is recycled. 7. Isolated thyroidal mitochondria synthesized phosphatidylinositol from [2-3H1]inositol only because of their contaminating microsomal component. 8. Some evidence has been obtained of a rapid transfer of phosphatidylinositol molecules from thyroidal microsomes to mitochondria when these were incubated together in the presence of a supernatant fraction. 9. Both phosphatidylinositol breakdown by the supernatant fraction of the gland and synthesis by the microsomes were totally inhibited by 1mm-chlorpromazine. This drug is known to suppress thyrotrophin-induced stimulation of activity in thyroid slices.
Phosphoenolpyruvate carboxykinase of kidney. Subcellular distribution and response to acid–base changes
Authors: Flores H. , Alleyne G. A. O.
Published: August 2015

1. A method for the assay of phosphoenolpyruvate carboxykinase is presented, based on the enzymic determination of the phosphoenolpyruvate produced by the enzyme reaction. 2. The subcellular distribution of phosphoenolpyruvate carboxykinase in the kidney of several animal species resembled the distribution in the liver. 3. The rise in enzyme activity in the kidney cortex of rats made acidotic by feeding with ammonium chloride was not prevented by administration of ethionine or actinomycin. 4. The possibility is suggested that in the kidney acidosis causes activation of an inactive form of the enzyme already present.
Amino acid sequence of the encephalitogenic basic protein from human myelin
Authors: Carnegie P. R.
Published: August 2015

Myelin from the central nervous system contains an unusual basic protein, which can induce experimental autoimmune encephalomyelitis. The basic protein from human brain was digested with trypsin and other enzymes and the sequence of the 170 amino acids was determined. The localization of the encephalitogenic determinants was described. Possible roles for the protein in the structure and function of myelin are discussed.
Metabolism of rat brain mitochondria. Studies on the potassium ion-stimulated oxidation of pyruvate
Authors: Nicklas W. J. , Clark J. B. , Williamson J. R.
Published: August 2015

1. Rat brain-cortex mitochondria were incubated in media containing 1, 5 or 100mm-K+ in the presence of ADP, uncoupler (FCCP, carbonyl cyanide p-trifluoro-methoxyphenylhydrazone) or valinomycin while metabolizing pyruvate and malate, or acetylcarnitine and malate or glutamate and malate as substrates. Both the uptake of oxygen and disappearance of substrate were measured under these conditions. 2. With pyruvate and malate as substrate in the presence of both ADP and valinomycin, both the uptake of oxygen and disappearance of pyruvate increased markedly on increasing the K+ content of the incubation medium from 5 to 100mm-K+. However, in the presence of uncoupler (FCCP), although the oxygen uptake doubled little change was observed in the rate of disappearance of pyruvate on increasing the K+ concentration. 3. Only small changes in uptake of substrate and oxygen were observed in the presence of ADP, uncoupler (FCCP) or valinomycin on increasing the K+ concentration when acetylcarnitine+malate or glutamate+malate were used as substrates by brain mitochondria. 4. Further, increasing the K+ concentration from 1 to 20mm when rat brain mitochondria were oxidizing a mixture of pyruvate and glutamate in the presence of malate and ADP caused a 30% increase in the respiration rate, 50% increase in the rate of disappearance of pyruvate and an 80% decrease in the rate of disappearance of glutamate. 5. Investigation of the redox state of the cytochromes and the nicotinamide nucleotides in various conditions with either pyruvate or acetylcarnitine as substrates suggested that the specific stimulation of metabolism of pyruvate by K+ could not be explained by a general stimulation of the electron-transport system. 6. Low-amplitude high-energy swelling of rat brain mitochondria was investigated in both Na+- and K+-containing media. Swelling of brain mitochondria was much greater in the Na+-containing medium and in this medium, the addition of Mg2+ caused a partial reversal of swelling together with an 85% decrease in the rate of utilization of pyruvate. However, in the K+-containing medium, the addition of Mg2+, although also causing a reversal of swelling, did not affect the rate of disappearance of pyruvate. 7. Measurements of the ATP, NADH/NAD+ and acetyl-CoA/CoA contents were made under various conditions and no evidence that K+ concentrations affected these parameters was obtained. 8. The results are discussed in relationship to the physiological significance of the stimulation of pyruvate metabolism by K+ in rat brain mitochondria. It is proposed that K+ causes its effects by a direct stimulation of the pyruvate dehydrogenase complex.
Sporulation in Bacillus subtilis 168. Comparison of alkaline phosphatase from sporulating and vegetative cells
Authors: Glenn A. R. , Mandelstam J.
Published: August 2015

1. The purification of the `vegetative' alkaline phosphatase of Bacillus subtilis 168 was simplified by ionic elution of the enzyme from intact cells. 2. The enzyme has a molecular weight of about 70000 and treatment of the enzyme with 10mm-hydrochloric acid or 6.0m-guanidine hydrochloride, β-mercaptoethanol (0.1m) gives rise to enzymically inactive subunits. 3. The amino acid composition of the enzyme was determined. The N-terminal residue determined by the DNS chloride method is glycine. 4. The properties of this enzyme were compared with the `sporulation' alkaline phosphatase of the same strain. 5. Although the `sporulation' enzyme differs from the `vegetative' enzyme in its physiology of appearance and apparent mRNA stability, an examination of properties of the enzymes revealed no differences. 6. The enzyme from both cell forms is bound to the particulate fraction of cell extracts, but can be solubilized by high concentrations of magnesium chloride; removal of the magnesium chloride, by dialysis, results in precipitation of both enzymes. Both enzymes can be removed from intact cells by ionic elution. 7. The `vegetative' and `sporulation' enzymes have identical pH optima, Km and Ki values and electrophoretic mobilities in cellulose acetate. 8. Their half-life is 28min at 65°C and their Q10 is 1.25. 9. The molecular size determined by gel filtration on Sephadex G-100 is about 69000. 10. `Vegetative' and `sporulation' forms gave precipitin lines that were continuous and non-spurred when tested against antiserum prepared against the `vegetative' enzyme. 11. The `sporulation' alkaline phosphatase appears to be associated with stage II of sporulation and appears to be induced by something specifically concerned in sporulation and not by phosphate starvation.
Protease and ribonuclease activities in bovine pituitary lobes
Authors: Pickup J. C. , Hope D. B.
Published: August 2015

1. Acid and alkaline protease activities in bovine anterior and posterior pituitary lobes were reinvestigated by measurement of u.v. and Folin–Ciocalteu colour values of trichloroacetic acid-soluble digestion products of denatured haemoglobin. 2. Both lobes of the pituitary gland contain a cathepsin with a pH optimum at 3.8. 3. When release of u.v.-absorbing material was used as the assay there was also an optimum at pH8.3–9.7, but this proved to be due to the release of nucleosides from an endogenous substrate. 4. The presence of a `cyclizing' ribonuclease active at alkaline pH on endogenous RNA was confirmed by the inhibitory effects of phosphate, arsenate and bentonite. The activity was unaffected by heat, EDTA or metal ions. The enzyme also acted on exogenous RNA. 5. A purified preparation of neurosecretory granules from fresh bovine posterior pituitary lobes was free from alkaline ribonuclease activity. Most of the activity present in the tissue was recovered in the supernatant plus microsomal material. 6. The distribution of RNA did not follow that of the alkaline ribonuclease.
The number and location of genes for 5S ribonucleic acid within the genome of Drosophila melanogaster
Authors: Quincey R. V.
Published: August 2015

DNA was prepared from wild-type and two mutant stocks of Drosophila melanogaster that differed in their dosage of the nucleolar organizer region. The relative amounts of DNA from the nucleolar organizer region in these preparations of DNA were determined by hybridization with 3H-labelled 28S rRNA. As expected, the amount of 3H-labelled 28S rRNA that hybridized was directly related to the dosage of nucleolar organizer region. No positive correlation was observed between the amount of 3H-labelled 5S RNA that hybridized and the dosage of nucleolar organizer region. Thus genes for 5S RNA are located primarily, if not exclusively, outside the nucleolar organizer region. The haploid genome of the wild-type D. melanogaster used in this work has 106 genes for 28S rRNA and 96–105 genes for 5S RNA.
Uptake of tetracycline by membrane preparations from Escherichia coli
Authors: Franklin T. J.
Published: August 2015

1. A membrane fraction from Escherichia coli has been prepared essentially free from ribosomes by treatment of the membranes with Triton X-100 at 0°C followed by differential centrifugation. 2. The ribosome-free membrane vesicles absorbed tetracycline by a reversible temperature-dependent process with an apparent Km of 0.029mm at pH7.5 and 37°C. 3. The absorption process was negligible below 25°C and had an optimum at 40°C; a pH optimum at 7.5 was observed. 4. The absorption of tetracycline was strongly inhibited by EDTA and ATP; ADP inhibited less strongly and AMP had no effect. 5. There was no significant difference in the rates or extent of uptake of tetracycline by membranes prepared from tetracycline-sensitive and tetracycline-resistant, R-factor-bearing E. coli.
The influence of the intracellular concentration on the uptake of l-[14C]valine by chopped tissue from cerebral cortex
Authors: Jones C. T. , Banks P.
Published: August 2015

1. The intracellular concentrations of sodium, potassium, ATP and creatine phosphate in chopped tissue from guinea-pig cerebral cortex were altered by a 10min period of electrical stimulation. 2. The ability of the tissue to take up l-[14C]valine and incorporate it into protein was tested at various times after the cessation of electrical stimulation. 3. The restoration of the ability of the tissue to accumulate l-[14C]valine correlated closely with the restoration of the resting intracellular concentrations of sodium and ATP.
Changes in the proportions of two mitochondrial populations during the development of embryonic chick liver
Authors: Pollak J. K. , Woog Maria
Published: August 2015

1. Two populations of morphologically intact mitochondria were isolated from embryonic, neonatal and adult chick liver by isopycnic centrifugation. 2. The protein/phospholipid ratio of the total mitochondrial fraction, the low-density mitochondria (B2, d1.176) and the high-density mitochondria (B3, d1.206) did not differ significantly. 3. During development there is a marked increase in the B2 fraction in relation to the B3 fraction. 4. Cytochrome oxidase and malate dehydrogenase activities as well as respiratory control increased during the embryonic development of the chick, though their rates of increase were not correlated. 5. In the three different embryonic stages that were investigated, as well as in the neonatal and adult chick, the protein/lipid as well as the protein/phospholipid ratio stayed constant and showed no progressive increase, as had been previously reported. 6. It was shown that forces greater than 18400gav. for 2h have to be used before chick liver mitochondria reach isopycnic equilibrium. 7. As for rat liver mitochondria, the constant protein/phospholipid ratio of the B2 and B3 fractions and their apparent morphological intactness leads one to conclude that the matrix space of B2 mitochondria is inaccessible to sucrose, whereas B3 mitochondria possess an inner membrane that is permeable to sucrose.
The hydrolysis by thermolysin of dipeptide derivatives that contain substituted cysteine
Authors: Damoglou A. P. , Lindley H. , Stapleton I. W.
Published: August 2015

1. The preparation of protected dipeptides of the form acetylglycylamino acid amides is described, where the amino acid is phenylalanine, leucine, valine, alanine, S-methylcysteine, S-ethylcysteine, S-benzylcysteine and S-phenylcysteine. 2. Kinetic parameters for the thermolytic hydrolysis of these blocked dipeptides are reported. The rate of hydrolysis was fastest when the amino acid was leucine or phenylalanine, slower when it was S-methylcysteine, valine or S-ethylcysteine, much slower when it was alanine, and negligible for S-phenylcysteine or S-benzylcysteine. 3. The results are compared with those for similar dipeptide derivatives with benzyloxycarbonyl and furylacryloyl blocking groups, which are hydrolysed faster.
Different effects of thymidine and 5-fluorouracil 2′-deoxyriboside on biosynthetic events in cultured P815Y mast cells
Authors: Bergeron J. J. M.
Published: August 2015

1. Addition of 2mm-thymidine, although resulting in the cessation of cell division, allows the continuation of phospholipid and protein synthesis and results in an increase in mean cell volume for at least 15h. 2. 5-Fluorouracil 2′-deoxyriboside inhibits cell division but differs from thymidine by inhibiting the synthesis of phospholipid and protein in a more marked manner. 3. The relation between these results and the P815Y cell cycle is discussed.
The isolation of ovomucoid variants differing in carbohydrate composition
Authors: Beeley J. G.
Published: August 2015

Three major and two minor species of ovomucoid were separated by chromatography on sulphoethyl-Sephadex. The predominant sialic acid-free species was further resolved into three fractions by DEAE-cellulose chromatography. Although all species of ovomucoid had closely similar trypsin-inhibiting activity, immunochemical properties and amino acid composition, they differ in carbohydrate composition. Wide variation was observed in the content of galactose, N-acetylglucosamine and sialic acid. Charge heterogeneity was related, in part, to variation in sialic acid content. The implications of variable carbohydrate composition for the structure and function of ovomucoid are discussed.
Isolation and nature of intracellular peptides from a cephalosporin C-producing Cephalosporium sp
Authors: Loder P. Bronwen , Abraham E. P.
Published: August 2015

1. Three peptides containing α-aminoadipic acid and cysteine have been obtained in small amounts from the mycelium of a Cephalosporium sp. 2. The peptides were precipitated as cuprous mercaptides together with glutathione and resolved from the latter and from each other by preparative paper electrophoresis and chromatography either in the sulphonic acid form or as S-sulphonyl derivatives. From the S-sulphonyl derivatives they were obtained in the thiol form. 3. One peptide (P3) was shown by amino acid analysis and the mass spectrum of the NS-ethoxycarbonyl derivative of its methyl ester to be δ-(l-α-aminoadipyl)-l-cysteinyl-d-valine. A second peptide (P2) contained α-aminoadipic acid, cysteine, valine and glycine, and the third peptide (P1) contained α-aminoadipic acid, cysteine, β-hydroxyvaline and glycine.
Evidence for the presence of a pool of glycerides with a rapid rate of turnover in brown fat from newborn rabbits
Authors: Knight B. L.
Published: August 2015

1. The specific radioactivity of [14C]glycerol released during the incubation of brown fat with [14C]glucose is much greater than that of the tissue lipid glycerol. 2. From a study of the release of [14C]glycerol from pre-labelled brown fat, it is concluded that the tissue contains a pool of glycerides with a higher rate of turnover than those in the main lipid store. 3. This pool contains newly synthesized glycerides, has a half-life of 25–30min and supplies about 25% of the glycerol liberated by brown fat. 4. Thus, a significant fraction of the total14C incorporated from glucose into brown-fat lipids is released as [14C]glycerol during an incubation.
Some relationships between R-factor and chromosomal β-lactamase in Gram-negative bacteria
Authors: Dale J. W. , Smith J. T.
Published: August 2015

1. The β-lactamases specified by Klebsiella aerogenes 418 and the R-factor R-7268 have been partially purified. 2. The molecular weights of the K. aerogenes strains 418 and 373, Aerobacter cloacae 53, R-7268 and R-TEM β-lactamases were all about 20000; that of the enzymes from Escherichia coli strains 419 and 214T was about 31000. 3. These enzymes were also compared by means of their Km values for benzylpenicillin and ampicillin, and their behaviour on starch-gel electrophoresis. 4. The β-lactamases specified by the two Klebsiella strains, the Aerobacter strain, and the R-factors R-TEM and R-7268 were found to comprise a broadly similar group. However, within this group, only two enzymes seemed to be identical, namely those specified by the two R-factors. The two E. coli strains produce identical β-lactamases which are very different from the ‘Klebsiella/Aerobacter-type’ enzymes.