bluuebunny/crossref_metadata_2025 huggingface dataset:
Metabolism of palmitate in perfused rat liver. Computer models of subcellular triacylglycerol metabolism
Authors: Kondrup Jens , Damgaard Stig E. , Fleron Peter
Published: August 2015

1. In the preceding paper [Kondrup (1979) Biochem. J.184, 63–71] the separation of two major fractions of hepatic triacylglycerol was described. One fraction contained triacylglycerol from the endoplasmic reticulum and from the Golgi apparatus. The other fraction contained triacylglycerol from the cytoplasmic lipid droplets. In the present paper possible precursor–product relationships between the two fractions were investigated by means of computer models. 2. The fatty acids present in di- and tri-acylglycerol in the fractions isolated in the time studies were analysed by gas chromatography. From this analysis the relative specific radioactivities, and contents, of palmitate in acylglycerols in the two fractions at the various time points were calculated. 3. A computer was used to predict relative specific radioactivities of pools in defined models of hepatic triacylglycerol metabolism. The acceptability of the models was evaluated by comparing predicted with measured relative specific radioactivities. 4. It is suggested that triacylglycerol in cytoplasmic lipid droplets does not originate (a) directly from triacylglycerol in the endoplasmic reticulum, (b) from a sub-pool of it or (c) directly from non-esterified fatty acids entering the cell. Rather, it is formed from diacylglycerol (and acyl-CoA) in the endoplasmic reticulum. Diacylglycerol, on the other hand, is furnished in part by hydrolysis of triacylglycerol in the endoplasmic reticulum. 5. This suggestion is discussed in relation to previous models of hepatic fatty acid metabolism.
Increase in liver nuclear tri-iodothyronine receptors associated with increased deoxyribonucleic acid by long-term administration of tri-iodothyronine in thyroidectomized rats
Authors: Nakamura Hirotoshi , Hamada Satoshi , Imura Hiroo
Published: August 2015

The effect of long-term administration of small amounts of tri-iodothyronine was examined on the nuclear tri-iodothyronine receptors in rat liver. The maximal binding capacity (Cmax.) and association constant (Ka) of the receptors were determined in thyroidectomized rats given vehicle alone (group A), 2μg of tri-iodothyronine/100g body wt. (group B) or 7μg of tri-iodothyronine/100g body wt. (group C) for 2 weeks. Scatchard analyses with correction for the amount of endogenous tri-iodothyronine revealed that Cmax. values per g of liver were increased to 1.5 and 2.7 times the control value in groups B and C respectively. Since concentrations of DNA per g of liver were significantly increased in the two groups of hormone-treated rats, Cmax. values per mg of DNA were nearly the same in group B, but still increased significantly in group C compared with group A. Ka values remained unchanged in all three groups of animals. Mitochondrial α-glycerophosphate dehydrogenase activity was 9.6 and 28.7 times as high in groups B and C, respectively, as in group A. Concentrations of endogenous tri-iodothyronine bound to non-histone protein were substantially increased in groups B and C, although concentrations of serum tri-iodothyronine remained rather low. The results obtained indicate that the long-term administration of tri-iodothyronine in small doses induces an increase in the nuclear receptors associated with increased DNA with and without accompanying a relative increase in the receptor concentration in thyroidectomized rats. Also the hormonal effect is closely related to the total number of the nuclear receptors and the concentrations of nuclear tri-iodothyronine bound to the receptors rather than the serum tri-iodothyronine concentrations.
Metabolism of cholecalciferol in land snails
Authors: Weiner S , Noff D , Meyer M S , Weisman Y , Edelstein S
Published: August 2015

1. Radioactively labelled cholecalciferol was injected into the land snails Levantina hiersolyma and Theba pisana. Three metabolites (C, D and E), more polar than cholecalciferol, were found. 2. Metabolite C was found to be identical with 25-hydroxycholecalciferol. On injection of 25-hydroxy[26,27-3H]cholecalciferol, metabolite E was predominantly formed. Metabolite D was predominantly formed from cholecalciferol. Metabolites D and E differ from any known cholecalciferol metabolites. 3. The intestine was found to be the tissue capable of carrying out the transformation of 25-hydroxycholecalciferol into metabolite E. 4. 25-Hydroxycholecalciferol and metabolite E were localized in the digestive gland of the snail, the tissue responsible for the absorption of Ca2+ and its storage. Metabolite D was not localized in any specific tissue.
Occurrence and operation of the glycollate–glyoxylate shuttle in mitochondria of Euglena gracilis Z
Authors: Yokota A , Kitaoka S
Published: August 2015

Both glyoxylate reductase (NADP+) and glycollate dehydrogenase were located exclusively in mitochondria in Euglena gracilis and constitute the glycollate–glyoxylate shuttle, whose existence in higher plants was thought doubtful, owing to different subcellular locations of the two enzymes. Disrupted Euglena mitochondria showed a glycollate-dependent NADPH oxidation, indicating actual operation of the shuttle in this protozoon.
Effects of phorbol 12-myristate 13-acetate and cortisol interaction on steroid-binding capacity in the rat
Authors: Janssens J P , de Loecker W
Published: August 2015

The specificity of the cortisol-receptor protein is examined in plasma and liver cytosol of rats. Phorbol 12-myristate 13-acetate does not inhibit the binding of cortisol to transcortin, nor does it affect the binding capacity of dexamethasone to the intracellular glucocorticoid receptor, but, by interacting with the cortisol molecule, it interferes with hormone-mediated processes in the cell.
Two forms of ‘malic’ enzyme with different regulatory properties in Trypanosoma cruzi
Authors: Cannata J J B , Frasch A C C , Cataldi de Flombaum M A , Segura E L , Cazzulo J J
Published: August 2015

1. Cell-free extracts from culture epimastigotes of Trypanosoma cruzi contained two forms of NADP+-linked ‘malic’ enzyme (EC 1.1.1.40), I and II, with the same molecular weight but different electrophoretic mobilities and kinetic and regulatory properties. 2. The apparent Km for L-malate was lower for ‘malic’ enzyme I, with hyperbolic kinetics, whereas the kinetic pattern for ‘malic’ enzyme II was slightly sigmoidal (h 1.4). The kinetics for NADPH were hyperbolic for ‘malic’ enzyme I, and very complex for ‘malic’ enzyme II, suggesting both positive and negative co-operativity. 3. ‘Malic’ enzyme II was markedly inhibited by adenine nucleotides; AMP was the the most effective, at least in the presence of an excess of MnCl2. ‘Malic’ enzyme I was much less affected by the nucleotides. Both enzyme forms were inhibited by oxaloacetate, competitively towards L-malate, but the apparent Ki for ‘malic’ enzyme I (9 microM) was 10-fold lower than the value for ‘malic’ enzyme II. ‘Malic’ enzyme II, but not ‘malic’ enzyme I, was activated by L-aspartate and succinate (apparent Ka of 0.12 and 0.5 mM respectively); the activators caused a decrease in the apparent Km for L-malate and, to a lesser extent, in the apparent Km for NADP+. L-Aspartate, but not succinate, increased the apparent Vmax. 4. The inhibition by AMP suggests regulation by energy charge, with the L-malate-decarboxylation reaction catalysed by ‘malic’ enzyme II fulfilling a biosynthetic role. The inhibition by oxaloacetate and the activation by succinate are probably involved in the regulation of the ‘partial aerobic fermentation’ of glucose which yields succinate as final product. The activation by L-aspartate would facilitate the catabolism of this amino acid, when present in excess in the growth medium.
The tricarboxylic acid cycle in Dictyostelium discoideum. A model of the cycle at preculmination and aggregation
Authors: Kelly P J , Kelleher J K , Wright B E
Published: August 2015

A preliminary model of tricarboxylic acid-cycle activity in Dictyostelium discoideum is presented. Specific-radioactivity labelling patterns of intra- and extra-mitochondrial pools are simulated by this model and compared with the experimental data. The model arrived at by this method shows the following features. (1) The cycle flux rate is approx. 0.4 mM/min. (2) Both fumarate and malate are compartmentalized at approx. 1:5 between cycle pools and non-cycle pools. These may represent mitochondrial and cytoplasmic pools. Citrate is compartmentalized at 1:10. Succinate appears to exist in three compartments, two of which become labelled by [14C]glutamate and only one by [14C]aspartate (3) Two pools of aspartate with two associated pools of oxaloacetate are necessary for simulation. (4) Exchange between the cycle and non-cycle pools of both citrate and fumarate occurs at very low rates of about 0.003 mM/min, whereas exchange between the malate pools is about 0.004 mM/min. The exchange reaction glutamate in equilibrium 2-oxoglutarate runs at approx. 15 times the cycle flux. (5) A reaction catalysed by “malic” enzyme is included in the model, as this reaction is necessary for complete oxidation of amino acid substrates. (6) Calculation of the ATP yield from the model is consistent with earlier estimates of ATP turnover if the activity of adenylate kinase is considered.
Changes in the nuclear polyamine content of chick erythrocytes during embryonic development
Authors: Goyns M H
Published: August 2015

The polyamine content of the circulating erythrocyte population in the embryonic chick was studied during its development. Total cellular polyamine content fell dramatically between 5 and 7 days of development, paralleling the decrease in metabolic activity exhibited by these cells. Nuclei were isolated from the erythrocytes by a non-aqueous technique, which not only eliminated the polyamine loss that occurred with aqueous isolation, but also prevented redistribution of the polyamines from the cytoplasm. Nuclear spermidine and spermine contents decreased markedly between 5 and 6 days of development from 31 to 10 pmol/microgram of DNA and from 33 to 18 pmol/microgram of DNA respectively. Thereafter the spermine content remained constant, but the spermidine content continued to decline. Good correlations between spermidine and RNA contents were observed in both cells and nuclei, and similarly between spermine and RNA contents in cells, but no such correlation was observed between spermine and RNA in nuclei.
Identification of a defence mechanism in vivo against the leakage of enterokinase into the blood
Authors: Grant D A W , Magee A I , Meeks D , Regan C , Bainbridge D R , Hermon-Taylor J
Published: August 2015

1. The serum proteinase inhibitors alpha 1-antitrypsin, alpha 2-macroglobulin, inter-alpha-trypsin inhibitor and C1-esterase inhibitor were found not to affect the catalytic activity of human enterokinase, whereas bovine trypsin activity was modified essentially as expected. Enterokinase was also not inhibited by Trasylol (trypsin inhibitor from bovine lung) or bovine pancreatic trypsin inhibitor. No other component in human or mouse serum complexing with enterokinase was identified. 2. Human enterokinase administered intravenously into mice was rapidly cleared from the circulation with a half-life of 2.5 min. This removal was not the result of the difference in species, since partially purified mouse enterokinase was cleared at the same rate as the human enzyme. Clearance was mediated by recognition of the carbohydrate portion of enterokinase and not through specific recognition of its catalytic site. Immunofluorescent staining showed that the enzyme accumulated in the liver. Attempts to block the clearance by the simultaneous infusion of competing glycoproteins suggested that enterokinase was taken up by hepatocytes. Of the glycoproteins tested only two, human lactoferrin (terminal fucosyl alpha 1 leads to 3 N-acetylglucosamine) and bovine asialo-fetuin (terminal galactosyl beta 1 leads to 4 N-acetylglucosamine) were weakly competitive. Two inhibitors of endocytosis, Intralipid and Triton WR1339, failed to delay the removal of enterokinase. It is proposed that enterokinase is cleared from the circulation by an as yet uncharacterized hepatocyte receptor.
Inhibition of collagen-induced platelet aggregation by antibodies to distinct types of collagens
Authors: Balleisen L , Nowack H , Gay S , Timpl R
Published: August 2015

Aggregation of platelets by fibrils formed from collagens type I, II and III could be inhibited by coating the fibrils with anti-collagen antibodies or Fab fragments. Similar results were obtained in a clot-retraction assay. Inhibition was achieved with stoichiometric amounts of antibodies and was specific for each type of collagen. Aggregation caused by a mixture of type-I and -III collagens could only be inhibited by a mixture of antibodies against both collagens. The data show that each interstitial collagen is capable of interacting with platelets and do not support the concept of an outstanding activity of type-III collagen.
Fate of bradykinin-potentiating peptide 9a after intravenous injection
Authors: Martin L C , Ryan J W , Fisher G H , Chung A , Epstein M , Stewart J M
Published: August 2015

The fat of less than Glu1-3H-labelled bradykinin-potentiating peptide 9a [BPP9a; less than Glu-Trp-Pro-Arg-Pro-Gln-Ile-Pro-Pro, an inhibitor of angiotensin-converting enzyme (peptidyl dipeptidase)] was studied in the rabbit. After intravenous injection, BPP9a was rapidly removed from blood and much of the associated radioactivity was excreted in urine. Approx. 8% of the radioactivity in urine collected 2h after drug administration occurred in the form of BPP9a itself, the remainder occurring in three lower homologues: less than Glu-Trp (60%), less Glu-Trp-Pro-Arg-Pro-Gln (20%) and less than Glu-Trp-Pro-Arg-Pro-Gln-Ile (12%). Hydrolysis was not accounted for by enzymes in blood or urine. Apparently hydrolysis occurred within the kidney, as less than Gl-Trp was obtained in 60% yield in urine of isolated rat kidney perfused with [less than Glu1-3H]BPP9a.
Specificity of P2 primary alkylsulphohydrolase induction in the detergent-degrading bacterium Pseudomonas C12B. Effects of alkanesulphonates, alkyl sulphates and other related compounds
Authors: Cloves J M , Dodgson K S , White G F , Fitzgerald J W
Published: August 2015

Primary alkanesulphonates were shown to serve as non-metabolizable (gratuitous) inducers of the P2 primary alkylsulphohydrolase enzyme in resting cell suspensions of Pseudomonas C12B. The effects of increasing concentrations of inducer on the production of enzyme were complex and suggestive of a multiphasic phenomenon. However, it was possible to determine Kinducer constants (analogous to Km or Ki) for alkanesulphonates of chain length from C7 to c12. these decreased with increasing chain length in a manner characteristic of an homologous series. Primary alkyl sulphates also served as good inducers of alkylsulphohydrolase, but valid kinetic values could not be obtained because these esters are good substrates for the enzyme and are therefore appreciably hydrolysed during the induction period. Small amounts of enzyme were also produced when cyprinol sulphate, dodecyltriethoxy sulphate C12H23-[O-CH2-CH2]3-O-SO3-Na+), Crag herbicide and some secondary alkyl sulphates were tested as inducers.
Characterization of the intermediates in the reaction of mixed-valence state soluble cytochrome oxidase with oxygen at low temperatures by optical and electron-paramagnetic-resonance spectroscopy
Authors: Clore G M , Andréasson L E , Karlsson B , Aasa R , Malmström B G
Published: August 2015

The reaction of soluble mixed-valence-state (a3+CuA2+.CuB+A32+) cytochrome oxidase with O2 at low temperature was studied by optical and e.p.r. spectroscopy. The existence of three intermediates [Clore & Chance (1978) Biochem. J. 173, 799-8101] was confirmed. From the e.p.r data it is clear that cytochrome a and CuA remain in the low-spin ferric and cupric states respectively throughout the reaction. No e.p.r. signals attributable to cytochrome a3 or CuB were seen in the intermediates. The difference spectra (intermediates minus unliganded mixed-valence-state cytochrome oxidase) and absolute spectra of the three intermediates were obtained. The chemcal nature of the three intermediates is discussed in terms of their spectroscopic properties. A catalytic cycle for cytochrome oxidase is proposed.
Sensitivity and other factors affecting biospecific desorption in chromatography of proteins. A study by computer simulation
Authors: Yon R J
Published: August 2015

Some theoretical aspects of the desorption of a column-bound protein by elution with its biospecific ligand are considered in cases where, in comparison with the unliganded protein, the protein-ligand complex has a diminished but finite affinity for the adsorbent. A quantity termed the biospecific sensitivity, B, is introduced to facilitate comparison between different systems. Biospecific sensitivity may be defined as the fractional change in standard free energy of adsorption on formation of the protein-ligand complex. The effects of a moderate-to-low biospecific sensitivity on theoretical desorption profiles have been examined by using a computer simulation of the classical multiple-plate column model. Desorption was simulated under various boundary conditions involving protein-adsorbent and protein-ligand affinities and the initial concentrations of adsorption sites, protein and ligand. These simulations suggest that, when the biospecific sensitivity is low, desorption is optimized if (a) the unliganded protein is adsorbed as weakly as possible, (b) the column is loaded to near-saturation with the required protein, (c) the free ligand concentration is many times greater than that giving near-saturation of the protein in free solution, and (d) protein contaminants with high affinity for the adsorbent, and present in large amount, are removed in preliminary purification steps.
Isoelectric-focusing properties and carbohydrate content of pea (Pisum sativum) legumin
Authors: Gatehouse John A. , Croy Ronald R. D. , Boulter Donald
Published: August 2015

Legumin from pea (Pisum sativum) is a molecule made up of six pairs of subunits, each pair consisting of an ‘acidic’ subunit (mol.wt. about 40000) and a ‘basic’ subunit (mol.wt. about 20000) linked by one or more disulphide bonds. The heterogeneity of legumin has been investigated by isoelectric focusing; undissociated legumin could not be focused satisfactorily, but legumin subunits could be analysed under dissociating conditions. 8m-Urea was not found to be a satisfactory medium for isoelectric focusing of legumin, as the ‘basic’ subunits showed a shift in pI with time of incubation in urea. A new dissociating medium for isoelectric focusing, namely 50% (v/v) formamide, was used for analysis of legumin, which gave pI values of 5.0–5.3 for the ‘acidic’ subunits, and 8.3–8.7 for the ‘basic’ subunits. Both types of subunits were shown to be heterogeneous in charge and molecular weight by two-dimensional analysis employing isoelectric focusing in the first dimension and sodium dodecyl sulphate/polyacrylamide gel electrophoresis in the second. The ‘basic’ and ‘acidic’ subunits of legumin were separated on the preparative scale by ion-exchange chromatography in 50% formamide. Carbohydrate attached to the protein was investigated as a possible cause of the heterogeneity of legumin subunits. However, both a fluorescent-labelling technique and a sensitive radioactive-labelling technique failed to show any carbohydrate bound to legumin subunits, and it was concluded that legumin is not a glycoprotein.
An adenosine triphosphate-dependent carbamoylphosphate-3-hydroxymethylcephem O-carbamoyltransferase from Streptomyces clavuligerus
Authors: Brewer S J , Taylor P M , Turner M K
Published: August 2015

Cell-free supernatants from cells of Streptomyces clavuligerus (N.R.R.L. 3585), which are actively synthesizing cephamycin C, transfer a carbamoyl group from carbamoylphosphate to a 3-hydroxymethylceph-3-em-4-carboxylic acid nucleus to form a 3-carbamoyloxymethylcephem. This reaction was stimulated by nucleoside triphosphates and by a mixture of Mn2+ and Mg2+ cations. The enzyme responsible was purified 40-fold by batch absorption onto DEAE-cellulose and hydroxyapatite. The purified O-carbamoyltransferase is most active at pH 6.8. It is stabilized by phosphate anions, but is inhibited by PPi anions, (NH4)2SO4 or NaCl. The enzyme is stimulated by ATP, but it is not known whether this nucleotide acts as an effector or as a substrate. Some activity is observed with dATP, but two other analogues of ATP, in which a methylene group replaced the oxygen atom between the alpha- and beta- or the beta- and gamma-phosphorus atoms, inhibit the action of ATP itself. The enzyme synthesizes a wide range of 3-carbamoyloxymethylcephems. The structure of some of these products, for example that of cefuroxime (3-carbamoyloxymethyl-7 beta-[2-(fur-2-yl)-2-syn-methoxyiminoacetamido]ceph-3-em-4-carboxylic acid), was confirmed by their proton-n.m.r. spectra.
The size of collagen fibrils that stimulate platelet aggregation in human plasma
Authors: Kronick P , Jimenez S A
Published: August 2015

Suspensions of collagen fibrils of different size were prepared from solutions of radioactive tropocollagen type I by either differential centrifugation or differential incubation at elevated temperature. The fractions were compared with respect to their ability to stimulate human blood platelet aggregation in plasma, their binding to human platelets, and their morphology, as seen in the electron microscope. Although small particles with a sedimentation coefficient as low as 4.5 S bound to platelets, aggregation was not observed in the presence of collagen multimers and protofibrils without visible cross-bands in stained specimens. The onset of platelet-aggregating activity before the appearance of turbidity in collagen solutions incubated at elevated temperature is due to the formation of a few banded fibrils; this early onset and the fibrils do not appear in collagen solutions that have been ulctracentrifuged before incubation.
Microbial metabolism of amino alcohols. Biosynthetic utilization of ethanolamine for lipid synthesis by bacteria
Authors: Shukla S D , Turner J M
Published: August 2015

1. Ten bacteria utilizing [2-14C]ethanol-2-amine as the sole or major source of nitrogen for growth on glycerol + salts medium incorporated radioactivity into a variety of bacterial substances. A high proportion was commonly found in lipid fractions, particularly in the case of Erwinia carotovora. 2. Detailed studies of [14C]ethanolamine incorporation into lipids by five bacteria, including E. carotovora, showed that all detectable lipids were labelled. Even where phosphatidylethanolamine was the major lipid labelled, radioactivity was predominantly in the fatty acid rather than the base moiety. The labelled fatty acids were identified in each case. 3. The addition of acetate to growth media decreased the incorporation of radioactivity from ethanolamine into both fatty acid and phosphatidyl-base fragments of lipids from all the bacteria except Mycobacterium smegmatis. Experiments with [3H]ethanolamine and [14C]acetate confirmed that unlabelled acetate decreased the incorporation of both radioactive isotopes into lipids, except in the case of M. smegmatis. 4. Enzyme studies suggested one of two metabolic routes between ethanolamine and acetyl-CoA for each of four bacteria. A role for ethanolamine O-phosphate was not obligatory for the incorporation of [14C]ethanolamine into phospholipids, but correlated with CoA-independent aldehyde dehydrogenase activity.
Glucose metabolism and recycling of radioactively labelled glucose in the Zucker genetically obese rat (fa/fa)
Authors: Wade A J
Published: August 2015

1. The glucose metabolism of conscious lean and obese rats of the Zucker strain was studied by using doubly labelled glucose ([6-3H,U-14C]glucose) given by intravenous injection as a single dose. Fed animals were used, allowing the study to be made in conditions favouring active lipogenesis. 2. At any given prior food intake (consumption during preceding 24 h), the irreversible glucose replacement rate, R0, was considerably higher in the growing obese rat (4-6 months old) when both of these variables were scaled in terms of the total body water of the animals. 3. When scaled in a similar way, the minimal mass of glucose (Mmin.) was also larger in the obese rats. The mean transit time, t, through the pool did not differ significantly between the two groups, but there was a tendency for this to be shorter in obese rats. 4. There was no difference in the proportion of 14C (derived from metabolized labelled glucose) that recycled as [14C]-glucose after passing through the pyruvate pool in the two groups of rats if the rate of recycling of radioactivity (Rc) was expressed as a percentage of R0.
The mother-cell-membrane adenosine triphosphatase of sporulating Clostridium pasteurianum
Authors: Clarke David J. , Morris J. Gareth
Published: August 2015

1. Sporulation of Clostridium pasteurianum effects several changes in its proton-translocating cell-membrane H+-ATPase. Notable among these are the acquisition of susceptibility to activation by trypsin and a changed protein subunit composition. 2. A protein was isolated from the mother-cell membrane that inhibited the ATP phosphohydrolase activity of purified vegetative-cell-membrane H+-ATPase [BF0F1 complex, which consists of soluble ATPase (BF1) and the proton-channel component (BF0)] and rendered it susceptible to trypsin activation. 3. This trypsin-sensitive inhibitor protein had a molecular weight of 10000 and on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis was indistinguishable from the novel protein subunit e of the mother-cell-membrane ATPase 4. In bacteriorhodopsin-containing everted membrane vesicles, the specific ATP synthetase activity of the mother-cell-membrane ATPase was significantly greater than that of the vegetative-cell-membrane ATPase. 5. Treatment with trypsin-sensitive inhibitor protein of artificial proteoliposomes containing bacteriorhodopsin and vegetative-cell-membrane H+-ATPase (BF0F1) significantly increased the specific ATP synthetase activity of this enzyme. 6. The ATP synthetase activity of crude cell-membrane preparations from cultures of Clostridium pasteurianum increased during that period in the course of sporulation when the membrane ATP phosphohydrolase was both most rapidly decreasing in specific activity and acquiring its susceptibility to activation by trypsin.
Characterization of rat hypothalamic progestin binding by spheroidal hydroxylapatite chromatography
Authors: Thrower S , Lim L
Published: August 2015

The progestin-high-affinity-binding components in rat target tissues have been assayed by a simple and precise procedure by using spheroidal hydroxylapatite. The progestin ‘receptors’ in the uterus and hypothalamus of female rats are highly specific for progestins, which they bind with high affinity (Kd for [3H]progesterone in hypothalamus is 1.9 nM and in uterus is 3.7 nM). The dissociation of [3H]progesterone from receptor in vitro is rapid: t1/2 6 degrees C = 45 min in uterine cytosol; t1/2 6 degrees C = 160 min in hypothalamic cytosol. The binding is destroyed by proteinase. In the cytosol of hypothalamus and cortex of developing rats, progestin ‘receptors’ were present in both male and female rats by 2-3 days after birth; subsequent changes in concentration of these ‘receptors’ appeared to be independent of sex. Concentrations of progestin ‘receptor’ were close to adult values by 8-9 days, and thereafter changed relatively little.
The effect of nitrofurantoin on deoxyribonucleic acid synthesis in plant mitochondria
Authors: Palit S , Goswami B B , Dube D K
Published: August 2015

Nitrofurantoin (1-([(5-nitrofuran-2-yl)methylene]amino)imidazolidine-2,4-dione), a widely used drug and also a well-known bacterial mutagen, inhibits DNA synthesis in mitochondria from 48 h etiolated seedlings of Vigna sinensis (Linn.) Savi (snake bean). The effect appears at the level of the uptake of radioactive deoxynucleoside triphosphates by the plant mitochondria. Nitrofurantoin does not inhibit DNA synthesis in vitro by homogeneous Escherichia coli DNA polymerase I and DNA polymerase from avian-myeloblastosis virus. No specific nitroreductase activity could be detected in mitochondria.
Contribution of rat liver and gastrointestinal tract to whole-body protein synthesis in the rat
Authors: McNurlan M A , Garlick P J
Published: August 2015

The rate of protein synthesis was assessed in liver, stomach, small and large intestine and in the whole body of rats by injection of 100 mumol of [14C]leucine/100 g body wt. In each of the tissues turnover was very rapid, so that taken together they accounted for 43% of the protein synthesized by the whole animal.
Restoration of respiratory electron-transport reactions in quinone-depleted particle preparations from Anacystis nidulans
Authors: Peschek G A
Published: August 2015

Electron transport from H2, NADPH, NADH and succinate to O2 or ferricytochrome c in respiratory particles isolated from Anacystis nidulans in which hydrogenase had been induced was abolished after extraction of the membranes with n-pentane; oxidation of ascorbate plus NNN‘N’-tetramethyl-p-phenylenediamine remained unaffected. Incorporation of authentic ubiquinone-10, plastoquinone-9, menaquinone-7 and phylloquinone (in order of increasing efficiency) restored the electron-transport reactions. ATP-dependent reversed electron flow from NNN‘N’-tetramethyl-p-phenylenediamine to NADP+ or, via the membrane-bound hydrogenase, to H+ was likewise abolished by pentane extraction and restored by incorporation of phylloquinone. Participation of the incorporated quinones in the respiratory electron-transport reactions of reconstituted particles was confirmed by measuring the degree of steady-state reduction of the quinones. Isolation and identification of the quinones present in native Anacystis membranes yielded mainly plastoquinone-9 and phylloquinone; neither menaquinone nor alpha-tocopherolquinone could be detected. Together with the results from reconstitution experiments this suggests that phylloquinone might function as the main respiratory quinone in Anacystis nidulans.
Fibronectin–collagen binding and requirement during cellular adhesion
Authors: Gold Leslie I. , Pearlstein Edward
Published: August 2015

Fibronectin isolated from human plasma and from the extracellular matrices of cell monolayers mediates the attachment in vitro and spreading of trypsin-treated cells on a collagen substratum. Fibronectin-dependent kinetics of cellular attachment to collagen were studied for several adherent cell types. It was shown that trypsin-treated human umbilical-cord cells, mouse sarcoma CMT81 cells, endothelial cells, and human fibroblasts from a patient with Glanzmann's disease were completely dependent on fibronectin for their attachment to collagen, whereas guinea-pig and monkey smooth-muscle cells and chick-embryo secondary fibroblasts displayed varying degrees of dependence on fibronectin for their attachment. Radiolabelled human plasma fibronectin possessed similar affinity for collagen types I, II and III from a variety of sources. The fibronectin bound equally well to the collagens with or without prior urea treatment. However, in the fibronectin-mediated adhesion assay using PyBHK fibroblasts, a greater number of cells adhered and more spreading was observed on urea-treated collagen. Fibronectin extracted from the extracellular matrix of chick-embryo fibroblasts and that purified from human plasma demonstrated very similar kinetics of complexing to collagencoated tissue-culture dishes. Fibronectin from both sources bound to collagen in the presence of 0.05–4.0m-NaCl and over the pH range 2.6–10.6. The binding was inhibited when fibronectin was incubated with 40–80% ethylene glycol, the ionic detergents sodium dodecyl sulphate and deoxycholate, and the non-ionic detergents Nonidet P-40, Tween 80 and Triton X-100, all at a concentration of 0.1%. From these results we proposed that fibronectin–collagen complexing is mainly attributable to hydrophobic interactions.
Copper metabolism in mottled mouse mutants. Defective placental transfer of 64Cu to foetal brindled (Mo
br) mice
Authors: Mann J R , Camakaris J , Danks D M
Published: August 2015

Decreased placental transfer of 64Cu into both Mobr/Y and Mobr/+ foetuses was apparent after its administration to pregnant Mobr/+ mice. The severity of impairment and the degree of retention and accumulation of 64Cu in the placenta was not as great as has been previously observed in the gut of the suckling Mobr/Y mice.
A species-non-specific liver plasma-membrane antigen and its involvement in chronic active hepatitis
Authors: De Kretser T A , McFarlane I G , Eddleston A L , Williams R
Published: August 2015

Rabbit liver plasma membranes were isolated and purified by using an aqueous two-phase polymer system. Examination of these preparations with respect to electron-microscopical appearance, distribution of marker enzymes and gross biochemical composition revealed them to be free from contamination by intracellular components. Sera from ten patients with chronic active hepatitis, four with and six without hepatitis B viral markers (HBsAg) in their sera, produced a single precipitin line on immunodiffusion against a detergent extract of the isolated plasma membranes. Sera from HBsAg-positive and HBsAg-negative patients reacted against the same antigen. This antigen was enriched in the plasma membrane preparations compared with whole-liver homogenates and was identical with a species-non-specific antigen in a macromolecular fraction of normal human liver, which has been previously described as liver-specific lipoprotein.
Evidence from rat liver nuclear preparations that latency of microsomal UDP-glucuronosyltransferase is associated with vesiculation
Authors: Wishart G J , Fry D J
Published: August 2015

1. Nuclear, nuclear-envelope and microsomal preparations were prepared from rat liver, and their purity and morphology monitored by electron microscopy. 2. UDP-glucuronosyltransferase activity in microsomal preparations, but not in standard nuclear or nuclear-envelope preparations, displays latency from the criterion of being enhanced (‘activated’) by a range of detergents or the endogenous activator UDP-N-acetyl-glucosamine. 3. Nuclear preparations resemble activated rather than native microsomal preparations in failing to transfer glucuronic acid from 4-nitrophenyl glucuronide to 2-aminophenol. 4. Electron microscopy indicates that membranes of nuclear preparations and of our standard nuclear-envelope preparations remain, as in vivo, in a cisternal arrangement, whereas those of microsomal preparations are vesiculated. 5. In nuclear-envelope preparations in which vesiculation has been encouraged, the transferase can be activated by detergents. 6. We suggest that latency of UDP-glucuronosyltransferase results from vesiculation of membranes during preparation and that the latency of the microsomal transferase is largely a preparative artefact.
Prolonged effect of insulin on glucose uptake by rat skeletal muscle
Authors: Lewis Stephen B. , Schultz Thomas A. , Daniels Ellen L. , Bliziotes Michael M. , Montague William
Published: August 2015

The fate of insulin, as it relates to its action on skeletal-muscle glucose uptake, was studied in non-cyclically perfused rat hindlimbs. Insulin (1m-i.u./ml) with and without 125I-labelled insulin was infused intra-arterially for 5 or 6min. Net glucose uptake and the release of 125I-labelled insulin into the venous effluent were evaluated by arteriovenous-difference measurements for an additional 24–32min. The infusion of insulin for 5min promoted glucose uptake, an effect that persisted throughout a subsequent 25min of perfusion in the absence of insulin. The addition of insulin antibody to the perfusate in the presence of insulin blocked the action of insulin on glucose uptake, but it failed to alter insulin action if the muscle tissue had been exposed to insulin before addition of antibody. When 125I-labelled albumin was infused for 6min, venous effluent radioactivity decayed rapidly and remained HClO4-insoluble and there was no significant tissue retension of radioactivity. Comparable experiments in which 125I-labelled insulin was infused for 6min revealed that the venous effluent radioactivity decayed more slowly, a significant amount of the 125I-labelled insulin appeared as fragments (HClO4-soluble) and there was a significant retention of radioactivity in the tissue. Radioactivity in muscle tissue biopsies obtained 28min after infusion of 125I-labelled insulin was associated largely with intact insulin and a peptide of mol.wt. 2400. The total radioactivity retained in the muscle at this time was 7% of the amount infused. An insulin bolus (1i.u.) failed to increase the discharge of this tissue-associated radioactivity. These results suggest that insulin and a product of insulin metabolism persists in muscle tissue long after the arterial presence of insulin ends. This tissue residence and processing of insulin may be important components of insulin's prolonged action on glucose uptake by skeletal muscle.
Actions of insulin and vitamin A on Sertoli cells
Authors: Karl A F , Griswold M D
Published: August 2015

The synthesis of androgen-binding protein by cultured Sertoli cells is increased by insulin, retinol, follitropin and testosterone. Only follitropin will stimulate an increase in cyclic AMP in these cells, yet each agent individually increased the synthesis of androgen-binding protein in short-term cultures. For long-term culture of Sertoli cells follitropin, testosterone, insulin, and retinol appeared to act synergistically to prolong the ability of the cells to secrete androgen-binding protein. These are the first reported results which suggest an action of insulin and retinol directly on Sertoli cells even though the importance of both factors in male reproduction is known.
Impaired lipogenesis in mammary glands of lactating rats fed on a cafeteria diet. Reversal of inhibition of glucose metabolism in vitro by insulin
Authors: Agius L , Rolls B J , Rowe E A , Williamson D H
Published: August 2015

In lactating rats fed on a cafeteria diet (chow plus palatable high-energy foods) the decreased glucose uptake and lipogenesis in vitro in acini correlated with the depressed mammary-gland lipogenesis in vivo. Insulin in vitro restored the rate of glucose uptake and its conversion to lipid to values approaching those for acini from rats fed on the chow diet alone.
Purification and characterization of α-l-fucosidase from the liver of a fucosidosis patient
Authors: Alhadeff Jack A. , Andrews-Smith Grai L.
Published: August 2015

α-l-Fucosidase was partially purified from the liver of a fucosidosis patient by column chromatography either on agarose–ε-aminohexanoylfucosamine or on concanavalin A–Sepharose, despite no apparent enzymic activity in the crude liver supernatant. Mixing studies indicated that the liver of the fucosidosis patient did not lack activators or contain inhibitors of α-l-fucosidase activity. The partially purified α-l-fucosidase from the liver of the fucosidosis patient exhibits a 4–5-fold-increased Michaelis constant for the 4-methylumbelliferyl substrate (700–750μm) and a greatly decreased thermostability at 55°C compared with the normal liver enzyme. The pH–activity curve is similar to that for the normal enzyme between pH5 and 8, but quite dissimilar in the acid region (pH3.0–4.5): below pH4.5 the α-l-fucosidase shows no activity, whereas the normal enzyme retains considerable activity (≥50% of maximal activity). Isoelectric focusing of the α-l-fucosidase revealed one major form with pI5.8 and other possible minor forms. No cross-reacting material was detected when the α-l-fucosidase was run in double-immunodiffusion experiments against the immunoglobulin-G fraction of anti-(α-l-fucosidase) antibodies, but the enzyme was immunoprecipitated by this immunoglobulin-G fraction. For at least the fucosidosis patient being studied here, all the data suggest retention of a thermolabile portion of normal α-l-fucosidase (with characteristic Michaelis constant and pH–activity curve) or production of a kinetically altered α-l-fucosidase with decreased catalytic activity but antigenic similarity to the normal enzyme.
Structures and apoprotein linkages of phycoerythrobilin and phycocyanobilin
Authors: Killilea S. Derek , O'Carra Padraig , Murphy Richard F.
Published: August 2015

Phycoerythrobilin and phycocyanobilin are covalently attached to the apoproteins of phycoerythrins and phycocyanins. One linkage consists of an ester bond between the hydroxy group of a serine residue and the propionate side chain on one of the inner pyrrole rings (probably ring C). The other linkage is a labile thioether bond between a cysteine residue and the two-carbon side chain on pyrrole ring A. This side chain and both of the α-positions of the ring A are in the reduced state. This constitutes an important structural revision, since, in the structures currently accepted for the phycobilins, the two-carbon side chain on ring A is depicted as an ethylidene grouping and this has been regarded not only as a very characteristic feature of the phycobilins, but also as a probable structural feature of the chromophore of phytochrome, largely on the basis of other analogies with the phycobilins. The ethylidene-containing structures apply instead to artefact forms of the pigments released from the apoproteins by treatment with hot methanol. Cleavage of the ring-A linkage involves an elimination reaction releasing the cysteine residue and generating a double bond in the ring-A side chain. During cleavage in methanol the direction of the elimination is towards the ring, generating the ethylidene double bond. Since this is linked to the conjugated system, the methanol-released pigments differ spectrally from the native phycobilins. During acid-catalysed release of the pigments, the elimination apparently goes in the opposite direction, generating a double bond at the outer position of the side chain. Since this double bond is not linked to the conjugated system, the acid-released pigments remain spectrally identical with their protein-bound counterparts.
Characteristics of solubilized human-somatotropin-binding protein from the liver of pregnant rabbits
Authors: Tsushima T , Sasaki N , Imai Y , Matsuzaki F , Friesen H G
Published: August 2015

A specific binding site for somatotropin was solubilized by 1% (v/v) Triton X-100 from a crude particulate membrane fraction of pregnant rabbit liver, partially purified and characterized. The solubilized binding site retained many of the characteristics observed in the original particulate fraction, indicating that extraction of the binding site with Triton X-100 does not cause any major changes in its properties. The binding of human 125I-labelled-somatotropin to the solubilized binding site is a saturable and reversible process, depending on temperature, incubation time, pH and ionic environment. Analysis of the kinetic data revealed a finite number of binding sites with an affinity constant of 0.32 × 10(10)M-1. The binding activity for human 125I-labelled-somatotropin was adsorbed to a concanavalin-A-Sepharose column and was dissociated from the column with alpha-methyl-D-glucoside, suggesting that the binding protein may be a glycoprotein. Using affinity chromatography on concanavalin-A-Sepharose, ion-exchange chromatography on DEAE-cellulose and gel filtration on Sepharose 6B, the binding protein was purified 1000-4000-fold from the original liver homogenate. When the partially purified preparation was chromatographed on Sepharose 6B, the binding protein eluted as a molecule with an apparent molecular weight of 200000, with a Stokes' radius of 4.9 nm. Sucrose-density-gradient centrifugation of the preparation showed that the sedimentation coefficient of the binding protein was 7.2S. Isoelectric focusing experiments revealed that a major part of the protein has an acidic pI (4.2-4.5). Exposure of the protein to trypsin decreased the binding activity for human 125I-labelled-somatotropin or bovine 125I-labelled-somatotropin, whereas ribonuclease, deoxyribonuclease, phospholipase C or neuraminidase had little or no effect.
The effects of modification with N-bromosuccinimide on the binding of ligands to dihydrofolate reductase
Authors: Thomson J W , Roberts G C , Burgen A S
Published: August 2015

The binding constants of substrate, inhibitors and coenzymes to native Lactobacillus casei dihydrofolate reductase and to the enzyme modified (at Trp-21) by N-bromosuccinimide have been determined using fluorimetric and spectrophotometric methods. The modification leads to only modest decreases (factors of 2-4) in the binding of substrate or substrate analogues, but the effects of coenzyme binding are much larger. The binding of NADPH is decreased by a factor of 200, but that of NADP+ by only a factor of 4, indicating a clear difference in their mode of interaction with the enzyme. The nature of this difference is discussed in the light of crystallographic and n.m.r. studies of the enzyme.
Acetyl-coenzyme A carboxylase from avocado (Persea americana) plastids and spinach (Spinacia oleracea) chloroplasts
Authors: Mohan S B , Kekwick R G
Published: August 2015

Preparations of acetyl-CoA carboxylase [acetyl-CoA-carbon-dioxide ligase (ADP-forming), EC 6.4.1.2] have been obtained from the plastids of avocado (Persea americana) fruit mesocarp and from spinach (Spinacia oleracea) chloroplasts. Both preparations required bovine serum albumin, HCO3-, citrate and glycerol for stabilization. The molecular weight of the avocado enzyme was about 6.5 × 10(5) on the basis of 1 mol of biotin/mol of enzyme, the behaviour of both enzymes on gel filtration being in accord with such a value. Removal of the stabilizing bovine serum albumin resulted in the loss of a biotin-containing fragment from the avocado enzyme. Citrate stabilized the enzyme at 10 mM and activated it optimally at 3.0 mM, effecting an approx. 2-fold increase in Vmax. It is suggested that in vivo the enzyme may be located within the chloroplast lamellae.
A stopped-flow study of the cupric ion oxidation of adult-human haemoglobin
Authors: Brittain T
Published: August 2015

The addition of excess Cu2+ to adult human haemoglobin leads to the production of alpha 2(2+) beta 2(3+), in both the oxy and deoxy forms of the protein. Stopped-flow studies of the oxidation process yields apparent second-order rate constants of 196M-1 X S-1 and 41M-1 X S-1 for the deoxy and oxy forms respectively. The rate of the deoxy-form oxidation is linearly dependent on [Cu2+], whereas that of the oxy form is rate-limited above 2 mM to 0.11 S-1. Arrhenius activation energies of the two processes are almost identical at 91 kJ X mol-1, as are the activation enthalpies of 89 kJ X mol-1. The activation entropies show small differences, being 31 entropy units and 48 entropy units for the oxy and deoxy forms respectively. ATP and glycerate 2,3-bisphosphate at saturating concentrations do not affect the rate of oxidation of the oxy form, but halve the rate found for the deoxy form. These data are discussed in terms of the previously proposed mechanism of oxidation in which slow Cu2+ binding is followed by rapid electron transfer.
Glutamine release from hindlimb and uptake by kidney in the acutely acidotic rat
Authors: Schröck H , Cha C J M , Goldstein L
Published: August 2015

The arteriovenous difference (release) for glutamine across the hindlimb increases significantly during acute HCl-induced acidosis. This additional amount release by muscle tissue can account for the extra glutamine taken up by the kidney.