bluuebunny/crossref_metadata_2025 huggingface dataset:
Properties of mitochondria isolated from cyanide-sensitive and cyanide-stimulated cultures of Acanthamoeba castellanii
Authors: Edwards Steven W. , Lloyd David
Published: August 2015

1. Mitochondria isolated from cultures of Acanthamoeba castellanii exhibit respiratory control and oxidize α-oxoglutarate, succinate and NADH with ADP:O ratios of about 2.4, 1.4 and 1.25 respectively. 2. Mitochondria from cultures of which the respiration was stimulated up to 50% by 1mm-cyanide (type-A mitochondria) and from cyanide-sensitive cultures (type-B mitochondria) had similar respiratory-control ratios and ADP:O ratios. 3. State-3 rates of respiration were generally more cyanide-sensitive than State-4 rates, and the respiration of type-A mitochondria was more cyanide-resistant than that of type-B mitochondria. 4. Salicylhydroxamic acid alone had little effect on respiratory activities of either type of mitochondria, but when added together with cyanide, irrespective of the order of addition, inhibition was almost complete. 5. Oxidation of externally added NADH by type-A mitochondria was mainly via an oxidase with a low affinity for oxygen (Km[unk]15μm), which was largely cyanide-sensitive and partially antimycin A-sensitive; this electron-transport pathway was inhibited by ADP. 6. Cyanide-insensitive but salicylhydroxamic acid-sensitive respiration was stimulated by AMP and ADP, and by ATP after incubation in the presence of MgCl2. 7. Addition of rotenone to mitochondria oxidizing α-oxoglutarate lowered the ADP:O ratios by about one-third and rendered inhibition by cyanide more complete. 8. The results suggest that mitochondria of A. castellanii possess branched pathways of electron transport which terminate in three separate oxidases; the proportions of electron fluxes via these pathways vary at different stages of growth.
Heterogeneity of lysosomes originating from rat liver parenchymal cells. Metabolic relationship of subpopulations separated by density-gradient centrifugation
Authors: Pertoft H , Wärmegård B , Höök M
Published: August 2015

1. A crude lysosomal fraction obtained by differential centrifugation of a rat liver homogenate was subjected to zonal centrifugation in iso-osmotic self-generating gradients composed of modified colloidal silica (Percoll). Analysis of relevant marker-enzyme activities shows a continuous band of considerably purified lysosomal particles in the density range 1.04–1.11 g/ml. 2. A relationship between age and buoyant density of the parenchymal lysosomal subpopulations is indicated by the distribution of 125I-labelled asialoglycoproteins in the heterogeneous lysosomes during the catabolism of the glycoprotein. The labelled asialoglycoprotein first appeared in lysosomal particles of low density, which with time progressively acquired a higher density. Furthermore, 30 min after administration the 125I-labelled asialocaeruloplasmin recovered in the light lysosomes was less degraded than the material recovered in the heavy lysosomes. 3. A lysosomal enzyme (arylsulphatase) was found to possess considerably higher isoelectric points in the heavy lysosomes than in the light lysosomes, which is consistent with a relationship between age and density of the lysosomes.
Is a urea cycle present in insects?
Authors: Pant R , Kumar S
Published: August 2015

1. The presence of appreciable activity of the urea-cycle enzymes in the tissues of Sarcophaga ruficornis, a carnivorous dipteran insect, all through its life-cycle appears significant in view of their total absence barring arginase (L-arginine ureohydrolase, EC 3.5.3.1) in the phytophagous lepidopteran eri silkwork Philosamia ricini at any stage of development. Further, the variation of all these enzymes all through its development suggests the possibility of the operation of the Krebs-Henseleit urea cycle in this carnivorous insect. 2. The almost parallel behaviour of arginase and ornithine delta-transaminase (L-ornithine-2-oxo acid aminotransferase, EC 2.6.1.13) in both the insects suggests another important role of the former in proline biosynthesis in insects. 3. High proteolytic activity accompanied with significant protein depletion and simultaneous increase in arginine is suggestive of the degradation of proteins and peptides.
The mode of regulation of pyruvate dehydrogenase of lactating rat mammary gland. Effects of starvation and insulin
Authors: Baxter Michael A. , Coore Haldane G.
Published: August 2015

1. The ‘initial activity’ of the pyruvate dehydrogenase enzyme complex in whole tissue or mitochondrial extracts of lactating rat mammary glands was greatly decreased by 24 or 48h starvation of the rats. Injection of insulin and glucose into starved rats 60min before removal of the glands abolished this difference in ‘initial activities’. 2. The ‘total activity’ of the enzyme complex in such extracts was revealed by incubation in the presence of free Mg2+ and Ca2+ ions (more than 10 and 0.1mm respectively) and a crude preparation of pig heart pyruvate dehydrogenase phosphatase. Starvation did not alter this ‘total activity’. It is assumed that the decline in ‘initial activity’ of the enzyme complex derived from the glands of starved animals was due to increased phosphorylation of its α-subunit by intrinsic pyruvate dehydrogenase kinase. 3. Starvation led to an increase in intrinsic pyruvate dehydrogenase kinase activity in both whole tissue and mitochondrial extracts. Injection of insulin into starved animals 30min before removal of the lactating mammary glands abolished the increase in pyruvate dehydrogenase kinase activity in whole-tissue extracts. 4. Pyruvate (1mm) prevented ATP-induced inactivation of the enzyme complex in mitochondrial extracts from glands of fed animals. In similar extracts from starved animals pyruvate was ineffective. 5. Starvation led to a decline in activity of pyruvate dehydrogenase phosphatase in mitochondrial extracts, but not in whole-tissue extracts. 6. These changes in activity of the intrinsic kinase and phosphatase of the pyruvate dehydrogenase complex of lactating rat mammary gland are not explicable by current theories of regulation of the complex.
Disialoganglioside GDla of rat brain subcellular particles during development
Authors: Yusuf H K M , Dickerson J W T
Published: August 2015

The increase observed in the amount of the disialoganglioside GDlof the rat cerebrum during development between 21 and 81 days of age accounted for nearly 40% of the overall increase in total ganglioside in the tissue during the same period. Subcellular fractionation showed the microsomal fraction to contribute by far the most towards this increase in Cerebral ganglioside GDla. It is suggested that microsomal ganglioside GDla may serve as a marker for dendritic arborization in the rat cerebrum.
The involvement of phosphatidate phosphohydrolase and phospholipase A activities in the control of hepatic glycerolipid synthesis. Effects of acute feeding with glucose, fructose, sorbitol, glycerol and ethanol
Authors: Sturton R G , Pritchard P H , Han L Y , Brindley D N
Published: August 2015

Rats were fed by stomach tube with a single dose of glucose, sorbitol, fructose, glycerol or ethanol of equivalent energy contents or with 0.15 M-NaCl. They were killed 6 h later and the relative rates of phosphatidate deacylation and dephosphorylation measured in the microsomal and supernatant fractions of the livers. Treatment with sorbitol, fructose, glycerol and ethanol increased phosphohydrolase activities in the microsomal and supernatant fractions. The only significant change in deacylase activity was an increase in the microsomal fraction produced by ethanol. It is proposed that hepatic triacylglycerol synthesis is partly controlled by the balance between phosphatidate phosphohydrolase and phospholipase A-type activities.
The interaction between mitochondrial NADH-ubiquinone oxidoreductase and ubiquinol-cytochrome c oxidoreductase. Evidence for stoicheiometric association
Authors: Ragan C I , Heron C
Published: August 2015

1. The NADH-ubiquinone oxidoreductase complex (Complex I) and the ubiquinol-cytochrome c oxidoreductase complex (Complex III) combine in a 1:1 molar ratio to give NADH-cytochrome c oxidoreductase (Complex I-Complex III). 2. Experiments on the inhibition of the NADH-cytochrome c oxidoreductase activity of mixtures of Complexes I and III by rotenone and antimycin indicate that electron transfer between a unit of Complex I-Complex III and extra molecules of Complexes I or III does not contribute to the overall rate of cytochrome c reduction. 3. The reduction by NADH of the cytochrome b of mixtures of Complexes I and III is biphasic. The extents of the fast and slow phases of reduction are determined by the proportion of the total Complex III specifically associated with Complex I. 4. Activation-energy measurements suggest that the structural features of the Complex I-Complex III unit promote oxidoreduction of endogenous ubiquinone-10.
Effect of zinc status of rats on the synthesis and degradation of copper-induced metallothioneins
Authors: Bremner I , Hoekstra W G , Davies N T , Young B W
Published: August 2015

Injection of Zn2+-adequate and Zn2+-deficient rats with Cu2+ stimulated the incorporation of l-[35S]cysteine into a low-molecular-weight Cu2+-binding protein in both liver and kidney. No significant incorporation of l-[4,5-3H]leucine into this protein occurred, confirming the previous claim that it was metallothionein and not some other leucine-rich protein. The half-life of the protein was found to be 16.9 +/- 1.0 (S.E.)h in the liver of Zn2+-adequate rats but only 12.3 +/- 0.5h in Zn2+-deficient animals. The degradation rate of the metallothionein was similar to the rate of disappearance of Cu2+ and Zn2+ from the protein, indicating that the release of mental from the protein and its catabolism occurred simultaneously. There was no significant difference in the half-lives of the hepatic or renal copper-thioneins in Zn2+-adequate rats.
Changes in the form of Arrhenius plots of the activity of glucagon-stimulated adenylate cyclase and other hamster liver plasma-membrane enzymes occurring on hibernation
Authors: Houslay M D , Palmer R W
Published: August 2015

1. Arrhenius plots of the glucagon-stimulated adenylate cyclase, 5′-nucleotidase, (Na+ + K+)-stimulated adenosine triphosphatase and Mg2+-dependent adenosine triphosphatase activities of control hamster liver plasma membranes exhibited two break points at around 25 and 13 degrees C, whereas Arrhenius plots of their activities in hibernating hamster liver plasma membranes exhibited two break points at around 25 and 4 degrees C. 2. A single break occurring between 25 and 26 degrees C was observed in Arrhenius plots of the activities of fluoride-stimulated adenylate cyclase, basal adenylate cyclase and cyclic AMP phosphodiesterase of liver plasma membranes from both control and hibernating animals. 3. Arrhenius plots of phosphodiesterase I activity showed a single break at 13 degrees C for membranes from control animals, and a single break at around 4 degrees C for liver plasma membranes from hibernating animals. 4. The temperature at which break points occurred in Arrhenius plots of glucagon- and fluoride-stimulated adenylate cyclase activity were decreased by about 7–8 degrees C by addition of 40 mm-benzyl alcohol to the assays. 5. Discontinuities in the Arrhenius plots of 4-anilinonaphthalene-1-sulphonic acid fluorescence occurred at around 24 and 13 degrees C for liver plasma membranes from control animals, and at around 25 and 4 degrees C for membranes from hibernating animals. 6. We suggest that in hamster liver plasma membranes from control animals a lipid phase separation occurs at around 25 degrees C in the inner half of the bilayer and at around 13 degrees C in the outer half of the bilayer. On hibernation a change in bilayer asymmetry occurs, which is expressed by a decrease in the temperature at which the lipid phase separation occurs in the outer half of the bilayer to around 4 degrees C. The assumption made is that enzymes expressing both lipid phase separations penetrate both halves of the bilayer, whereas those experiencing a single break penetrate one half of the bilayer only.
Protection of the pancreatic β-cell glucoreceptor mechanism for insulin secretion during culture in chemically defined medium
Authors: Gylfe Erik
Published: August 2015

High concentrations of glucose have a protective effect on the glucoreceptor mechanism for insulin secretion during culture of pancreatic islets in chemically defined media. To study at what level glucose exerts this effect, insulin secretion from β-cell-rich mouse pancreatic islets was measured before and after culture for 1 week in the presence of different substances. Before culture, glucose and inosine were potent stimulators, mannose and fructose were less potent and xylitol had no effect on secretion. Culture in 3mm-glucose resulted in a 10-fold decrease in the insulin response to glucose stimulation. A less marked decrease was noted after culture in 20mm- or 30mm-glucose. Inosine-stimulated secretion was much decreased after culture in high concentrations of glucose, whereas the responses to mannose or fructose were unchanged. After culture in 30mm-mannose, glucose-stimulated secretion was similar to that observed after culture in high concentrations of glucose, whereas the response to mannose had much decreased. There were no secretory responses to glucose or fructose after culture in 30mm-fructose, or to glucose or xylitol after culture in 30mm-xylitol. Culture in 10mm-inosine did not preserve any significant response to glucose or inosine. The insulin contents of islets and culture media were higher after culture in high concentrations of glucose, mannose or inosine than after culture in fructose, xylitol or low concentrations of glucose. It is suggested that glucose, and to some extent mannose, preserves the glucoreceptor mechanism for insulin secretion by influencing an early stage in glucose metabolism, presumably glucokinase activity.
Control of sterol metabolism in rat adrenal mitochondria
Authors: Mason J. Ian , Arthur John R. , Boyd George S.
Published: August 2015

Steroidogenesis by adrenal mitochondria from endogenous precursors is stimulated by corticotropin (ACTH) and is sensitive to the protein-synthesis inhibitor cycloheximide. In the present investigation the effect of cycloheximide treatment on the metabolism of a number of analogues of the normal steroidogenic substrate, i.e. cholesterol, by rat adrenal mitochondria was studied. It was observed that the metabolism of analogues such as desmosterol, 26-norcholest-5-en-3β-ol and 5-cholen-3β-ol (that is with non-polar alkyl side chains like cholesterol), was sensitive to cycloheximide treatment. By contrast, the metabolism of those analogues with polar groupings on the side chain, i.e., 20α-, 24-, 25- and 26-hydroxycholesterols was insensitive to pretreatment with cycloheximide. The binding of added sterol to the cytochrome P-450 component of the mitochondrial sterol desmolase was studied. Similar studies on the equilibration time on addition of exogenous sterols to achieve maximum rates of pregnenolone production were also made. Both studies show that cholesterol, a non-polar sterol, penetrated slowly through the mitochondrial milieu to reach the cytochrome P-450 reaction centre whereas 24- and 26-hydroxycholesterols rapidly attained the enzymic environment. The cycloheximide-sensitive process in sterol metabolism appeared related to the transfer of non-polar sterols such as cholesterol within the mitochondria to a region in close proximity to the enzyme. The importance, and possible mechanism of action, of the cycloheximide-sensitive factor in the control of adrenal steroidogenesis is discussed.
Enhancement of the viscosity of mucin by serum albumin
Authors: List S J , Findlay B P , Forstner G G , Forstner J F
Published: August 2015

The interaction of serum albumin with a model epithelial mucin from pig stomach was explored by rotary viscometry. During 30 min of incubation of human serum albumin(20mg/ml) and pig gastric mucin (8mg/ml) in iso-osmotic buffers at 37 degrees C, the solution became markedly viscous. Viscosity enhancement was proportional to albumin concentration (2-40mg/ml), was most pronounced under conditions of low shear rate (less than 45S-1), and was considerably greater than the additive or multiplicative viscosity values calculated from albumin or mucin solutions measured separately. The viscous mucin-albumin complex was destroyed by high shear rates (greater than 90S-1), but slowly re-formed under zero shear conditions. Elevation of pH (7 to 9), ionic strength (0.1 to 1.0), and addition of disodium EDTA (5mM) did not cause marked or specific alterations in the viscosity of the mixture, suggesting that electrostatic interactions probably do not stabilize mucin-albumin complexes. Urea (7M) and heating (35 to 55 degrees C) caused a major increase in the viscosity of mucin and mucin-albumin mixtures, suggesting that rupture of hydrogen bonds, unfolding and partial denaturation of mucin promotes greater intertangling (possibly hydrophobic interactions) between mucin and albumin molecules. The implications of mucin-albumin interaction in diseases associated with mucus obstruction are briefly discussed.
Purification and properties of β-N-acetylhexosaminidase from the mollusc Helicella ericetorum Müller
Authors: Calvo P , Reglero A , Cabezas J A
Published: August 2015

1. A beta-N-acetylhexosaminidase was purified 330-fold from the digestive gland of the terrestrial mollusc Helicella ericetorum Müller. 2. Its pH optimum is 4.5 for both beta-N-acetylglucosaminidase and beta-N-acetylgalactosaminidase activities in two buffer solutions; it is fully stable at 37 degrees C for 2h in the pH range 3.8–4.6 and shows one isoelectric point (pH 4.83). 3. The estimated mol.wt. is between 120,000 and 145,000. 4. The enzyme shows an endo-beta-N-acetylhexosaminidase activity on natural substrates such as ovalbumin, ovomucoid, chondroitin 4-sulphate, chitin and hyaluronic acid. 5. Two forms of the enzyme were separated by preparative polyacrylamide-gel electrophoresis. 6. Km and Vmax. for p-nitrophenyl 2-acetamido-2-deoxy-beta-D-glucopyranoside and p-nitrophenyl 2-acetamide-2-deoxy-beta-D-galactopyranoside are 0.43 mM, 30.1 micronmol of p-nitrophenol/min per mg and 0.19 mM, 8.6 micronmol of p-nitrophenol/min per mg respectively. 7. It is inhibited by Hg2+, Fe3+, acetate, some lactones, N-acetylgalactosamine, N-acetylglucosamine and mannose. 8. Mixed-substrates analysis and Ki values for competitive inhibitors indicated that beta-N-acetylglucosaminidase and beta-N-acetylgalactosaminidase activities are catalysed by the enzyme at the same active site.
Partial reversal of the acetaldehyde and butyraldehyde oxidation reactions catalysed by aldehyde dehydrogenases from sheep liver
Authors: Hart G J , Dickinson F M
Published: August 2015

In the presence of acetic anhydride or butyric anhydride, liver aldehyde dehydrogenases catalyse the oxidation of NADH at pH 7.0 and 25 degrees C. The maximum velocities and Michaelis constants for NADH at saturating anhydride concentrations are independent of which anhydride is used, the values being V′max. = 12 min-1 and Km for NADH = 9 micrometer for the mitochondrial enzyme and V′max = 25 min-1 and Km for NADH = 20 micrometer for the cytoplasmic enzyme. Substitution of [4A-2H]NADH for NADH resulted in 2-fold and 4-fold decreases in rate for the mitochondrial and cytoplasmic enzymes respectively.
Purification and properties of peroxisomal pyruvate (glyoxylate) aminotransferase from rat liver
Authors: Noguchi T , Takada Y
Published: August 2015

Pyruvate (glyoxylate) aminotransferase from rat liver peroxisomes was highly purified and characterized. The enzyme preparation has a mol.wt. of approx. 80,000 with two identical subunits, and isoelectric point of 8.0 and a pH optimum between 8.0 and 8.5. The enzyme catalysed transamination between a number of L-amino acids and pyruvate or glyoxylate. The effective amino acceptors were pyruvate, phenylpyruvate and glyoxylate with serine, and glyoxylate and phenylpyruvate with alanine as amino donor. These properties and kinetic parameters of the enzyme are remarkably similar to those previously described for mitochondrial alanine-glyoxylate aminotransferase isoenzyme 1 from glucagon-injected rat liver [Noguchi, Okuno, Takada, Minatogawa, Okai & Kido (1978, Biochem. J. 169, 113-122].
Isolation, characterization and distribution of adenosine 3′:5′-cyclic monophosphate from Pinus radiata
Authors: Wilson T , Moustafa E , Renwick A G C
Published: August 2015

Cyclic AMP was extracted in 0.1 M-HCl from tissues of Pinus radiata and purified by gel filtration on Sephadex G-10, and chromatography on Dowex AG1 (X2) and polyethyleneimine-cellulose in two separate solvent systems. Presumptive cyclic AMP from 10kg batches of pine needles was characterized by countercurrent distribution in the presence of cyclic [8-3H]AMP. Statistical analysis of the curves for radioactivity and mass (determined by the Gilman competitive-binding assay) showed that the fit of the curves was highly significant for seven degrees of freedom. The distribution of cyclic AMP within P. radiata and various other plant tissues was determined by the Gilman procedure. The results suggest that there is no relationship between variations in cyclic AMP concentrations and the known function of the tissue in which it was measured.
A possible role for 5-hydroxytryptamine as a mediator for calcitonin actions on the gastrointestinal tract and pancreas in rats
Authors: Nakhla A M , Latif A
Published: August 2015

Administration of pig calcitonin (10 M.R.C. units/kg body wt.) produced an immediate decrease in 5-hydroxytryptamine content in the antrum-duodenum region and ileum. In contrast, the hormone treatment rapidly increased the concentration of 5-hydroxytryptamine in the blood and pancreas. Serum immunoreactive gastrin did not change over a period of 3h after calcitonin administration. The results suggest that the release of 5-hydroxytryptamine probably mediates calcitonin actions on the gastrointestinal tract and pancreas.
Control of rat skeletal-muscle phosphorylase phosphatase activity by adrenaline
Authors: Tao S H , Huang F L , Lynch A , Glinsmann W H
Published: August 2015

Administration of adrenaline to an isolated rat hindlimb preparation rapidly decreased muscle phosphorylase phosphatase (EC 3.1.3.17) activity and increased heat-stable and trypsin-labile phosphatase inhibitor activity. This was associated with increased tissue cyclic AMP concentrations, phosphorylase (EC 2.4.1.1) activation and glycogen synthase (EC 2.4.1.11) inactivation.
A rapid method for the preparation of relatively pure metabolically competent synaptosomes from rat brain
Authors: Booth R F G , Clark J B
Published: August 2015

A rapid (less than 2h) method is described for the preparation of synaptosomes from rat brain by using a discontinuous Ficoll/sucrose gradient by a flotation technique. These synaptosomes are metabolically active and minimally (less than 5%) contaminated with ‘free’ mitochondria as judged by marker-enzyme assays and electron microscopy.
Inhibition of bacterial transport by uncouplers of oxidative phosphorylation. Effects of pentachlorophenol and analogues in Bacillus subtilis
Authors: Nicholas R A , Ordal G W
Published: August 2015

Analogues of the potent uncoupler of oxidative phosphorylation pentachlorophenol were tested as inhibitors of proline and glycine transport by Bacillus subtilis. These analogues included less highly substituted chlorophenols and pentachlorothiophenol. Like pentachlorophenol, they are non-competitive inhibitors of proline transport and uncompetitive inhibitors of glycine transport. However, the less highly substituted chlorophenols are weaker acids than pentachlorophenol and also weaker inhibitors. Analysis indicated that the anionic form of the uncouplers is the inhibiting species. Pentachlorothiophenol, a water-insoluble anion, is also a potent inhibitor. These results support previous studies that concluded that uncouplers of oxidative phosphorylation inhibit amino acid transport by binding at specific sites on proteins, the free energy of interaction stabilizing ‘unproductive’ conformations. Such specific interactions of uncoupler with protein are probably commonplace.
Endocytosis of sulphated proteoglycans by cultured skin fibroblasts
Authors: Prinz R , Schwermann J , Buddecke E , von Figura K
Published: August 2015

1. Human skin fibroblasts internalize homologous sulphated proteoglycans by adsorptive endocytosis. Endocytosis rate is half maximal when the concentration of the proteoglycans is 0.1 nM. At saturation, a single fibroblast may endocytose up to 8 × 10(6) proteoglycan molecules/h. 2. The kinetics of prote;glycan binding to the cell surface suggest the presence of 6 × 10(5) high-affinity binding sites per cell. The bulk of sulphated proteoglycans associates to low-affinity binding sites on the cell surface. 3. Glycosaminoglycans and other anionic macromolecules inhibit endocytosis of sulphated proteoglycans non-competitively. The lack of interaction of glycosaminoglycans with the cell-surface receptors for sulphated proteoglycans suggests that the protein core of proteoglycans is essential for binding to the cell surface. 4. The effects of trypsin, cell density, serum concentration and medium pH on endocytosis and degradation of endocytosed sulphated proteoglycans is described. 5. A comparison of the number of the high-affinity binding sites and the number of molecules endocytosed with respect to time suggests a recycling of the proteoglycan receptors between the cell surface and the endocytotic vesicles and/or the lysosomes.
Photosensitivity of respiration in Neurospora mitochondria. A protective role for carotenoid
Authors: Ramadan-Talib Z , Prebble J
Published: August 2015

1. The effect of visible light on respiratory activity was studied in two strains of Neurospora crassa, one a wild-type strain able to synthesize carotenoid and the other an albino mutant lacking carotenogenic activity. Light had no effect on growth under the conditions studied, but inhibited respiration of hyphal suspensions. the degree of inhibition being dependent on the carotenoid content of the hyphae. 2. In studies of respiration of isolated mitochondria, three types of photosensitive site were detected. These were the flavo-protein dehydrogenases themselves, a site separate from the latter also associated with the dehydrogenase but re-activatable by treatment with a thiol reagent, and the respiratory quinone, ubiquinone. Cytochrome oxidase, previously reported as photosensitive from many sources, was not appreciably affected by light in these preparations. 3. The degree of inactivation of the respiratory quinone was dependent on the amount of carotenoid in the preparation, high concentrations of the pigment in the mitochondrial membranes providing substantial protection against the effect of light. 4. Separation of the inner and outer membranes of mitochondria showed that under conditions where carotenoid appears to protect quinone, significant amounts are found in the inner mitochondrial membrane, oterhwise carotenoid is restricted to the outer membrane.
Responsiveness to glucagon by isolated rat hepatocytes controlled by the redox state of the cytosolic nicotinamide—adenine dinucleotide couple acting on adenosine 3′:5′-cyclic monophosphate phosphodiesterase
Authors: Clark M G , Jarrett I G
Published: August 2015

1. The effects of changes in the cytoplasmic [NADH]/[NAD+] ratio on the efficacy of glucagon to alter rates of metabolism in isolated rat hepatocytes were examined. 2. Under reduced conditions (with 10mM-lactate), 10nM-glucagon stimulated both gluconeogenesis and urea synthesis in isolated hepatocytes from 48h-starved rats; under oxidized conditions (with 10mM-pyruvate), 10nM-glucagon had no effect on either of these rates. 3. The ability of glucagon to alter the concentration of 3′:5′-cyclic AMP and the rates of glucose output, glycogen breakdown and glycolysis in cells from fed rats were each affected by a change in the extracellular [lactate]/[pyruvate] ratio; minimal effects of glucagon occurred at low [lactate]/[pyruvate] ratios. 4. Dose-response curves for glucagon-mediated changes in cyclic AMP concentration and glucose output indicated that under oxidized conditions the ability of glucagon to alter each parameter was decreased without affecting the concentration of hormone at which half-maximal effects occurred. 5. The phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine (0.05 mM) significantly reversed the inhibitory effects of pyruvate on glucagon-stimulated glucose output. 6. For exogenously added cyclic [3H]AMP(0.1 mM), oxidized conditions decreased the stimulatory effect on glucose output as well as the intracellular concentration of cyclic AMP attained, but did not alter the amount of cyclic [3H]AMP taken up. 7. The effects of lactate, pyruvate, NAD+ and NADH on cyclic AMP phosphodiesterase activities of rat hepatocytes were examined. 8. NADH (0.01–1 MM) inhibited the low-Km enzyme, particularly that which was associated with the plasma membrane. 9. The inhibition of membrane-bound cyclic AMP phosphodiesterase by NADH was specific, reversible and resulted in a decrease in the maximal velocity of the enzyme. 10. It is proposed that regulation of the membrane-bound low-Km cyclic AMP phosphodiesterase by nicotinamide nucleotides provides the molecular basis for the effect of redox state on the hormonal control of hepatocyte metabolism by glucagon.
Thyroid-hormone modulation of the number of β-adrenergic receptors in rat fat-cell membranes
Authors: Giudicelli Y
Published: August 2015

Adipocytes from thyroidectomized rats contain 3 times less [3H]dihydroalprenolol-binding sites (beta-adrenergic receptors) than adipocytes from euthyroid animals. This alteration is not solely due to cell-size differences, but also to a thyroidectomy-induced defect in beta-adrenergic receptor density per adipocyte surface area, a defect that is furthermore corrected by tri-iodothyronine treatment.
Some magnetic properties of Pseudomonas cytochrome oxidase
Authors: Walsh T A , Johnson M K , Greenwood C , Barber D , Springall J P , Thomson A J
Published: August 2015

The magnetic properties of the haem groups of Pseudomonas cytochrome oxidase and its cyanide-bound derivatives were studied in both the oxidized and reduced states by means of m.c.d. (magnetic circular dichroism) at low temperatures. In addition, the oxidized forms of the enzyme were also investigated by e.p.r. (electron-paramagnetic-resonance) spectroscopy, and a parallel study, using both e.p.r. and m.c.d., was made on Pseudomonas cytochrome c-551 to aid spectral assignments. For ascorbate-reduced Pseudomonas cytochrome oxidase, the temperature-independence of those features in the m.c.d. spectrum corresponding to the haem c, and the temperature-dependence of those signals corresponding to the haem d1, showed the former to be low-spin and the latter to be high-spin (s = 2). However, addition of cyanide to the reduced enzyme gave a form of the protein that was completely low-spin. The e.p.r. and m.c.d. sectra of oxidized Pseudomonas cytochrome oxidase and its cyanide derivative were consistent with the haem c and d1 components being low-spin in both cases. Pseudomonas cytochrome c-551 was found to be low-spin in both its oxidized and reduced redox states.
Derivation of molecular pK values from pH-dependences
Authors: Dixon H B F
Published: August 2015

If the titration pK values of a dibasic acid are called pK +/- logp, then its molecular pK values are pK +/- log(p+1/p). If the pH values at which the concentration of its monoprotonated form is half-maximal are called pK +/- logq, then its molecular pK values are pK +/- log(q-4+1/q).
Properties of the cupric sites in bovine superoxide dismutase studied by nuclear-magnetic-relaxation measurements
Authors: Boden N , Holmes M C , Knowles P F
Published: August 2015

Proton nuclear-relaxation rates have been measured as a function of frequency, temperature, pH and cyanide concentration in aqueous solutions of superoxide dismutase from bovine erythrocytes. The results show that, whereas for pH less than or equal to 9 only one water molecule is bound to each Cu2+ ion, at higher pH a second co-ordination site for OH- becomes available; it is proposed that this involves cleavage of the bond between Cu2+ and the histidine residue that bridges to Zn2+.
Kinetic studies of dogfish liver glutamate dehydrogenase
Authors: Electricwala A H , Dickinson F M
Published: August 2015

Initial-rate studies were made of the oxidation of L-glutamate by NAD+ and NADP+ catalysed by highly purified preparations of dogfish liver glutamate dehydrogenase. With NAD+ as coenzyme the kinetics show the same features of coenzyme activation as seen with the bovine liver enzyme [Engel & Dalziel (1969) Biochem. J. 115, 621–631]. With NADP+ as coenzyme, initial rates are much slower than with NAD+, and Lineweaver–Burk plots are linear over extended ranges of substrate and coenzyme concentration. Stopped-flow studies with NADP+ as coenzyme give no evidence for the accumulation of significant concentrations of NADPH-containing complexes with the enzyme in the steady state. Protection studies against inactivation by pyridoxal 5′-phosphate indicate that NAD+ and NADP+ give the same degree of protection in the presence of sodium glutarate. The results are used to deduce information about the mechanism of glutamate oxidation by the enzyme. Initial-rate studies of the reductive amination of 2-oxoglutarate by NADH and NADPH catalysed by dogfish liver glutamate dehydrogenase showed that the kinetic features of the reaction are very similar with both coenzymes, but reactions with NADH are much faster. The data show that a number of possible mechanisms for the reaction may be discarded, including the compulsory mechanism (previously proposed for the enzyme) in which the sequence of binding is NAD(P)H, NH4+ and 2-oxoglutarate. The kinetic data suggest either a rapid-equilibrium random mechanism or the compulsory mechanism with the binding sequence NH4+, NAD(P)H, 2-oxoglutarate. However, binding studies and protection studies indicate that coenzyme and 2-oxoglutarate do bind to the free enzyme.
A four-straight-line model for the proteinase-binding characteristics of human blood serum
Authors: Topping R M , Seilman S
Published: August 2015

Kinetic evaluation of the capacity of human blood serum to form complexes with bovine trypsin generated partition profiles that may be approximated by a series of four intersecting straight lines. Such profiles are suggested to reflect the binding of trypsin to alpha 2-macroglobulin in a kinetically preferred mode (alpha-sites), followed by a subsidiary mode (beta-sites) and finally to alpha 1-antitrypsin. The form of the profile, in addition to revealing a hitherto unreported proteinase-binding capability of alpha 2-macroglobulin (beta-sites), also indicates that saturation of alpha-sites corresponds to a molar binding ratio of alpha 2-macroglobulin/trypsin of 1:2. Finally the profile provides, for certain pathological states, a clinically valuable characteristic.
Structure and synthesis of naturally occurring anhydrovitamin A2
Authors: Barua A B , Verma K , Das S R
Published: August 2015

The structure of naturally occurring anhydrovitamin A2 was elucidated as 3-hydroxyanhydroretinol from study of its u.v.–visible, i.r., n.m.r. and mass spectra. The structure has been confirmed by a chemical synthesis. Saccobranchus fossils, a freshwater fish, can convert 3-hydroxyretinol into 3-hydroxyanhydroretinol, which in turn is converted into-3-dehydroretinol. Chemical conversion of 3-hydroxyretinol into 3-dehydroretinol was also carried out.
Effects of food deprivation on ketonaemia, ketogenesis and hepatic intermediary metabolism in the non-lactating dairy cow
Authors: Baird G D , Heitzman R J , Reid I M , Symonds H W , Lomax M A
Published: August 2015

1. The aim of this work was to investigate why non-lactating dairy cows are less susceptible to the development of ketonaemia during food deprivation than are dairy cows in early lactation. 2. The first experiment (Expt. A) consisted of determining the effect of 6 days of food deprivation on the concentrations of ketone bodies, and of metabolites related to the regulation of ketogenesis, in jugular blood and liver of non-lactating cows. 3. During the food deprivation, blood ketone-body concentrations rose significantly, but to a value that was only 16% of that achieved in lactating cows deprived of food for 6 days [Baird, Heitzman & Hibbitt (1972) Biochem. J. 128, 1311–1318]. 4. In the liver, food deprivation caused: a rise in ketone-body concentrations; a fall in the concentration of glycogen and of various intermediates of the Embden-Meyerhof pathway and the tricarboxylic acid cycle; an increase in cytoplasmic reduction; a decrease in the [total NAD+]/[total NADH] ratio; a decrease in energy charge. These changes were all qualitatively similar to those previously observed in the livers of the food-deprived lactating cows. 5. There appeared therefore to be a discrepancy in the food-deprived non-lactating cows between the absence of marked ketonaemia and the occurrence of metabolic changes within the liver suggesting increased hepatic ketogenesis. This discrepancy was partially resolved in Expt. B by the observation in two catheterized non-lactating cows that, although there was a 2-fold increase in hepatic ketogenesis during 6 days of food deprivation, ketogenesis from the splanchnic bed as a whole (i.e. gut and liver combined) declined slightly owing to cessation of gut ketogenesis.
Glucose transport of Escherichia coli growing in glucose-limited continuous culture
Authors: Hunter I S , Kornberg H L
Published: August 2015

Dilute cultures of wild-type Escherichia coli K12 and of derivatives impaired in one or other Enzyme-II component of the glucose phosphotransferase system were grown in continuous culture under glucose limitation. Cells harvested from the chemostat took up [U-14C]glucose from 0.1 mM solutions at rates directly related to the rates at which those cells had grown; the activity of the phosphotransferase system in those cells, rendered permeable with optimal accounts of toluene, parallels the ability of the cells to take up glucose. The capacity of these systems was rate-limiting for growth under the negligibly low glucose concentration in the chemostat, but was adequate to account for the stimulation of respiration observed when the cells were presented suddenly with excess glucose.
Polarization of chlorotetracycline fluorescence in pancreatic islet cells and its response to calcium ions and D-glucose
Authors: Täljedal I B
Published: August 2015

Suspensions rich in pancreatic beta-cells were prepared from non-inbred ob/ob-mice, incubated with 10 micrometer-chlorotetracycline, and analysed for fluorescence polarization in a microscope. Throughout the temperature range 16–38 degrees C, fluorescence was enhanced by 5 mM-Ca2+ in the incubation medium; 20 mM-D-glucose decreased the fluorescence measured in the presence of Ca2+. Fluorescence showed a curvilinear negative regression on temperature. The curves were rectified to a virtually ideal degree by Arrhenius transformations of data. Non-parametric testing of differences between linearized regression lines forms the basis for the following conclusions. The temperature-dependence of fluorescence intensity appeared to be smaller for Ca2+-specific signals than for the background fluorescence of chlorotetracycline in Ca2+-deficient cells. D-Glucose significantly diminished the polarization of fluorescence in cells incubated with Ca2+. It is suggested that D-glucose increases the mobility of Ca2+ in beta-cell plasma membranes; this mobility increase may help to explain previously reported effects of D-glucose on 45Ca2+ fluxes and membrane electric potential.
Fungal metabolism of biphenyl
Authors: Dodge R H , Cerniglia C E , Gibson D T
Published: August 2015

Cunninghamella elegans grown on Sabouraud dextrose broth transformed biphenyl to produce 2-, 3- and 4-hydroxybiphenyl, as well as 4,4′-dihydroxybiphenyl as free phenols. A compound tentatively identified as 2,4′-dihydroxybiphenyl was also produced. When 4-hydroxybiphenyl or 2-hydroxybiphenyl replaced biphenyl as the substrate, C. elegans produced 4,4′-dihydroxybiphenyl and 2,5-dihydroxybiphenyl respectively. The compound identified as 2,4′-dihydroxybiphenyl was produced from both substrates. A survey of 11 species of fungi known to degrade hydrocarbons revealed two species that were comparable to C. elegans in their ability to convert biphenyl into free phenols. In addition to free phenolic metabolites, deconjugation experiments indicated that 44% of the known metabolites present in the culture filtrate were present in the form of conjugates. These results suggest that the transformation of biphenyl by C. elegans is similar to that found in mammalian systems.
Involvement of cytoskeletal structures in nerve-growth-factor-mediated induction of ornithine decarboxylase
Authors: Lakshmanan J
Published: August 2015

Induction of ornithine decarboxylase elicited in response to nerve-growth factor in target organs is greatly decreased by preincubation of these tissues with cytoskeletal poisons such as vinblastine, diamide, cytochalasin B and colchicine. These results are interpreted as evidence for the involvement of receptor-associated cytoskeletal structures in mediating the nerve-growth-factor-specific induction of ornithine decarboxylase.
The effects of lipid phase transitions on the interaction of mitochondrial NADH–ubiquinone oxidoreductase with ubiquinol–cytochrome c oxidoreductase
Authors: Heron C , Gore M G , Ragan C I
Published: August 2015

1. The endogenous phosphatidylcholine and phosphatidylethanolamine of Complexes I and III from bovine heart mitochondria may be completely replaced with 1,2-ditetradecanoyl-sn-glycero-3-phosphocholine with at least partial retention of activity. 2. The lipid-replaced enzymes associate in 1:1 molar ratio to give a Complex I–III unit catalysing NADH-cytochrome c oxidoreductase activity. 3. On increasing the concentration of ubiquinone-10 and the synthetic phospholipid, the lipid-replaced Complexes appear to operate independently of each other as in the natural membrane. Thus the lipid-replaced enzymes associate in exactly the same ways as the enzymes containing natural phospholipids. 4. Arrhenius plots of NADH–cytochrome c oxidoreductase activity reconstituted from lipid-replaced Complexes I and III exhibit changes in slope at 24 degrees C. When the concentrations of phospholipid and ubiquinone-10 are increased, the Arrhenius plots show discontinuities at 24 degrees C as well as changes in slope. 5. The kinetics of cytochrome b reduction by NADH were measured in mixtures containing 2 mol of Complex III/mol of Complex I. When the enzymes contained natural phospholipids. the reduction kinetics were biphasic. When the enzymes had been supplemented with further phospholipid and ubiquinone-10 the kinetics were monophasic. When lipid-replaced enzymes were supplemented with 1,2-ditetradecanoyl-sn-glycero-3-phosphocholine and ubiquinone-10, reduction of cytochrome b was monophasic above the phase-transition temperature of the lipid but biphasic below it. 6. These findings are interpreted in terms of the model for the interaction of Complexes in the natural membrane proposed by Heron, Ragan & Trum-power [(1978) Biochem. J. 174, 791–800].
The influence of vasopressin and related peptides on glycogen phosphorylase activity and phosphatidylinositol metabolism in hepatocytes
Authors: Kirk C J , Rodrigues L M , Hems D A
Published: August 2015

The relative abilities of seven vasopressin-like peptides to activate hepatic glycogen phosphorylase and stimulate phosphate incorporation into phosphatidylinositol were compared. Although the individual peptides differed in their potencies, the concentrations required to stimulate phosphatidylinositol metabolism were always greater (about 10 times) than those needed to activate phosphorylase. The molecular specificity of the hepatic vasopressin receptor and the role of vasopressin-stimulated phosphatidylinositol turnover are discussed.
Energy-dependence of calcium accumulation during sporulation of Bacillus megaterium KM
Authors: Hogarth C , Ellar D J
Published: August 2015

Ca2+ accumulation and endogenous respiration of sporulating Bacillus megaterium are inhibited to the same extent by electron-transport of inhibitors and the uncoupler carbonyl cyanide p-trifluoromethoxyphenylhydrazone, suggesting that Ca2+ is accumulated by an active transport process. Forespores isolated in stage V of sporulation demonstrated Ca2+-specific carrier-mediated Ca2+ uptake, consistent with downhill transfer [Hogarth & Ellar (1978) Biochem. J. 176, 197-203]. In the present studies forespore Ca2+ uptake was unaffected by carbonyl cyanide p-trifluoromethoxyphenylhydrazone and by concentrations of respiratory inhibitor that inhibited forespore endogenous respiration by 85%. These data suggest that Ca2+ enters the isolated forespore by facilitated diffusion. Ca2+ uptake into sporulating protoplasts was completely inhibited by concentrations of respiratory inhibitors that had no effect on either Ca2+ uptake or respiration of stage-V forespores, but which resulted in inhibition of mother-cell membrane NADH oxidase. These results indicate that the mother-cell membrane is a site for active transport of Ca2+ into the sporulating cell. The effects of the adenosine triphosphatase inhibitor dicyclohexylcarbodi-imide on mother-cell membrane adenosine triphosphatase, NADH oxidase and protoplast Ca2+ uptake were examined.
Effect of metabolic conditions on protein turnover in yeast
Authors: López S , Gancedo J M
Published: August 2015

1. In yeast growing on ethanol a turnover rate of up to 2%/h was measured. As much as 80% of the protein was subject to turnover, and no marked heterogeneity in the rate of degradation of protein was observed. When the yeast grew on glucose, the protein was degraded at a lower rate (0.5-1%/h). 2. Starvation for a nitrogen source increased the rate of protein degradation severalfold, whereas deprivation of phosphate had only a marginal effect (30% increase). Removal of glucose from a medium containing 50mM-phosphate did not cause marked changes in the rate of protein degradation. In contrast, when the media were low in phosphate (0.1 mM) removal of glucose increased the rate of turnover 2-4-fold. 3. Protein degradation proceeded unimpaired when the intracellular concentration of ATP decreased from 4 to 1 mM, but stopped completely when it decreased below 0.3 mM.
Calmodulin and myosin light-chain kinase of rabbit fast skeletal muscle
Authors: Nairn A C , Perry S V
Published: August 2015

1. It is confirmed that myosin light-chain kinase is a protein of mol.wt. about 80,000 that is inactive in the absence of calmodulin. 2. In the presence of 1 mol of calmodulin/mol of kinase 80-90% of the maximal activity is obtained. 3. Crude preparations of the whole light-chain fraction of rabbit fast-skeletal-muscle myosin contain enough calmodulin to activate the enzyme. A method for the preparation of calmodulin-free P light chain is described. 4. A procedure is described for the isolation of calmodulin from rabbit fast skeletal muscle. 5. Rabbit fast-skeletal-muscle calmodulin is indistinguishable from bovine brain calmodulin in its ability to activate myosin light-chain kinase. The other properties of these two proteins are also very similar. 6. Rabbit fast-skeletal-muscle troponin C was about 10% as effective as calmodulin as activator for myosin light-chain kinase. 7. By chromatography on a Sepharose-calmodulin affinity column evidence was obtained for the formation of a Ca2+-dependent complex between calmodulin and myosin light-chain kinase. 8. Troponin I from rabbit fast skeletal muscle and histone IIAS were phosphorylated by fully activated myosin light-chain kinase at about 1% of the rate of the P light chain.