bluuebunny/crossref_metadata_2025 huggingface dataset:
The effect of temperature on catalytic and regulatory functions of pyruvate kinases of the rainbow trout and the Antarctic fish Trematomus bernacchii
Authors: Somero G. N. , Hochachka P. W.
Published: August 2015

1. The effects of temperature on the catalytic and regulatory properties of pyruvate kinases from the temperate-zone rainbow trout and the Antarctic fish Trematomus bernacchii were examined. 2. The Km value of pyruvate kinase for one of its two substrates, phosphoenolpyruvate, is temperature-dependent, and is lowest at temperatures that closely coincide with the habitat temperatures of the two fishes. 3. Two regulatory functions of pyruvate kinase, feedforward activation by fructose diphosphate and feedback inhibition by ATP, are temperature-independent. Enzyme–ADP interaction is also temperature-independent. 4. It is concluded that enzyme–substrate and enzyme–modulator interactions are important factors in short-term and in evolutionary adaptations by poikilotherms to changes in temperature. Though the Km for substrate may vary in apparently adaptive manners, the regulatory functions of an enzyme appear to be unchanged over the range of temperatures experienced by the organism in Nature.
The distribution of the components of mixed-function oxidase between the rough and the smooth endoplasmic reticulum of liver cells
Authors: Holtzman J. L. , Gram T. E. , Gigon P. L. , Gillette J. R.
Published: August 2015

Mixed-function oxidase activity, when measured by the N-demethylation of ethylmorphine or the hydroxylation of aniline, is significantly higher in the smooth hepatic endoplasmic reticulum than in the rough. In the rabbit the smooth membrane/rough membrane activity ratios are significantly greater than 1 whether the activities are expressed per g. of liver (ratio 5), per mg. of protein (ratio 3–5), per μg. of phospholipid phosphorus (ratio 2), per unit of cytochrome P-450 (ratio 1·7) or per unit of NADPH–cytochrome c reductase activity (ratio 2). On the other hand, if the activities are normalized to the NADPH–cytochrome P-450 reductase, there is no significant difference between the rough and smooth membranes. These results suggest that, in the rabbit, the rate-limiting step is the reduction of cytochrome P-450. In contrast, in the rat the difference in activities can be explained by differences in the concentration of cytochrome P-450.
A new method for the quantitative determination of monosaccharides, amino sugars and N-acetylneuraminic acid and of 6-deoxyhexose (fucose) in the presence of other sugars
Authors: Brearley Gwyneth M. , Weiss Jacqueline B.
Published: August 2015

1. Monosaccharides, amino sugars and N-acetylneuraminic acid were determined by using an original colorimetric assay procedure, based on the detection of formaldehyde released after periodate oxidation. A range of these compounds was investigated by this method and they were all found to obey Beer's law within the concentration range 0–0·6μmole/ml. 2. A simple method for the determination of 6-deoxyhexose concentration in the presence of other monosaccharides is also described. 3. The optimum pH for the release of formaldehyde from sugars by periodate oxidation was 7·0–7·5. 4. The methods described have considerable advantages over existing assay systems and their particlar value in automatic colorimetry, where the use of concentrated acids is undesirable, is discussed.
The inhibition, by intermediary metabolites, of isocitrate lyase from Chlorella pyrenoidosa
Authors: John P. C. L. , Syrett P. J.
Published: August 2015

1. Intermediary metabolites, arising from the glyoxylate by-pass, were tested for their effect on the activity of isocitrate lyase, the first enzyme of the by-pass. 2. Oxaloacetate and pyruvate inhibited the enzyme when present at concentrations similar to that of the substrate. 3. Inhibition was competitive and did not depend on a non-optimum pH. The affinities determined at the pH optimum were: Ki (oxaloacetate), 3·7×10−5m; Ki (pyruvate), 6·6×10−5m. 4. The significance of the inhibitions is discussed with particular reference to the known inhibition by phosphoenolpyruvate.
The glycerol teichoic acid of walls of Staphylococcus lactis I3
Authors: Archibald A. R. , Baddiley J , Button D.
Published: August 2015

1. The teichoic acid from walls of Staphylococcus lactis I3 was isolated by extraction with trichloroacetic acid and shown to contain glycerol, N-acetylglucosamine, phosphate and d-alanine in the molecular proportions 1:1:2:1. The alanine is attached to the polymer through ester linkages. 2. Hydrolysis with acid gave alanine, glucosamine and glycerol diphosphates. Under mild acid conditions a repeating unit was produced; this consists of glycerol diphosphate joined through a phosphodiester group to N-acetylglucosamine. 3. Hydrolysis with alkali gave glycerol diphosphates, saccharinic acid and two phosphodiesters containing glucosamine whose structures were elucidated; these both contain glucosamine 1-phosphate, and N-acetylglucosamine 1-phosphate was isolated by a degradative procedure. 4. The unusual properties of the teichoic acid are explained by a polymeric structure in which N-acetylglucosamine 1-phosphate is attached through its phosphate to glycerol phosphate. 5. The biosynthetic implications of this structure are discussed.
The catalase–hydrogen peroxide system. Kinetics of catalatic action at high substrate concentrations
Authors: Jones Peter , Suggett A.
Published: August 2015

1. A re-examination of the catalase–hydrogen peroxide reaction at high substrate concentrations, by using the quenched-flow technique, reveals a more complex kinetic behaviour than that previously reported. At constant reaction time the catalatic process obeys Michaelis–Menten kinetics, but the apparent Michaelis constant is markedly time-dependent, whereas the conventional catalase activity is independent of time. 2. The kinetics of the ‘time effect’ were analysed and it is suggested that the effect derives from the formation of an inactive species (thought to be catalase Compound II). The process shows Michaelis–Menten kinetics, with a Michaelis constant equal to that for the catalatic reaction in the limit of zero reaction time. 3. It has been confirmed that certain buffer components have marked inhibitory effects on the catalatic reaction and that, in unbuffered systems, catalatic activity is substantially independent of pH in the range 4·7–10·5.
The catalase–hydrogen peroxide system. A theoretical appraisal of the mechanism of catalase action
Authors: Jones Peter , Suggett A.
Published: August 2015

1. The mechanisms of catalase action advanced by Jones & Wynne-Jones (1962) and by Nicholls (1964) are compared in terms of their relative plausibilities and their utility for extension to accommodate more recent experimental information. 2. A revised formal mechanism is advanced that avoids the less satisfactory features of these mechanisms and attempts to account for the roles of catalase sub-units in both reversible and irreversible deactivation phenomena. 3. Theoretical studies of the redox chemistry of peroxides are used to provide the basis for a discussion of the mechanism of the redox act in catalatic action at the molecular level. It is suggested that an important feature of catalase action may be a mediation of the formation of a reactive intermediate by stereospecifically located acid–base functions in the active site. 4. A more detailed statement of this concept is attempted in terms of a hypothetical partial molecular model for the composition and stereochemistry of the active site of catalase. The utility of this model in describing the catalatic and peroxidatic actions of catalase is assessed.
Turnover rates of ketone bodies in normal, starved and alloxan-diabetic rats
Authors: Bates Margaret W. , Krebs H. A. , Williamson D. H.
Published: August 2015

1. Rates of appearance and disappearance of total ketone bodies were determined in normal, starved and alloxan-diabetic rats by measuring specific radioactivities and concentrations of blood acetoacetate and 3-hydroxybutyrate at different times after injection of 3-hydroxy[14C]butyrate. 2. The mean rates of appearance were 1·7, 4·2 and 10·9μmoles/min./100g. body wt. respectively for normal, starved and alloxan-diabetic rats. The rates of disappearance were of the same order of magnitude as the rates of appearance. 3. There was a direct correlation between the rates of appearance and disappearance and the blood concentrations of the ketone bodies. 4. The results indicate that in the rat increased ketone-body production is paralleled by increased ketone-body utilization and that the raised ketone-body concentration in the blood in starvation and alloxan-diabetes is due to a slight imbalance between the rates of production and utilization. 5. The findings are discussed in relation to the concept that ketone bodies can serve as fuels of respiration when the supply of carbohydrate is limited.
The pyridoxal- and pyridoxal 5′-phosphate-catalysed non-enzymic degradations of l-serine O-sulphate and related compounds
Authors: Thomas J H , Dodgson K S , Tudball N.
Published: August 2015

1. l-Serine O-sulphate and l-threonine O-sulphate are degraded in the presence of pyridoxal 5′-phosphate to yield equimolar amounts of the corresponding keto acid, ammonia and sulphate. 2. Pyridoxal catalyses the same reactions at a faster rate. 3. One of a number of bi- or ter-valent metal ions must be present for these degradations to proceed. The reaction rates are dependent on a number of factors including pH and the nature of the metal ion used. 4. Studies with related sulphate esters indicate that the α-hydrogen atom and the amino group are essential for activity. 5. Spectral changes during the pyridoxal and pyridoxal 5′-phosphate catalysis of l-serine O-sulphate breakdown suggest the formation of a Schiff base. 6. The mechanism for these reactions appears to be in accordance with the general mechanism proposed for pyridoxal-catalysed αβ-elimination reactions.
Studies on vision. The nature of the retinal–opsin linkage
Authors: Akhtar M , Blosse P. T. , Dewhurst P. B.
Published: August 2015

1. Convenient methods for the preparation of tritiated 11-cis-retinol, 11-cis-retinal and rhodopsin are described. Irradiation of labelled rhodopsin in the presence of sodium borohydride resulted in the irreversible binding of the retinyl moiety to the active site. Degradative studies established that the retinyl moiety in this reduced derivative of rhodopsin was attached to the ∈-amino group of lysine. In connexion with this investigation the synthesis of a number of N-retinylidene- and N-retinyl-amino acids was achieved. Derivatives of lysine with the retinyl moiety attached either to the α-amino group or to the ∈-amino group were also synthesized and characterized. 2. It is suggested that the involvement of a charge-transfer interaction between the retinylidene chromophore and a suitable group −X or −X·H on the opsin best explains the spectroscopic properties of rhodopsin and other visual proteins.
Purification and properties of esterases characteristic of adult rat brain
Authors: Dabich Danica , Chakrapani Bhargavan , Syner Frank N.
Published: August 2015

1. A method for the partial purification of an esterase fraction, present in the brain of the adult but not the newborn rat, is described. A 54-fold purification was achieved in three steps. 2. When subjected to starch-gel electrophoresis, the purified fraction resolved into three bands of esterase activity. Two of these bands migrate close together and faster than other esterases in the brain. These two esterases are inhibited by p-hydroxymercuribenzoate but not by di-isopropyl phosphorofluoridate. The third band is di-isopropyl phosphorofluoridate-sensitive and migrates just behind the two leading esterases. 3. After treatment with di-isopropyl phosphorofluoridate, to obviate the effects of the di-isopropyl phosphorofluoridate-sensitive esterase, the enzyme preparation hydrolyses α-naphthyl acetate, α-naphthyl propionate and α-naphthyl butyrate, but not cholesteryl acetate. The Vmax. for the naphthyl esters decreased with increase in chain length of the acyl group. The acetate ester is hydrolysed 34 times as fast as the butyrate and about seven times as fast as the propionate derivative. The Km values for these three esters, measured at pH7·2 and 37°, are 2·8×10−4m, 3·1×10−4m and 7·3×10−5m for the acetate, propionate and butyrate derivatives respectively. 4. The Hofstee (1952) plots for the kinetic data show a single line, indicating that the two most-rapidly migrating esterases, although electrophoretically separable, are not kinetically distinguishable in the substrate ranges examined.
Preparation of isolated multiple forms of rabbit brain aldolase
Authors: Foxwell C. J. , Buck G. Maureen , Baron D. N.
Published: August 2015

1. A method is described for partial purification of the total aldolase of rabbit brain by ammonium sulphate precipitation followed by fractionation of the aldolase into five electrophoretically distinct forms. 2. The method allows a laboratory-scale preparation of the five aldolases. 3. With the exception of band 3 the fructose 1,6-diphosphate/fructose 1-phosphate activity ratios of the isolated multiple forms show a continuous increase from isoenzyme 1 to isoenzyme 5; this evidence supports the four-sub-unit model.
Naturally occurring low-molecular-weight peptides from the blowfly Phormia regina
Authors: Bodnaryk R. P. , Levenbook L.
Published: August 2015

Freely occurring α-l-aspartyl-l-histidine, β-alanyl-l-histidine (carnosine), α-l-aspartyl-α-l-lysine, l-asparaginyl-α-l-lysine, α-l-glutamyl-α-l-lysine, l-isoleucyl-α-l-lysine, l-seryl-α-l-lysine, l-valyl-α-l-lysine and glycyl-α-l-lysyl-α-l-lysine were identified in young larvae (second and third instars) of the blowfly (Phormia regina).
The neurotoxicity of β-N-oxalyl-l-αβ-diaminopropionic acid, the neurotoxin from the pulse Lathyrus sativus
Authors: Cheema P. S. , Malathi K. , Padmanaban G. , Sarma P. S.
Published: August 2015

Intraperitoneal administration of β-N-oxalyl-l-αβ-diaminopropionic acid, the neurotoxin from Lathyrus sativus, to 12-day-old rats causes typical convulsions within 10min. There is a striking accumulation of glutamine in the brain, and chronic ammonia toxicity is indicated. There are no changes in the amounts of urea, aspartic acid and glutamic acid in the brain. Adult rats, even when injected with a dose of excess of β-N-oxalyl-l-αβ-diaminopropionic acid, do not develop symptoms, and there are no changes in the amounts of glutamine or ammonia in the brain. A significant concentration of β-N-oxalyl-l-αβ-diaminopropionic acid can be detected in the brain of the young rat but not in that of the adult animal. It is concluded that β-N-oxalyl-l-αβ-diaminopropionic acid interferes with the ammonia-generating or -fixing mechanisms in the brain and leads to chronic ammonia toxicity.
Hormonal steroids in the tissue of induced rat mammary tumours
Authors: Briziarelli G. , Lusignani A. , Pedrali C.
Published: August 2015

The possible presence of steroids in the tissue of induced hormone-dependent rat mammary tumours was investigated. The method used involves a preliminary extraction of tumours followed by chemical separation and thin-layer chromatography. The identified compounds were cholesterol, androst-4-ene-3,17-dione, 5β-androst-1-ene-3,17-dione, androsta-1,4-diene-3,17-dione and oestrone. This is the first report of the presence of these steroids in the tissue of an experimental tumour of a non-endocrine organ. In particular 5β-androst-1-ene-3,17-dione has not previously been identified from natural sources.
Intestinal disaccharidase activities in the chick
Authors: Siddons R. C.
Published: August 2015

1. Disaccharidase activities of the small and large intestines of the chick were studied. 2. Homogenates of the small intestine readily hydrolysed maltose, sucrose and palatinose (6-O-α-d-glucopyranosyl-d-fructose), hydrolysed lactose slowly and did not hydrolyse trehalose and cellobiose. 3. Within the small intestine the disaccharidases were located mainly in the intestinal wall; the activity in the contents accounted for less than 5% of the total activity. 4. The disaccharidases were non-uniformly distributed along the small intestine, the activities being greatest in the middle section. 5. The disaccharidase activities increased with age between 1 and 43 days. 6. Homogenates of the large intestine and contents readily hydrolysed maltose, sucrose, palatinose and lactose and hydrolysed cellobiose and trehalose slowly. 7. The large-intestinal disaccharidases were located mainly in the contents. 8. Similar Km and pH optimum values were found for the maltase, sucrase and palatinase activities of the large and small intestines. 9. The lactase activity of the large intestine was markedly affected by diet and had different Km and pH values from the small intestinal lactase. 10. Low activities of intestinal disaccharidase were found in 12-day-old embryos and marked increases in the intestinal disaccharidases of the developing embryo occurred 2–3 days before hatching.
Incorporation of [32P]orthophosphate into brain-slice phospholipids and their precursors. Effects of electrical stimulation
Authors: Pumphrey A. M.
Published: August 2015

1. The incorporation of [32P]phosphate into phospholipids was measured in slices cut from the pial surface of guinea-pig cerebral cortex; incorporation into the phosphorus of some water-soluble precursors of phospholipid was measured under similar conditions. 2. Slices subjected to overall electrical stimulation at a frequency of 5pulses/sec. differed from control slices in their pattern of phospholipid labelling. After 1hr. of stimulation, incorporation of [32P]phosphate into phosphatidylcholine, ethanolamine phospholipid and cardiolipin was respectively 54, 55 and 58% of the control value, and that into phosphatidylinositol was 186% of control. Phosphatidic acid labelling tended to increase with electrical stimulation, but the statistical significance of this change was marginal. Labelling of phosphatidylglycerol and di- and tri-phosphoinositides was not affected significantly by electrical stimulation. 3. Electrical stimulation of the tissue altered the specific radioactivities of water-soluble precursors of phospholipid. 4. The turnover rates of the phosphate groups of phospholipids were estimated approximately from the specific radioactivities of phospholipids and their precursors. Phosphatidylinositol (and its lipid-soluble precursors) showed the largest change in turnover rate in response to electrical stimulation of the tissue; the turnover rates of other lipids were also affected. Changes in the specific radioactivity of phospholipids did not correspond to changes in turnover in these experiments.
Inhibition of hepatic gluconeogenesis by ethanol
Authors: Krebs H. A. , Freedland R. A. , Hems R. , Stubbs Marion
Published: August 2015

1. Gluconeogenesis from 10mm-lactate in the perfused liver of starved rats is inhibited by ethanol. The degree of inhibition reached a maximum of 66% at 10mm-ethanol under the test conditions and decreased at higher ethanol concentrations. The concentration-dependence of the inhibition is paralleled by the concentration-dependence of the activity of alcohol dehydrogenase. The enzyme is also inhibited by ethanol concentrations above 10mm. 2. Gluconeogenesis from pyruvate is not inhibited by ethanol. 3. The degree of the inhibition of gluconeogenesis from lactate by ethanol depends on the concentration of lactate and other oxidizable substances, e.g. oleate, in the perfusion medium. 4. Ethanol also inhibits, to different degrees, gluconeogenesis from glycerol, dihydroxyacetone, proline, serine, alanine, fructose and galactose. 5. The inhibition of gluconeogenesis from lactate by ethanol is reversed by acetaldehyde. 6. Pyrazole, a specific inhibitor of alcohol dehydrogenase, also reverses the inhibition of gluconeogenesis by ethanol. 7. Gluconeogenesis in kidney cortex, where the activity of alcohol dehydrogenase is very low, is not inhibited by ethanol. 8. Kidney cortex, testis, ovary, uterus and certain tissues of the alimentary tract were the only rat tissues, apart from the liver, that showed measurable alcohol dehydrogenase activity. 9. The concentrations of pyruvate in the liver were decreased to about one-fifth by ethanol. 10. The concentration of lactate in the perfused liver was about 3mm below that of the perfusion medium 30min. after the addition of 10mm-lactate. 11. The great majority of the findings support the view that the inhibition of gluconeogensis by ethanol is caused by the alcohol dehydrogenase reaction, which decreases the [free NAD+]/[free NADH] ratio. The decrease lowers the concentration of pyruvate and this is the immediate cause of the inhibition of gluconeogenesis from lactate, alanine and serine: the fall in the concentration of pyruvate lowers the rate of the pyruvate carboxylase reaction, one of the rate-limiting reactions of gluconeogenesis. The cause of the inhibition of gluconeogenesis from other substrates is discussed.
The heparin–protein complex of ox liver capsule. Isolation and chemical characterization
Authors: Serafini-Fracassini A , Durward J J , Floreani L.
Published: August 2015

1. A procedure for the isolation and purification of the heparin–protein complex from ox liver capsule, based on the solubility properties of mucopolysaccharide–cetylpyridinium complexes, is described. 2. The yield of the heparin–protein complex with this method averages 35mg./100g. of dry ox liver capsule. 3. The results of analyses on the polysaccharide show good agreement with values previously published for purified heparin fractions. 4. The amino acid analysis of the protein component shows several similarities to that of chondromucoprotein. 5. The results of β-carbonyl elimination, either with or without catalytic hydrogenation, and column chromatography after β-elimination, show that the polysaccharide is covalently bound to the protein.
Studies on the reduction of a human 19s immunoglobulin M
Authors: Beale D , Feinstein A.
Published: August 2015

1. Reduction of a 19s immunoglobulin M with 3mm-mercaptoethanol or 0·05–0·5mm-dithiothreitol followed by alkylation gave sedimentation patterns indicating products compatible with structures consisting of one, two, three, four and five 7s sub-units. This supports the concept of a five-sub-unit structure for immunoglobulin M. 2. Reduction with 0·125mm-dithiothreitol or 20mm-cysteine produced 7s sub-units that could not be dissociated into chains in m-propionic acid. 3. By labelling (with iodo[2−14C]acetic acid) the thiol groups liberated during reduction with 0·125mm-dithiothreitol, it was possible to identify the tryptic peptides involved in the disulphide bridges that link the 7s sub-units together (inter-sub-unit bridges). 4. By further reducing and labelling (with iodo[2−14C]acetic acid) the 7s sub-units produced by 0·125mm-dithiothreitol, it was possible to identify tryptic peptides derived from intra-sub-unit bridges. 5. Sub-units produced by reduction with 20mm-cysteine proved to be unsuitable for distinguishing between inter-sub-unit bridges and intra-sub-unit bridges. 6. The possible arrangement of the interchain disulphide bridges was deduced.
Measurement of adipose-tissue metabolites in vivo
Authors: Ballard F. J. , Hanson R. W.
Published: August 2015

1. The concentrations of glucose, pyruvate, lactate, citrate, glutamate, malate and aspartate were measured in epididymal adipose tissue from starved, fed and starved–re-fed rats. 2. To measure these intermediates it was necessary to correct for their concentration in the extracellular tissue space, which was considered to be most satisfactorily equated with the glucose space. This space in vivo was 7·42, 4·90 and 7·54ml./100g. wet wt. of tissue in adipose tissue taken from starved, fed and starved–re-fed rats respectively. After correction for the glucose space, the concentrations of metabolites (nmoles/g. of cells) in epididymal adipose tissue of fed rats were: pyruvate, 8·5; lactate, 50·3; citrate, 18·5; glutamate, 100·0; malate, 6·4; aspartate, 34·2. 3. Starvation for 72hr. resulted in a fall in pyruvate and aspartate concentrations to 3·57 and 25·1nmoles/g.; starvation for 72hr. followed by re-feeding for 72hr. caused an increase in glutamate and aspartate concentrations to 140 and 67·6nmoles/g. 4. These changes are interpreted with regard to the simultaneous alteration in lipogenesis that occurs during the starvation–re-feeding cycle.
Reactions of vitamin A with acceptors of electrons. Interactions with iodine and the formation of iodide
Authors: Lucy J. A. , Lichti F. Ulrike
Published: August 2015

1. The reactions of retinol and retinoic acid with iodine were investigated since knowledge of the chemical reactions of vitamin A with acceptors of electrons may shed light on its biochemical mode of action. 2. Colloidal retinol, but not retinoic acid, reacts with iodine to yield a blue–green complex that rapidly decomposes, giving iodide and an unknown species with λmax. at 870mμ. 3. In addition, both retinol and retinoic acid reduce iodine to iodide by a reaction that does not involve an intermediate coloured complex; this reaction appears to yield unstable carbonium ion derivatives of the vitamin. 4. The presence of water greatly facilitates the production of iodide from vitamin A and iodine. 5. Possible chemical pathways involved in these reactions are discussed. 6. It is suggested that the chemical properties of retinol and retinoic acid that underlie their biochemical behaviour might be apparent only when the molecules are at a lipid–water interface, and that vitamin A might be expected to react with a number of different electron acceptors in vivo.
The effect of low temperatures on fatty acid biosynthesis in plants
Authors: Harris P. , James A T
Published: August 2015

1. Of three systems, bulb tissue, plant leaf tissue and intact green algal (Chlorella vulgaris) cells, only the former shows an increase in rate of formation of unsaturated fatty acids with decrease in temperature. 2. In bulb tissue the oxygen concentration is rate-limiting for synthesis of unsaturated fatty acids at temperatures down to 10°. 3. At elevated oxygen concentrations the formation of unsaturated fatty acids in bulb tissue increases with temperature. 4. The failure of photosynthetic tissues to respond to either lower temperatures or increased oxygen concentrations in the presence of light is attributed to photosynthetic production of excess of oxygen. This is supported by the fact that in the dark a potentiating oxygen effect on the formation of unsaturated fatty acids can be demonstrated. 5. The HCO3− ion concentration has a small effect on the formation of unsaturated fatty acids. 6. Elevated content of unsaturated acids at lower temperatures in plants is attributed to increases in oxygen concentration in solution.
Purification and general properties of argininosuccinate lyase from jack bean, Canavalia ensiformis (L.) DC
Authors: Rosenthal Gerald A. , Naylor Aubrey W.
Published: August 2015

1. Argininosuccinate lyase (EC 4.3.2.1) from jack bean [Canavalia ensiformis (L.) DC] seeds was purified 532-fold from an acetone–butanol-dried powder. 2. The enzyme functions reversibly and exhibits maximum stability at 16°. 3. At 16° it has a half-life (t½) of 263min. 4. The enzyme is both cold-labile (t½ 131min. at 0°) and heat-inactivated (t½ 74min. at 38°); inactivation appears to be irreversible. 5. Treatment of the acetone–butanol-extracted powder with sodium dodecyl sulphate increased the sensitivity of the enzyme to temperature (t½ 70min. at 0°; t½ 23min. at 38°). 6. Addition, to the purified enzyme, of a fraction containing lipid from the seed increased the half-life to about 510min. at either 0° or 38°. 7. Arginine or homoarginine, and to a smaller extent some other amino acids or fumarate, protected the enzyme from cold-inactivation. 8. Reactivation attempts with both the cold- and heat-inactivated enzyme failed. 9. The Km value for argininosuccinate at pH7·5 is 1·3×10−4. 10. The enzyme was inactivated completely within 15min. at 16° by 0·5mm-p-hydroxymercuribenzoate, and subsequent exposure to 5mm-cysteine had no restorative effect.
Cataract produced by tyrosinase and tyrosine systems in rabbit lens in vitro
Authors: Srivastava Satish K. , Beutler Ernest
Published: August 2015

1. Cataracts were developed by incubating rabbit lenses for 22hr. at 37° in a culture medium containing tyrosine and tyrosinase (EC 1.10.3.1). 2. A 45% diminution in the content of GSH and significant inhibition of glucose 6-phosphate dehydrogenase (EC 1.1.1.49) activity were observed in the cataractous lenses compared with controls. 3. GSSG accumulated in both cataractous and control lenses. Significant amounts of GSSG were transported outward from the cataractous lenses and small amounts from control lenses. 4. Transport of GSSG from rabbit lens incubated in a diffusate of plasma from a naphthalene-fed rabbit was also observed. 5. GSSG was found in the aqueous humour obtained between 2 and 24hr. after feeding of naphthalene to rabbits; subsequently the GSSG in the aqueous humour decreased to almost undetectable amounts in 48hr.; in controls, GSSG was not detectable. 6. A possible mechanism of formation of experimental and senile cataract is briefly discussed.
Some properties of hepatic glycerol kinase and their relation to the control of glycerol utilization
Authors: Robinson Janice , Newsholme E A
Published: August 2015

1. Glycerol kinase (EC 2.7.1.30) is shown to catalyse a non-equilibrium reaction in rat liver; and, as it is the first enzyme in the pathway metabolizing glycerol, its properties may be pertinent to the metabolic regulation of glycerol uptake and utilization by this tissue. 2. The properties of hepatic glycerol kinase were studied by using a radiochemical technique to measure the enzyme activity. When the concentration of ATP is low the activity of glycerol kinase is inhibited by high concentrations of glycerol; but when the concentration of ATP is high there is no inhibition and the double-reciprocal plot is linear, providing a Km for glycerol of 3·16×10−6m. Glycerol kinase is activated by high ATP concentrations provided that the concentration of the second substrate (glycerol) is high; at low concentrations of glycerol ATP does not activate the enzyme so that the double-reciprocal plot is linear, providing a Km for ATP of 5·8×10−5m. It is suggested that these kinetics may be explained by a model similar to that described by Ferdinand (1966) for phosphofructokinase. 3. Hepatic glycerol kinase is inhibited by ADP and AMP, and raising the Mg2+ concentration increases the inhibition by these two compounds; this suggests that ADP–Mg2+ and AMP–Mg2+ complexes are the inhibitory species. The physiological significance of these inhibitions may be to prevent phosphorylation of glycerol when the hepatic ATP concentration is low. It is suggested that this inhibition may provide an approach to the problem of measurement of rates of lipolysis by glycerol release in tissues that contain glycerol kinase (e.g. liver, kidney, muscle, adipose tissue). 4. Hepatic glycerol kinase is inhibited by l-3-glycerophosphate competitively with respect to glycerol. The physiological significance of this inhibition may be that factors that change the intracellular concentration of l-3-glycerophosphate could change glycerol uptake by the tissue. Thus it is suggested that thyroxine treatment or feeding rats on a diet high in glycerol, which increase the activity of glycerophosphate oxidase in liver and kidney cortex respectively, lead to an increased glycerol uptake through a decrease in the concentration of glycerophosphate in these tissues. It is known that ethanol administration decreases glycerol uptake by liver, and this can be explained by the increased concentration of l-3-glycerophosphate causing inhibition of glycerol kinase.
The precipitation of toroidal collagen fibrils
Authors: Cooper Alan
Published: August 2015

The morphology of aggregates of calf-skin tropocollagen, precipitated by continuous injection into neutral phosphate buffers at 35°, has been studied by electron microscopy. Although most of the collagen is precipitated as normal native fibrils, a small proportion forms closed toroidal structures having the usual native band–interband pattern. Theoretical considerations, based on elastic energies in a general microfibril model, predict that the toroids should have a simple super-helical structure, and this is not inconsistent with the observations. From the theoretical energies it was possible to estimate a crude lower limit of 3kcal./mole for the free energy of association of the tropocollagen macromolecules.
The conversion of 4-hydroxypyrazolo-[3,4-d]pyrimidine (allopurinol) into 4,6-dihydroxypyrazolo[3,4-d]pyrimidine (oxipurinol) in vivo in the absence of xanthine–oxygen oxidoreductase
Authors: Chalmers R. A. , Parker R. , Simmonds H A , Snedden W. , Watts R. W. E.
Published: August 2015

1. A patient with congenital deficiency of xanthine oxidase (EC 1.2.3.2) (xanthinuria) excreted the xanthine isomer 4,6-dihydroxypyrazolo[3,4-d]pyrimidine (oxipurinol) in his urine when the hypoxanthine isomer 4-hydroxypyrazolo[3,4-d]pyrimidine (allopurinol) was given by mouth. 2. The identity of the oxipurinol that the patient excreted was established by mass spectrometry. 3. The mass spectra and infrared spectra of allopurinol, oxipurinol, hypoxanthine and xanthine are compared. 4. A mechanism for the fragmentation of these compounds that occurs during their mass-spectrometric investigation is proposed. 5. A possible metabolic pathway for the oxidation of allopurinol to oxipurinol in the absence of xanthine oxidase is discussed.
Effect of potassium ions on the attachment of polyribosomes to the membranes in lysates of exponential-phase cells of Bacillus amyloliquefaciens
Authors: Coleman Geoffrey
Published: August 2015

1. The distribution of ribosomal components between the soluble and membrane fractions of a preparation of exponential-phase cells of Bacillus amyloliquefaciens lysed with lysozyme in 0·05m-tris buffer, pH7·6, containing 0·01m-Mg2+, was strongly influenced by the addition of K+ to the buffer, in the range 0–0·1m. 2. In the absence of K+, 37% of the ribosomal material was bound to the membrane and was not removed by repeated washing with the lysing buffer. The amount of bound material was progressively decreased on increasing the K+ concentration to 0·1m, when only 5% of ribosomal components remained attached to the membrane. 3. About 87% of the material that remained bound to the washed membranes prepared in the absence of added K+ was removed on suspension of the membranes in a buffer containing 0·1m-potassium chloride. 4. In the absence of K+, washed membranes, containing no detectable ribosomal material, were able to re-attach no more than half as much material as was found associated with membranes in the same buffer immediately after lysis. 5. There was no evidence of binding of specific components to the membrane. 6. In the presence of 0·05m-tris buffer, pH7·6, maximum incorporation of amino acids into protein by B. amyloliquefaciens polyribosomes is effected in the presence of 0·01m-Mg2+ and 0·07–0·1m-K+, under which conditions less than 10% of the ribosomal material of a cell lysate would be membrane-bound.
The reduction of cystine during its transport into rat jejunal everted segments and sacs
Authors: Crawhall J. C. , Davis M. G.
Published: August 2015

1. Everted segments and sacs of rat jejunum were incubated in buffer containing [35S]cystine. 2. Concentration gradients were achieved by both segments and sacs, and the effects of duration of incubation and of cystine concentration on the isotope distribution ratios were determined. 3. Kinetic constants were determined for the uptake of cystine by both segments and sacs, and the differences between the two systems are discussed. 4. Reduction to cysteine was virtually complete intracellularly and in the sac lumen. Extensive reduction in the medium occurred only when segments were incubated. 5. Anaerobiosis prevented a concentration gradient being obtained between the medium and the tissue, but had little effect on the extent of reduction to cysteine in the tissue and sac lumen. 6. It is concluded that cystine is transported by an active process into rat jejunum, where it is present almost entirely in the reduced form, and that efflux of cysteine occurs through the serosal surface.
Rates of ketone-body formation in the perfused rat liver
Authors: Krebs H. A. , Wallace Patricia G. , Hems R. , Freedland R. A.
Published: August 2015

1. The rates of formation of acetoacetate and β-hydroxybutyrate by the isolated perfused rat liver were measured under various conditions. 2. The rates found after addition of butyrate, octanoate, oleate and linoleate were about 100μmoles/hr./g. wet wt. in the liver of starved rats. These rates are much higher than those found with rat liver slices. 3. The differences between the rates given by slices and by the perfused organ were much higher with the long-chain than with short-chain fatty acids. The increments caused by oleate and linoleate were 12 and 16 times as large in the perfused organ as in the slices, whereas the increments caused by butyrate and octanoate were about four times as large. 4. The rates of ketogenesis in the unsupplemented perfused liver of well-fed rats, and the increments caused by the addition of fatty acids, were about half of those in the liver from starved rats. 5. The value of the [β-hydroxybutyrate]/[acetoacetate] ratio of the medium was raised by octanoate, oleate and linoleate. 6. Carnitine did not significantly accelerate ketogenesis from fatty acids. 7. Oleate formed up to 82% of the expected yield of ketone bodies. 8. In the liver of alloxan-diabetic rats the endogenous rates of ketogenesis were raised, in some cases as high as in the liver from starved rats, after addition of oleate. 9. On addition of either β-hydroxybutyrate or acetoacetate to the perfusion medium the liver gradually adjusted the [β-hydroxybutyrate]/[acetoacetate] ratio towards the normal range. 10. The [β-hydroxybutyrate]/[acetoacetate] ratio of the medium was about 0·4 when slices were incubated, but near the physiological value of 2 when the liver was perfused. 11. The experiments demonstrate that for the study of ketogenesis slices are in many ways grossly inferior to the perfused liver.
Inhibition by oestrogen of collagen breakdown in the involuting rat uterus
Authors: Woessner J. F.
Published: August 2015

1. Both the post-partum involution of the rat uterus and the rapid breakdown of collagen that accompanies it are extensively inhibited by oestrogenic hormones. In the normal rat, 85% of the uterine collagen is degraded within 4 days after parturition; in rats treated with 100μg. of 17β-oestradiol/day, only 35% of uterine collagen is broken down in the same period. 2. Similar effects are produced by diethylstilboestrol if the dose is increased tenfold. 3. Collagen breakdown is inhibited to a greater extent than is the loss of wet weight by oestradiol but not by diethylstilboestrol. 4. The oestrogens appear to act by blocking the breakdown of collagen. There is a greatly decreased concentration of free hydroxyproline in the uterus of treated animals. 5. Acid hydrolase concentrations (β-glucuronidase, β-galactosidase, cathepsin D and acid phosphatase) in the uterus are decreased by oestrogen treatment compared with controls, but the total amounts of these enzymes in the uterus are somewhat elevated. Oestrogens do not appear to inhibit collagen breakdown by altering the concentration and total amount of acid hydrolases.
Utilization of l-threonine by a species of Arthrobacter. A novel catabolic role for ‘aminoacetone synthase’
Authors: McGilvray D. , Morris J G
Published: August 2015

1. A species of Arthrobacter (designated Arthrobacter 9759) was isolated from soil by its ability to grow aerobically on l-threonine as sole source of carbon atoms, nitrogen atoms and energy; the organism also grew well on other sources of carbon atoms including glycine, but no growth was obtainable on aminoacetone or dl-1-aminopropan-2-ol. 2. During growth on threonine, 14C from l-[U−14C]threonine was rapidly incorporated into glycine and citrate, and thereafter into serine, alanine, aspartate and glutamate. 3. With extracts of threonine-grown cells supplied with l-[U−14C]threonine, evidence was obtained of the NAD and CoA-dependent catabolism of l-threonine to produce acetyl-CoA plus glycine. Short-term incorporation studies in which [2−14C]acetate and [2−14C]glycine were supplied (a) to cultures growing on threonine, and (b) to extracts of threonine-grown cells, showed that the acetyl-CoA was metabolized via the tricarboxylic acid cycle and glyoxylate cycle whereas the glycine was converted into pyruvate via the folate-dependent ‘serine pathway’. 4. The threonine-grown organism contained ‘biosynthetic’ threonine dehydratase and a potent NAD-linked l-threonine dehydrogenase but possessed no l-threonine aldolase activity. 5. Evidence was obtained that the acetyl-CoA and glycine produced from l-threonine had their immediate origin in the α-amino-β-oxobutyrate formed by the threonine dehydrogenase; the CoA-dependent cleavage of this compound was catalysed by an α-amino-β-oxobutyrate CoA-ligase, which was identified with ‘aminoacetone synthase’. A continuous spectrophotometric assay of this enzyme was developed, and it was found to be inducibly synthesized only during growth on threonine and not during growth on acetate plus glycine. 6. By using a reconstituted mixture of separately purified l-threonine dehydrogenase and α-amino-β-oxobutyrate CoA-ligase (i.e. ‘aminoacetone synthase’), l-[U−14C]threonine was broken down to [14C]glycine plus [14C]acetyl-CoA (trapped as [14C]citrate). 7. There was no evidence of aminoacetone metabolism by Arthrobacter 9759 even though a small amount of this amino ketone appeared in the culture medium during growth on threonine.
The influence of metal ions on the orthophosphatase and inorganic pyrophosphatase activities of human alkaline phosphatase
Authors: Moss D W
Published: August 2015

1. The differential effects of adding Zn2+ and Mg2+ on the orthophosphatase and inorganic pyrophosphatase activities of human intestinal alkaline phosphatase were studied. 2. In the presence of excess of Zn2+, inorganic pyrophosphatase activity is inhibited. At higher concentrations of pyrophosphate, hydrolysis of this substrate takes place, but is inhibited competitively by the Zn2+–pyrophosphate complex. This complex also acts as a competitive inhibitor of orthophosphate hydrolysis. 3. Excess of Mg2+ also inhibits pyrophosphatase action by removal of substrate; at low concentrations, this ion activates pyrophosphatase, as is the case with orthophosphatase. 4. It is concluded that, when interactions between metal ions and pyrophosphate are taken into account, the effects of these ions are consistent with the view that alkaline phosphatases possess both orthophosphatase and inorganic pyrophosphatase activities.
The subcellular distribution of 32P-labelled phospholipids, 32P-labelled ribonucleic acid and 125I-labelled iodoprotein in pig thyroid slices. Effect in vitro of thyrotrophic hormone and dibutyryl-3′,5′-(cyclic)-adenosine monophosphate
Authors: Kerkof P. R. , Tata J. R.
Published: August 2015

1. The incorporation in vitro of [32P]phosphate into phospholipids and RNA and of [125I]iodide into protein-bound iodine by pig thyroid slices incubated for up to 6hr. was studied. The subcellular distribution of the labelled products formed after incubation with radioactive precursor in the nuclear, mitochondrial, smooth-microsomal, rough-microsomal and cell-sap fractions was also studied. 2. Pig thyroid slices actively took up [32P]phosphate from the medium during 6hr. of incubation; the rate of incorporation of 32P into phospholipids was two to five times that into RNA. 3. The uptake of [125I]iodide by the slices from the medium was rapid for 4hr. of incubation, 6–10% of the label being incorporated into iodoprotein. 4. Much of the 32P-labelled phospholipid accumulated in mitochondria and microsomes, whereas the nuclear fraction contained most of the 32P-labelled RNA. After 2hr. of incubation most of the 32P-labelled cytoplasmic RNA accumulated in the rough-microsomal fraction. The major site of localization of proteinbound 125I was the smooth-microsomal fraction, and gradually increasing amounts appeared in the soluble cytoplasm fraction, suggesting a vectorial discharge of [125I]iodoprotein (presumably thyroglobulin) from smooth vesicles into the colloid. 5. The addition of 0·1–0·4 unit of thyrotrophic hormone/ml. of incubation medium markedly enhanced the accumulation of 32P-labelled phospholipids in the microsomal fractions and to a much smaller extent that of 32P-labelled RNA without any increase in the total uptake of the label. Almost simultaneously the hormone increased the uptake of [125I]iodide by the slices and enhanced the accumulation of protein-bound 125I in the smooth-microsomal fraction. 6. As a function of time of incubation, thyrotrophic hormone had a biphasic effect on [125I]iodide uptake and protein-bound 125I formation, the stimulatory effect being reversed after 4hr. of incubation. 7. 6-N-2′-O-Dibutyryl-3′,5′-(cyclic)-AMP, but not 3′,5′-(cyclic)-AMP or 5′-AMP, mimicked the action of thyrotrophic hormone on iodine uptake as well as on iodination of protein. On the other hand, the mimicry by 6-N-2′-O-dibutyryl-3′,5′-(cyclic)-AMP of the stimulatory effect of thyrotrophic hormone on the formation of labelled thyroid phospholipids and RNA was only an apparent one resulting from an enhanced uptake of [32P]phosphate. 8. It is concluded that thyrotrophic hormone causes a co-ordinated increase in the formation or accumulation of phospholipids, RNA and iodoprotein associated with the endoplasmic reticulum, and that 6-N-2′-O-dibutyryl-3′,5′-(cyclic)-AMP mimics the more rapid effects of thyrotrophic hormone on transport and metabolic functions of thyroid cells, but does not influence their slower biosynthetic responses to the hormone.
Effects of adenosine phosphates and nicotinamide nucleotides on pyruvate carboxylase from baker's yeast
Authors: Cazzulo J. J. , Stoppani A. O. M.
Published: August 2015

1. Pyruvate carboxylase from baker's yeast is inhibited by ADP, AMP and adenosine at pH8·0 in the presence of magnesium chloride concentrations equal to or higher than the ATP concentration. The adenine moiety is essential for the inhibitory effect. 2. In the absence of acetyl-CoA (an allosteric activator) ADP, AMP and adenosine are competitive inhibitors with respect to ATP. In the presence of acetyl-CoA, besides the effect with respect to ATP, AMP competes with acetyl-CoA, whereas ADP and adenosine are non-competitive inhibitors with respect to the activator. 3. Pyruvate carboxylase is inhibited by NADH. The inhibition is competitive with respect to acetyl-CoA and specific with respect to NADH, since NAD+, NADP+ and NADPH do not affect the enzyme activity. In the absence of acetyl-CoA, NAD+, NADH, NADP+ and NADPH do not inhibit pyruvate carboxylase. 4. Pyruvate carboxylase is inhibited by ADP, AMP and NADH at pH6·5, in the presence of 12mm-Mg2+, 0·75mm-Mn2+ and 0·5mm-ATP, medium conditions similar to those existing inside the yeast cell. The ADP and NADH effects are consistent with a regulation of enzyme activity by the intracellular [ATP]/[ADP] ratio and secondarily by NADH concentration. These mechanisms would supplement the already known control of yeast pyruvate carboxylase by acetyl-CoA and l-aspartate. Inhibition by AMP is less marked and its physiological role is perhaps limited.