bluuebunny/crossref_metadata_2025 huggingface dataset:
Species differences in the aromatization of quinic acid in vivo and the role of gut bacteria
Authors: Adamson R. H. , Bridges J. W. , Evans M. E. , Williams R. T.
Published: August 2015

1. The fate of (-)-quinic acid has been investigated in 22 species of animals including man. 2. In man and three species of Old World monkeys, i.e. rhesus monkey, baboon and green monkey, oral quinic acid was extensively aromatized (20–60%) and excreted in the urine as hippuric acid, which was determined fluorimetrically. 3. In three species of New World monkeys, i.e. squirrel monkey, spider monkey and capuchin, in three species of lemurs, i.e. bushbaby, slow loris and tree shrew, in the dog, cat, ferret, rabbit, rat, mouse, guinea pig, hamster, lemming, fruit bat, hedgehog and pigeon, oral quinic acid was not extensively aromatized (0–5%). 4. In the rhesus monkey, injected quinic acid was not aromatized, but largely excreted unchanged. 5. In rhesus monkeys pretreated with neomycin to suppress gut flora, the aromatization of oral quinic acid was considerably suppressed. 6. In rats and rhesus monkeys [14C]quinic acid was used and this confirmed its low aromatization in rats and its high aromatization in the monkeys. 7. Shikimic acid given orally was excreted as hippuric acid (26–56%) in rhesus monkeys, but not in rats. 8. The results support the view that quinic acid and shikimic acid are aromatized by the gut flora in man and the Old World monkeys.
Appendix: Nuclear-magnetic-resonance and mass-spectral examination of the principal bile alcohol from Catostomus commersoni and its anhydro derivative
Authors: Tökés Lászlo
Published: August 2015

On the basis of i.r. and chromatographic evidence 5α-cholestane-3α,7α,12α,24ξ,26-pentol(I) and 24ξ,26-oxido-5α-cholestane-3α,7α,12α-triol (IV) structures have been proposed for the principal cleavage and alkaline-hydrolysis products of the bile salts from Catostomus commersoni (Anderson & Haslewood, 1969). N.m.r.- and mass-spectral analyses of these substances and the tetra- (II) and penta-acetate (III) derivatives of (I) provided supporting evidence for the structural assignments.
Inhibition of fly head acetylcholinesterase by bis-[(m-hydroxyphenyl)trimethylammonium iodide] esters of polymethylenedicarbamic acids
Authors: Davies J. H. , Campbell W. R. , Kearns C. W.
Published: August 2015

A series of bis-[(m-hydroxyphenyl)trimethylammonium iodide] esters of polymethylenedicarbamic acids and a number of (m-hydroxyphenyl)trimethylammonium iodide esters of straight-chain N-alkylcarbamic acids have been examined as inhibitors of acetylcholinesterase from fly head. Evidence is presented suggesting that inhibition of acetylcholinesterase by the bis-carbamates is due to carbamoylation of the enzyme, as is generally thought to be the case with esters of N-alkylcarbamic acids. Inhibition is irreversible. The (m-hydroxyphenyl)trimethylammonium iodide ester of N-hexylcarbamic acid also inhibits fly head acetylcholinesterase irreversibly. There is therefore no need to implicate a second functional group in bis-carbamate esters to explain the irreversible inhibition of the enzyme. An unusual feature of the inhibition is that inhibition lines do not pass through 100% enzyme activity at t=0, except for rather low concentrations of inhibitor (<10μm for the octamethylene compound). Also, inhibition lines tend towards a maximum slope as inhibitor concentration is increased. The first observation indicates complex-formation, even in the presence of high concentrations of substrate, and by using measurements of inhibition at relatively high inhibitor concentrations, affinity constants K′a have been calculated. K′a varies from 0.1μm for the dodecamethylene compound to 10μm for the tetramethylene compound, in the presence of 3.75mm-acetylthiocholine, indicating high affinity for the enzyme. The second observation shows that, owing to this high affinity, the enzyme becomes saturated with inhibitor under the experimental conditions employed, and from the limiting slope values of the carbamoylation rate constant (k2) have been calculated. k2 varies from 0.15min−1 for the tetramethylene compound to 1min−1 for the decamethylene compound. Variations of potency in this series are therefore mainly due to changes in affinity (100-fold) rather than in carbamoylation rate (sevenfold). The observation that large molecules may acylate the enzyme raises certain problems, which are discussed.
Equilibrium relations between the cytoplasmic adenine nucleotide system and nicotinamide–adenine nucleotide system in rat liver
Authors: Veech R. L. , Raijman L. , Krebs H. A.
Published: August 2015

1. The ratio [ATP]/[ADP][Pi], as measured by direct determination of the three components in rat liver, was found in various nutritional states to have approximately the same value as the ratio [ATP]/[ADP][Pi] calculated from the concentrations of lactate, pyruvate, glyceraldehyde phosphate and 3-phosphoglycerate on the assumption that lactate dehydrogenase, glyceraldehyde phosphate dehydrogenase and 3-phosphoglycerate kinase are at near-equilibrium in the liver. This implies that the redox state of the NAD couple in the cytoplasm is linked to, and partially controlled by, the phosphorylation state of the adenine nucleotides. 2. The combined equilibrium constant of the glyceraldehyde 3-phosphate dehydrogenase and 3-phosphoglycerate kinase reactions at 38°C and I0.25, was found to be 5.9×10−6. 3. The fall of the [NAD+]/[NADH] ratio in starvation and other situations is taken to be the consequence of a primary fall of the [ATP]/[ADP][HPO42−] ratio.
The biosynthesis of androst-16-enes in boar testis tissue
Authors: Katkov Tatyana , Gower D. B.
Published: August 2015

1. The metabolism of [4-14C]pregnenolone to androst-16-enes has been studied in short-term incubations of boar testis tissue. With fresh tissue androsta-5,16-dien-3β-ol (8%) and 5α-androst-16-en-3β-ol (2%) were formed. Tissue that had been stored at −20°C was still capable of metabolizing pregnenolone to androsta-5,16-dien-3β-ol. 2. NADPH was essential for the formation of androsta-5,16-dien-3β-ol from pregnenolone; NADH had less activity and ATP was not necessary for the reaction. 3. [4-14C]Androsta-5,16-dien-3β-ol, prepared biosynthetically from [4-14C]pregnenolone, was shown to be converted by boar testis preparations into androsta-4,16-dien-3-one (31%) if NAD+ was present or into 5α-androst-16-en-3β-ol (4%) if NADPH was present. 4. 17α-Hydroxyandrost-4-en-3-one and 3β,17α-dihydroxypregn-5-en-20-one were considered as possible precursors for androst-16-ene formation, but both were shown to be ineffective. 5. No radioactivity was incorporated into androst-5-en-3β-ol used to trap any corresponding 14C-labelled compound formed from [4-14C]pregnenolone.
Histone changes during degeneration and regeneration of the pancreas
Authors: Fitzgerald P. J. , Marsh W. H. , Ord Margery G. , Stocken L. A.
Published: August 2015

1. Changes in the microstructure of histones from the pancreas were followed in rats given ethionine while on a protein-free diet. The lysine-rich histone F1 and the other histones extracted subsequently with 250mm-hydrochloric acid were isolated. When the ethionine-treated rats returned to the stock diet, regeneration of the pancreas was accompanied by an immediate increase in phosphate content of the total histone extract from which histone F1 had been removed. After 3–4 days there was a further increase in the phosphate content of this extract, and also in that of histone F1. 2. Similar changes in the phosphate and thiol+disulphide content of pancreas histones other than F1 were produced by starvation and re-feeding.
The metabolism in vitro and hepatic microsomal interactions of some enantiomeric drug substrates
Authors: Hewick David S. , Fouts James R.
Published: August 2015

1. The metabolism in vitro and microsomal interactions of (+)-amphetamine, (−)-amphetamine, (+)-benzphetamine and (−)-benzphetamine were studied with hepatic microsomes from phenobarbitone-pretreated male rabbits. 2. (+)-Benzphetamine was N-demethylated 30–35% faster than (−)-benzphetamine, but the apparent Michaelis constants for the two enantiomers were similar. 3. (−)-Amphetamine was deaminated about 200% faster than (+)-amphetamine. 4. The benzphetamine enantiomers gave qualitatively and quantitatively identical type I microsomal difference spectra (peak, 390nm; trough, 425nm) indicating identical apparent binding affinities for microsomes and identical spectral changes at maxima (ΔEmax. values). 5. The amphetamine enantiomers gave qualitatively identical type II microsomal difference spectra (peak, 433nm; trough, 395nm). However, the type II spectral data indicated that (+)-amphetamine had a markedly higher apparent binding affinity than (−)-amphetamine for microsomes. The amphetamine enantiomers gave identical ΔEmax. values. 6. The benzphetamine enantiomers (0.5mm) enhanced the rate of microsomal cytochrome P-450 reduction by NADPH by 400–500%, (+)-benzphetamine enhancing the rate 20–25% more than (−)-benzphetamine. 7. The amphetamine enantiomers decreased the rate of microsomal cytochrome P-450 reduction by NADPH. At a concentration of 2mm, (+)-amphetamine decreased the rate more than (−)-amphetamine. 7. All four enantiomers enhanced microsomal NADPH oxidation.
Studies on the phospholipases of rat intestinal mucosa
Authors: Subbaiah P. V. , Ganguly J.
Published: August 2015

1. Subcellular distribution and characteristics of different phospholipases of rat intestinal mucosa were studied. 2. The presence of free fatty acid was necessary for the maximal hydrolysis of lecithin (phosphatidylcholine), but there was no accumulation of lysolecithin (1 or 2-acylglycerophosphorylcholine);lysolecithin accumulated when the reaction was carried out in the presence of sodium deoxycholate and at or above pH8.0. 3. The fatty acid-activated phospholipase B as well as lysolecithinase showed optimum activity at pH6.5, whereas for the phospholipase A it was about pH8.6. 4. The bulk of the phospholipase A was present in the microsomal fraction, whereas the phospholipase B and lysolecithinase activities were distributed between the microsomal and soluble fractions of the mucosal homogenate. 5. Phospholipase A was equally distributed between the brush border and brush-border-free particulate fraction, with the brush border having highest specific activity, whereas the other two activities were distributed between the brush-border-free particulate and soluble fractions. 6. Various treatments showed marked differences between the phospholipase A and phospholipase B activities, but not between phospholipase B and lysolecithinase activities. 7. By using (β[1-14C]-oleoyl) lecithin it was shown that the mucosal phospholipase A was specific for the β-ester linkage of the lecithin molecule.
Iodination of glyceraldehyde 3-phosphate dehydrogenase
Authors: Thomas Jean O. , Harris J. Ieuan
Published: August 2015

1. A high degree of homology in the positions of tyrosine residues in glyceraldehyde 3-phosphate dehydrogenase from lobster and pig muscle, and from yeast, prompted an examination of the reactivity of tyrosine residues in the enzyme. 2. Iodination of the enzyme from lobster muscle with low concentrations of potassium tri-[125I]-iodide led to the identification of tyrosine residues of differing reactivity. Tyrosine-46 appeared to be the most reactive in the native enzyme. 3. When the monocarboxymethylated enzyme was briefly treated with small amounts of iodine, iodination could be confined almost entirely to tyrosine-46 in the lobster enzyme; tyrosine-39 or tyrosine-42, or both, were also beginning to react. 4. These three tyrosine residues were also those that reacted most readily in the carboxymethylated pig and yeast enzymes. 5. The difficulties in attaining specific reaction of the native enzyme are considered. 6. The differences between our results and those of other workers are discussed.
A convenient synthesis of labelled rhodopsin and studies on its active site
Authors: Hirtenstein M. D. , Akhtar M.
Published: August 2015

Digitonin solutions of labelled rhodopsin, containing 3H in the retinyl moiety, were prepared by two related methods. Labelled rhodopsin was also prepared for the first time in cetyltrimethylammonium bromide and purified by column chromatography. It was shown that only certain rhodopsin preparations on denaturation in the dark and the reduction with sodium borohydride gave up to 60% of the radioactivity in a fraction characterized as N-retinylphosphatidylethanolamine. Such preparations also gave a lipid-linked retinyl moiety at the metarhodopsin-I stage, but, as expected, a protein-linked retinyl moiety at the metarhodopsin-II stage. Other preparations however, gave exclusively protein-bound radioactivity at the native-rhodopsin, metarhodopsin-I and metarhodopsin-II stages. It is therefore conceivable that the formation of N-retinylphosphatidylethanolamine is due to a non-enzymic reaction resulting from the transfer of the retinyl moiety from its native site to an amino group of a favourably oriented phospholipid molecule. The only firmly established aspect of the rhodopsin active site remains the demonstration in our previous work that at the metarhodopsin-II stage the retinyl moiety is linked to an ∈-amino group of lysine. On the basis of chemical reactivity it is argued that the light-induced conversion of rhodopsin into metarhodopsin II involves a profound conformational change resulting in the dislocation of the retinylideneiminium chromophore from a non-polar environment in rhodopsin to a polar environment in metarhodopsin II.
Fatty acid metabolism in the perfused rat liver
Authors: Krebs H. A. , Hems R.
Published: August 2015

1. The formation of acetoacetate, β-hydroxybutyrate and glucose was measured in the isolated perfused rat liver after addition of fatty acids. 2. The rates of ketone-body formation from ten fatty acids were approximately equal and independent of chain length (90–132μmol/h per g), with the exception of pentanoate, which reacted at one-third of this rate. The [β-hydroxybutyrate]/[acetoacetate] ratio in the perfusion medium was increased by long-chain fatty acids. 3. Glucose was formed from all odd-numbered fatty acids tested. 4. The rate of ketone-body formation in the livers of rats kept on a high-fat diet was up to 50% higher than in the livers of rats starved for 48h. In the livers of fat-fed rats almost all the O2 consumed was accounted for by the formation of ketone bodies. 5. The ketone-body concentration in the blood of fat-fed rats rose to 4–5mm and the [β-hydroxybutyrate]/[acetoacetate] ratio rose to 11.5. 6. When the activity of the microsomal mixed-function oxidase system, which can bring about ω-oxidation of fatty acids, was induced by treatment of the rat with phenobarbitone, there was no change in the ketone-body production from fatty acids, nor was there a production of glucose from even-numbered fatty acids. The latter would be expected if ω-oxidation occurred. Thus ω-oxidation did not play a significant role in the metabolism of fatty acids. 7. Arachidonate was almost quantitatively converted into ketone bodies and yielded no glucose, demonstrating that gluconeogenesis from poly-unsaturated fatty acids with an even number of carbon atoms does not occur. 8. The rates of ketogenesis from unsaturated fatty acids (sorbate, undecylenate, crotonate, vinylacetate) were similar to those from the corresponding saturated fatty acids. 9. Addition of oleate together with shorter-chain fatty acids gave only a slightly higher rate of ketone-body formation than oleate alone. 10. Glucose, lactate, fructose, glycerol and other known antiketogenic substances strongly inhibited endogenous ketogenesis but had no effects on the rate of ketone-body formation in the presence of 2mm-oleate. Thus the concentrations of free fatty acids and of other oxidizable substances in the liver are key factors determining the rate of ketogenesis.
Nitrite and hydroxylamine reduction in higher plants. Fractionation, electron donor and substrate specificity of leaf enzymes, principally from vegetable marrow (Cucurbita pepo L.)
Authors: Hucklesby D. P. , Hewitt E. J.
Published: August 2015

Nitrite reductase was purified between 760- and 1300-fold from vegetable marrow (Cucurbita pepo L.) and residual hydroxylamine reductase activity was low or negligible by comparison. With ferredoxin as electron donor, nitrite loss and ammonia formation at pH7.5 were stoicheiometrically equivalent. Crude nitrite reductase preparations showed negligible activity with NADPH as electron donor maintained in the reduced state by glucose 6-phosphate, whereas by comparison, activity was high when either ferredoxin or benzyl viologen were also present and reduced by the NADPH–glucose 6-phosphate system, whereas FMNH2 produced variable and relatively low activity under the same conditions. At pH values below 7, non-enzymic reactions occurred between reduced benzyl viologen and nitrite, and intermediate reduction products were inferred to be produced instead of ammonia. Activity with ferredoxin (0.1mm), reduced by chloroplast grana in the light, was 25 times that produced with ferredoxin (40μm) reduced with NADPH and glucose 6-phosphate. For an approximate molecular weight 61000–63000 derived by chromatography on Sephadex G-100 and G-200, and a specific activity of 46μmol of nitrite reduced/min per mg of protein with light and chloroplast grana, a minimum turnover number of 3×103mol of nitrite reduced/min per mol of enzyme was found. Two hydroxylamine reductases were separated on Sephadex gels. One (HR1) was initially associated with nitrite reductase during gel filtration but disappeared during later fractionation. This HR1 fraction showed nearly comparable activity with reduced benzyl viologen, ferredoxin or FMNH2. The other (HR2), of molecular weight approx. 35000, reacted with reduced benzyl viologen but showed negligible activity with ferredoxin or NADPH. Activity with FMNH2 was associated with an irregular trailing boundary during gel filtration, with much diminished activity in the HR2 region. Activity with NADPH was about 30% of that with FMNH2, reduced benzyl viologen or ferredoxin and was considered to reside in fraction HR1. Hydroxylamine yielded ammonia under all assay conditions. No activity with hyponitrite or sulphite was observed with reduced benzyl viologen as electron donor in either the nitrite reductase or the hydroxylamine reductase systems, but pyruvic oxime produced about 4% of the activity of hydroxylamine.
Phospholipid metabolism during regeneration of rat liver
Authors: Fex Göran
Published: August 2015

The concentration and composition of phospholipids and mitotic activity in regenerating rat liver were studied. (1) The total amount of liver phospholipid increased approximately linearly during 48h after operation but without change in the relative concentrations of individual phospholipids. (2) The appearance of mitoses 30h after operation was accompanied by an increased incorporation of 32P into the liver phospholipids. (3) The regenerating livers incorporated a higher percentage of the label into the phosphatidylserine+phosphatidylinositol fraction than those of control rats. The percentage of the label incorporated into phosphatidylethanolamine in these livers increased but decreased in the phosphatidylcholine.
The glycosaminoglycans of human tracheobronchial cartilage
Authors: Mason R. M. , Wusteman F. S.
Published: August 2015

1. The glycosaminoglycans of human tracheobronchial cartilages from subjects of various ages were liberated by proteolysis of the tissue and purified by ion-exchange chromatography. Purified glycosaminoglycans were fractionated on Dowex 1 resin and cetylpyridinium chloride was used to separate chondroitin sulphates and keratan sulphates occurring in the same fraction. 2. The total chondroitin sulphate content of the cartilages decreased linearly with increasing age. Age-dependent changes in the chemical heterogeneity of chondroitin sulphate were observed, a low-sulphated compound making up 25% of the total glycosaminoglycan at birth but rapidly diminishing in content during the first 6 months of life. Of the total chondroitin sulphate the 6-isomer became rather more prominent than the 4-isomer with increasing age. 3. The total keratan sulphate content of the cartilages increased from trace amounts only at birth to a plateau value by the beginning of the fifth decade. Of the total keratan sulphate approx. 70% was due to a high-molecular-weight compound with a sulphate/hexosamine ratio of 1.5–1.8: 1.0. The degree of sulphation varied between compounds isolated from different individuals. The remaining 30% of the keratan sulphate appeared to be intimately associated with chondroitin 6-sulphate and could only be separated from it after treatment with 0.45m-potassium hydroxide. The hybrid glycosaminoglycans were of lower molecular weight and had a lower sulphate/hexosamine ratio than the major keratan sulphate compound.
Galactose-specific messenger ribonucleic acid contents in Escherichia coli
Authors: Gosden J. R. , Irving M. I. , Bishop J. O.
Published: August 2015

A method is described for measuring the proportion of galactose-specific mRNA (gal-mRNA) in the total RNA extracted from pulse-labelled cells of Escherichia coli K12, by DNA–RNA hybridization with DNA prepared from bacteriophage λdg. RNA from wild-type E. coli was compared with RNA from a homogenote carrying the gal operon both in the chromosome and in a substituted sex-factor, and with RNA from a deletion strain that carried the galactose operon only in the exogenote. In each case the cultures were induced with fucose. Under these conditions the amount of gal-mRNA was found to be proportional to the content of galactokinase in the different cultures, and to the gene frequency. The amounts of gal-mRNA in an Oc mutant and an R− mutant were also proportional to the observed contents of galactokinase. In cultures repressed for the enzymes of the galactose operon with thiomethylgalactoside, the content of gal-mRNA was higher than expected from the content of galactokinase. Possible explanations of this finding are discussed.
The chemistry of the collagen cross-links. The absence of reduction of dehydrolysinonorleucine and dehydrohydroxylysinonorleucine in vivo
Authors: Bailey A. J. , Peach Catherine M.
Published: August 2015

Two aldimine bonds have been shown to be present as stabilizing cross-links in intact collagen fibres from soft tissues: dehydrohydroxylysinonorleucine as a major component and dehydrolysinonorleucine being present in trace quantities. In the highly insoluble collagens less dehydrohydroxylysinonorleucine is present but the proportion of dehydrolysinonorleucine increases. In elastin the latter aldimine is reduced in vivo to give a more stable cross-link but no comparable reduction could be detected with either of the aldimines present in collagen.
Formation of malate from glyoxylate in animal tissues
Authors: Liang Chi-Chin , Ou Lo-Chang
Published: August 2015

1. Incubation of rat liver homogenate with [1-14C]glyoxylate, ATP and acetate shows a rapid sequential incorporation of radioactivity into malate, oxaloacetate and citrate. 2. In liver from normal rats the rate of the formation of each substance in question is higher than that in liver from thiamin-deficient rats. 3. The net accumulation of malate is greater with liver from thiamin-deficient rats. Its further metabolism is retarded, it is suggested, by inhibitors formed by a condensation of glyoxylate and oxaloacetate.
Haem synthesis during mitochondrogenesis in yeast
Authors: Jayaraman J. , Padmanaban G. , Malathi K. , Sarma P. S.
Published: August 2015

The activities of δ-aminolaevulate synthetase and δ-aminolaevulate dehydratase have been assayed in Saccharomyces cerevisiae during glucose repression and de-repression. δ-Aminolaevulate dehydratase increased concomitantly with the increase in oxygen uptake during the de-repression phase caused by the depletion of glucose in the medium. δ-Aminolaevulate synthetase showed an oscillatory behaviour and a spurt in its activity always preceded the increase in oxygen uptake. The activity of both the enzymes was lowered if the cells were incubated with glucose or cycloheximide, but not with chloramphenicol.
The effects of salicylate on the activity of rat liver tyrosine–2-oxoglutarate aminotransferase in vitro and in vivo
Authors: Badawy A. A.-B.
Published: August 2015

1. Salicylate, in concentrations of 0.25mm and above, enhances the basal activity of tyrosine–2-oxoglutarate aminotransferase in homogenates of rat liver incubated in the absence of added pyridoxal 5′-phosphate (endogenous activity). The effect is decreased by increasing the concentration of the cofactor. 2. The intraperitoneal administration of sodium salicylate enhances the activity of rat liver tyrosine aminotransferase; the major effect during the first hour being on the enzyme in the absence of added pyridoxal phosphate. Actinomycin D prevents the induction of the enzyme by cortisol and tryptophan. Induction by pyridoxine or salicylate is 50% inhibited by actinomycin D. The effects of the injections of various combinations of cortisol, pyridoxine and salicylate were also studied in the absence or presence of actinomycin D. 3. It is suggested that salicylate induces rat liver tyrosine aminotransferase by displacing its protein-bound cofactor and that a cofactor-type induction of the hepatic enzyme occurs in pyridoxine-treated rats.
Folic acid and the methylation of homocysteine by Bacillus subtilis
Authors: Salem A. R. , Pattison J. R. , Foster M. A.
Published: August 2015

1. Cell-free extracts of Bacillus subtilis synthesize methionine from serine and homocysteine without added folate. The endogenous folate may be replaced by tetrahydropteroyltriglutamate or an extract of heated Escherichia coli for the overall C1 transfer, but tetrahydropteroylmonoglutamate is relatively inactive. 2. Extracts of B. subtilis contain serine transhydroxymethylase and 5,10-methylenetetrahydrofolate reductase, which are non-specific with respect to the glutamate content of the folate substrates. Methyl transfer to homocysteine requires a polyglutamate folate as methyl donor. These properties are not affected by growth of the organism with added vitamin B12. 3. The synthesis of methionine from 5-methyltetrahydropteroyltriglutamate and homocysteine has the characteristics of the cobalamin-independent reaction of E. coli. No evidence for a cobalamin-dependent transmethylation was obtained. 4. S-Adenosylmethionine was not a significant precursor of the methyl group of methionine with cell-free extracts, neither was S-adenosylmethionine generated by methylation of S-adenosylhomocysteine by 5-methyltetrahydrofolate. 5. A procedure for the isolation and analysis of folic acid derivatives from natural sources is described. 6. The folates isolated from lysozyme extracts of B. subtilis are sensitive to folic acid conjugase. One has been identified as 5-formyltetrahydropteroyltriglutamate; the other is possibly a diglutamate folate. 7. A sequence is proposed for methionine biosynthesis in B. subtilis in which methyl groups are generated from serine and transferred to homocysteine by means of a cobalamin-independent pathway mediated by conjugated folate coenzymes.
Studies of the ethylation of rat liver transfer ribonucleic acid after administration of l-ethionine
Authors: Pegg A. E.
Published: August 2015

1. The ethylated nucleosides present in tRNA isolated from the livers of rats treated with 0.5g of l-ethionine/kg body wt. were investigated. Evidence that this tRNA contained N2-ethylguanine, N2N2-diethylguanine, N2-ethyl-N2-methylguanine, 7-ethylguanine, two ethylated pyrimidines and ethylated ribose groups was obtained. 2. Ethylation of bacterial tRNA was catalysed by extracts containing tRNA methylases prepared from rat liver by using S-adenosyl-l-ethionine as an ethyl donor, but the rate of ethylation was 20 times less than the rate of methylation with S-adenosyl-l-methionine as a methyl donor. 3. The principal product of such ethylation in vitro was N2-ethylguanine and traces of the other ethylated guanines and pyrimidines found in tRNA isolated from rats treated with ethionine in vivo were also found. 1-Ethyladenine was not formed, although 1-methyl-adenine is a major product of methylation of bacterial tRNA by these extracts, and 1-ethyladenine was not present in the rat liver tRNA isolated from ethionine-treated animals. 4. After injection of actinomycin D (15mg/kg body wt.) or l-methionine (1.0g/kg body wt.) before the ethionine, ethylation of tRNA was diminished by about 80% but not completely abolished. Administration of 1-aminocyclopentanecarboxylic acid (2.5g/kg body wt.) to inhibit the formation of S-adenosyl-l-ethionine inhibited ethylation of tRNA by 44%. 5. These results suggest that not all of the ethylation of tRNA that occurs in the livers of rats treated with ethionine is mediated by the action of tRNA methylases acting with S-adenosyl-l-ethionine as a substrate, but that this pathway does occur and accounts for a major part of the observed ethylation. 6. The results are discussed with reference to ethionine-induced hepatocarcinogenesis.
The use of deoxyfluoro-d-glucopyranoses and related compounds in a study of yeast hexokinase specificity
Authors: Bessell E. M. , Foster A. B. , Westwood J. H.
Published: August 2015

1. 2-Deoxy-2-fluoro-d-glucose, 2-deoxy-2-fluoro-d-mannose and 2-deoxy-2,2-difluoro-d-arabino-hexose are good substrates for yeast hexokinase. 2. 3-Deoxy-3-fluoro-d-glucose and 4-deoxy-4-fluoro-d-glucose are poor substrates and have very similar Km values (8×10−2m). 3. Neither α- nor β-d-glucopyranosyl fluoride is a substrate or inhibitor. 4. Studies with 2-chloro-2-deoxy- and 2-O-methyl derivatives of d-glucose and d-mannose have revealed that little chemical modification is possible at position 2 without substantial loss in substrate binding. 5. The variation in the value of Km for the d-hexose derivatives was associated with a corresponding change in the value of Km for MgATP2− showing that the binding of MgATP2− is modified by the binding of the sugar.
Properties of the ribonucleic acid bacteriophage ZIK/1 coat protein and its synthesis in an Escherichia coli cell-free system
Authors: Robinson J. W.
Published: August 2015

The coat protein subunit of the RNA bacteriophage ZIK/1 has a molecular weight of 12100 and does not contain histidine, methionine and cysteine. The amino acid composition of the coat protein is different from that of other RNA bacteriophage coat proteins. Bacteriophage ZIK/1 belongs to a class of RNA bacteriophages distinct from the f2 type, which lack histidine in their coat proteins, and the Qβ type, which lack histidine and methionine. Bacteriophage ZIK/1 RNA is an efficient template in the Escherichia coli cell-free system producing coat protein as the major product and a number of non-coat proteins. This result is similar to that obtained with RNA from f2-type bacteriophages. It is probable that the genomes of RNA bacteriophages are structurally similar and that differences between the types of RNA bacteriophage arise from minor differences in RNA sequence.
The control of ribonucleic acid synthesis in bacteria. Polymerization rates for ribonucleic acids in amino acid-starved relaxed and stringent auxotrophs of Escherichia coli
Authors: Gray W. J. H. , Vickers T. G. , Midgley J. E. M.
Published: August 2015

Polymerization rates of newly formed chains of various RNA fractions were measured in Escherichia coli CP78 (RCstr) and CP79 (RCrel) multiple amino acid auxotrophs, deprived of four amino acids essential for growth. Immediately after the onset of severe amino acid deprivation, in RCstr strains the rate of labelling of RNA by exogenous nucleotide bases was greatly diminished. At first, the initiation of new RNA chains declined faster than the rate of polymerization in RCstr organisms, but as starvation proceeded the rate of polymerization was eventually lowered to about 10% of that found during normal growth. In strain CP79 (RCrel), on the other hand, chain-polymerization rates were unaffected by amino acid withdrawal. Artificial depletion of the intracellular purine nucleotide pools in RCstr or RCrel strains by trimethoprim, before the onset of amino acid deprivation, showed that in the RCstr, but not the RCrel strain, amino acid withdrawal gave rise to an inability of the cells to utilize exogenously supplied purine or pyrimidine bases for RNA synthesis. During a prolonged starvation, the observed 100-fold decrease in the total rate of incorporation of exogenous nucleotide bases into the RNA of RCstr organisms was ascribed to a combination of a tenfold decrease in the overall rate of RNA chain polymerization, at least a fivefold decrease in the ability of the cells to utilize exogenous bases and a preferential inhibition of initiation of stable RNA chains. None of these changes occurred in the corresponding RCrel strain.
Interrelationship and control of glucose metabolism and lipogenesis in isolated fat-cells. Effect of the amount of glucose uptake on the rates of the pentose phosphate cycle and of fatty acid synthesis
Authors: Kather H. , Rivera M. , Brand K.
Published: August 2015

In order to study the quantitative relationship between fatty acid synthesis and pentose phosphate-cycle activity under different hormonal and dietary conditions affecting the extent of glucose uptake, cells isolated from rat epididymal adipose tissue were incubated in bicarbonate buffer containing [U-14C]-, [1-14C]- or [6-14C]-glucose. From the amount of glucose taken up, the production of lactate and pyruvate, and the incorporation of 14C from differently labelled [14C]glucose into CO2, fatty acids and glyceride glycerol, the rates of glucose metabolism via different pathways and the extent of lipogenesis under various experimental conditions were determined. The contribution of the pentose phosphate-cycle to glucose metabolism under normal conditions was calculated to be 8%. Starvation and re-feeding, and the presence of insulin, caused an enhancement of glucose uptake, pentose phosphate-cycle activity and fatty acid synthesis. Plots of both pentose phosphate-cycle activity and fatty acid synthesis versus glucose uptake revealed that the extent of glucose uptake, over a wide range, determines the rates of fatty acid synthesis and glucose metabolism via the pentose phosphate cycle. A balance of formation and production of nicotinamide nucleotides in the cytoplasm was established. The total amount of cytoplasmic NADH and NADPH formed was only in slight excess over the hydrogen equivalents required for the synthesis of fatty acids, glyceride glycerol and lactate. Except in cells from starved animals, the pentose phosphate cycle was found to provide only about 60% of the NADPH required for fatty acid synthesis. The results are discussed with respect to an overall control of the different metabolic and biosynthetic reactions in the fat-cells by the amount of glucose transported into the cell.
Inhibition of hepatic deoxyribonucleic acid-dependent ribonucleic acid polymerases by the exotoxin of Bacillus thuringiensis in comparison with the effects of α-amanitin and cordycepin
Authors: Smuckler Edward A. , Hadjiolov Asen A.
Published: August 2015

The action of Bacillus thuringiensis exotoxin, a structural analogue of ATP, on mouse liver DNA-dependent RNA polymerases was studied and its effects were compared with those of α-amanitin and cordycepin. (1) Administration of exotoxin in vivo caused a marked decrease in RNA polymerase activity of isolated nuclei at various concentrations of Mg2+, Mn2+and (NH4)2SO4. A similar action was recorded after addition of exotoxin to isolated nuclei from control or exotoxin-treated mice. (2) Chromatographic separation of nuclear RNA polymerases from mice treated in vivo with exotoxin showed a drastic decrease of the peak of nucleoplasmic RNA polymerase, whereas the peak of nucleolar RNA polymerase remained unaltered. The same effect was observed after administration of α-amanitin in vivo, but cordycepin did not alter the relative amounts of the two main RNA polymerase peaks. (3) Administration of exotoxin in vivo did not alter the template activity of isolated DNA or chromatin tested with different fractions of RNA polymerase from control or exotoxin-treated mice. (4) Addition of exotoxin to isolated liver RNA polymerases inhibited both enzyme fractions. However, the α-amanitin-sensitive RNA polymerase was also 50–100-fold more sensitive to exotoxin inhibition than was the α-amanitin-insensitive RNA polymerase. Kinetic analysis indicated the exotoxin produces a competitive inhibition with ATP on the nucleolar enzyme, but a mixed type of inhibition with nucleoplasmic enzyme. The results obtained indicate that the B. thuringiensis exotoxin inhibits liver RNA synthesis by affecting nuclear RNA polymerases, showing a preferential inhibition of the nucleoplasmic α-amanitin-sensitive RNA polymerase.
Uptake of amino acids and nucleic acid precursors by regenerating rat liver
Authors: Ord Margery G. , Stocken L. A.
Published: August 2015

1. At 0.5–1.0h after partial hepatectomy the intracellular acid-soluble fraction of rat liver took up twice as much radioactivity from [3H]orotic acid, [3H]uridine and [3H]thymidine as did similar fractions from sham-operated animals. This increase in penetration was not prevented by adrenalectomy or actinomycin, both of which decreased precursor uptake into nuclear RNA at this time. The increase in entry was still shown by thymidine at 22h after operation, at the height of the S period. Adenine penetration was not increased 1h after partial hepatectomy. 2. Plasma concentrations of ornithine and possibly methionine, tyrosine and lysine were raised 1.5h after partial hepatectomy. [3H]Lysine entry into regenerating liver at this time was increased by 60% [3H]valine uptake was unaffected. Intracellular amounts of tyrosine, phenylalanine and ornithine in the liver were also increased. 3. The relation of these events to the start of liver regeneration is discussed.
Membrane proteins of chromaffin granules. Dopamine β-hydroxylase, a major constituent
Authors: Hörtnagl Heide , Winkler H. , Lochs H.
Published: August 2015

1. Soluble lysates and membranes were prepared from chromaffin granules isolated from bovine adrenal medulla. The detergent N-cetylpyridinium chloride was used for solubilizing the membrane proteins, including the membrane-bound dopamine (2,4-dihydroxyphenethylamine) β-hydroxylase. The solubilized proteins were fractionated by Sephadex chromatography in the presence of N-cetylpyridinium chloride. The major component of the membrane proteins, i.e. chromomembrin A, was identified as the enzyme dopamine β-hydroxylase. 2. The addition of N-cetylpyridinium chloride to the soluble lysate caused precipitation of up to 96% of the proteins, but only a small proportion of the dopamine β-hydroxylase activity was precipitated. The only protein demonstrable in the supernatant by polyacrylamide-gel electrophoresis was the protein that has a lower mobility than chromogranin A in disc gel electrophoresis. This component has been identified previously as dopamine β-hydroxylase. Thus, this method provides an extremely simple isolation procedure for dopamine β-hydroxylase. 3. A comparison of the membrane-bound and soluble dopamine β-hydroxylases revealed the identity of these two preparations. Both were activated by N-cetylpyridinium chloride, they migrated identically in polyacrylamide-gel electrophoresis, their amino acid composition was very similar and an immunological cross-reaction could be demonstrated.
A reaction of glucose with peptides
Authors: Dixon Henry B. F.
Published: August 2015

Valylhistidine (Val-His) reacts with glucose (Glc) in a mixture of pyridine and acetic acid to form glucosylvalylhistidine (Glc-Val-His). The pK of the α-amino group is thereby lowered to about 5.6 as judged by electrophoretic mobility. The reaction: [Formula: see text] also occurs in an aqueous solution of pyridine and acetic acid of pH6.2 at 50°C, in which it exhibits a half-time of about 30h and a dissociation constant of about 0.3m. Isoleucyltyrosine and glucose react similarly in aqueous solution. The Glc-Val-His has the chromatographic, electrophoretic and chemical properties reported by Holmquist & Schroeder (1966a) for the substance released by proteolysis from the N-terminus of the β-chains of haemoglobin AIc; the value of the dissociation constant means that the concentration of haemoglobin AIc found naturally could be explained by reaction of haemoglobin A with the blood glucose.
A study of the subunit structure and the thiol reactivity of bovine liver uridine diphosphate glucose dehydrogenase
Authors: Gainey P. A. , Pestell T. C. , Phelps C. F.
Published: August 2015

1. The amino acid analysis of UDP-glucose dehydrogenase is reported. 2. N-Terminal-group analysis indicates only one type of N-terminal amino acid, methionine, to be present. 3. Peptide ‘mapping’ in conjunction with the amino acid analysis indicates that the subunits of the enzyme are similar if not identical. 4. The various kinetic classes of thiol group were investigated by reaction with 5,5′-dithiobis-(2-nitrobenzoate). 5. NAD+, UDP-glucose and UDP-xylose protect the two rapidly reacting thiol groups of the hexameric enzyme. 6. Inactivation of the enzyme with 5,5′-dithiobis-(2-nitrobenzoate) indicates the involvement of six thiol groups in the maintenance of enzymic activity. 7. The pH-dependence of UDP-xylose inhibition of the enzyme was investigated. 8. The group involved in the binding of UDP-xylose to the protein has a heat of ionization of about 33kJ/mol and a pK of 8.4–8.6. 9. It is suggested that UDP-xylose has a cooperative homotropic effect on the enzyme.
The metabolites of cyclohexylamine in man and certain animals
Authors: Renwick A. G. , Williams R. T.
Published: August 2015

1. [1-14C]Cyclohexylamine hydrochloride was synthesized and given orally or intraperitoneally to rats, rabbits and guinea pigs (dose 50–500mg/kg) and orally to humans (dose 25 or 200mg/person). The 14C is excreted mainly in the urine, most of the excretion occurring in the first day after dosing. Only small amounts (1–7%) are found in the faeces. 2. In the rat, guinea pig and man, the amine is largely excreted unchanged, only 4–5% of the dose being metabolized in 24h in the rat and guinea pig and 1–2% in man. In the rabbit about two-thirds of the dose is excreted unchanged and about 30% is metabolized. 3. In the rat, five minor metabolites were found, namely cyclohexanol (0.05%), trans-3- (2.2%), cis-4- (1.7%), trans-4- (0.5%) and cis-3-aminocyclohexanol (0.1% of the dose in 24h). 4. In the rabbit, eight metabolites were identified, namely cyclohexanol (9.3%), trans-cyclohexane-1,2-diol (4.7%), cyclohexanone (0.2%), cyclohexylhydroxylamine (0.2%) and trans-3- (11.3%), cis-3- (0.6%), trans-4- (0.4%) and cis-4-aminocyclohexanol (0.2%). 5. In the guinea pig, six minor metabolites were found, namely cyclohexanol (0.5%), trans-cyclohexane-1,2-diol (2.5%) and trans-3- (1.2%), cis-3- (0.2%), trans-4- (0.2%) and cis-4-aminocyclohexanol (0.2%). 6. In man only two metabolites were definitely identified, namely cyclohexanol (0.2%) and trans-cyclohexane-1,2-diol (1.4% of the dose), but man had been given a smaller dose (3mg/kg) than the other species (50mg/kg). 7. The hydroxylated metabolites of cyclohexylamine were excreted in the urine in both free and conjugated forms. 8. Although cyclohexylamine is metabolized to only a minor extent, in rats the metabolism was mainly through hydroxylation of the cyclohexane ring, in man by deamination and in guinea pigs and rabbits by ring hydroxylation and deamination.
Statistical analysis of radioimmunoassay data
Authors: Healy Michael J. R.
Published: August 2015

The statistical processing of radioimmunoassay data is discussed, with special emphasis on fitting the standard curve, screening the data for aberrant readings and combining separate estimations from a single sample.
Effect of temperature and pH on carbamoylation and phosphorylation of serum cholinesterases. Theoretical interpretation of activation energies in complex reactions
Authors: Simeon V. , Reiner E. , Vernon C. A.
Published: August 2015

1. The effect of temperature and pH was studied on the kinetics of inhibition of horse serum and human serum cholinesterase by four organophosphorus compounds and five carbamates. 2. For all compounds, and at each pH and temperature, the inhibition followed the kinetics of a bimolecular reaction with the inhibitor in excess, and with a negligible concentration of the Michaelis complex. 3. The second-order rate constants (ka) for inhibition of human serum cholinesterase by one organophosphate and one carbamate increased from 5° to 40°C with an apparent activation energy of 46kJ/mol (11kcal/mol). 4. The ka constant for inhibition of horse serum cholinesterase increased with temperature from 5° to 30°C, and then decreased from 30° to 40°C. The theoretical interpretation of such an unusual effect of temperature is derived. 5. The increase of ka with pH (human serum cholinesterase) followed the dissociation curve for a single group on the enzyme (pK7.5). 6. Rate constants for decarbamoylation (k+3) were determined, and the time-course of inhibition was calculated from the ka and k+3 constants.
Quantitative aspects of fatty acid biohydrogenation, absorption and transfer into milk fat in the lactating goat, with special reference to the cis- and trans-isomers of octadecenoate and linoleate
Authors: Bickerstaffe R. , Noakes D. E. , Annison E. F.
Published: August 2015

1. Surgically prepared lactating goats were used to obtain quantitative information on the biohydrogenation and absorption of dietary fat, and on the mammary uptake and transfer into milk fat of the complex mixture of cis- and trans-isomers of octadecenoate that arise during ruminal biohydrogenation. 2. About 90% of dietary linolenate, linoleate and oleate was hydrogenated in the rumen, and the availability to the animals of the essential fatty acid, linoleate, represented only 0.5–1.5% of the total dietary energy. 3. The intra-ruminal administration of 14C-labelled linolenate and linoleate showed that these acids were not absorbed from the rumen, in agreement with previous work. 4. No selectivity was observed in the metabolism of the geometrical and positional isomers of octadecenoate: their rates of absorption from the small intestine, transfer into lymph, uptake by the mammary gland and appearance in milk fat were similar. 5. The desaturase activity of intestinal epithelium was demonstrated by the appearance in lymph of [1-14C]oleate after the addition of [1-14C]stearate to the small intestine.
Anthranilate synthase/anthranilate 5-phosphoribosyl 1-pyrophosphate phosphoribosyltransferase from Aerobacter aerogenes
Authors: Egan A. F. , Gibson F.
Published: August 2015

1. Anthranilate synthase and phosphoribosyltransferase from Aerobacter aerogenes purify simultaneously and sediment together on sucrose gradients, showing that they occur as an enzyme aggregate. Both activities of the intact aggregate are subject to inhibition by tryptophan. 2. By using appropriate auxotrophic mutants it was shown that an intact active enzyme aggregate is formed when the components come from separate mutant strains. An intact active aggregate can also be formed when one component is from Escherichia coli and the other from A. aerogenes. 3. Phosphoribosyltransferase of A. aerogenes is active when not in an aggregate with anthranilate synthase, but is not subject to tryptophan inhibition, indicating that the inhibitor site is on the anthranilate synthase component. 4. Anthranilate synthase can be active and sensitive to tryptophan inhibition when complexed with an inactive phosphoribosyltransferase. 5. Kinetic studies on the anthranilate synthase activity show that tryptophan is a competitive inhibitor with respect to chorismate and a non-competitive inhibitor with respect to either glutamine or NH4+ ions. This is consistent with a sequential mechanism of the ordered type in which chorismate is the first reactant.