bluuebunny/crossref_metadata_2025 huggingface dataset:
The arrangement of proteins on the deoxyribonucleic acid in chromatin
Authors: Itzhaki Ruth F.
Published: August 2015

The arrangement of the protein component on the DNA of the chromatin complex was studied by comparing the rate of release of oligonucleotides and of protein after addition of deoxyribonuclease I and deoxyribonuclease II to rat thymus chromatin. Also the action of deoxyribonuclease I on normal chromatin and on chromatin depleted of non-histone protein was compared, to elucidate the role of the latter protein in chromatin structure. As a preliminary to the above, the rate of action of deoxyribonuclease I on DNA and on chromatin at the same DNA concentration, and the dependence of the action of this enzyme on the Mg2+ concentration, were studied. It was found that: (1) little if any DNA in chromatin is present in extensive, truly ‘free’ zones, i.e. completely uncovered by protein; (2) at relatively low concentrations of added Mg2+, deoxyribonuclease I degrades chromatin more rapidly than DNA; (3) the non-histone protein is not attached directly to the DNA in chromatin.
Biosynthesis of the carbohydrate portions of immunoglobulin M
Authors: Parkhouse R. M. E. , Melchers Fritz
Published: August 2015

Incorporations of radioactive mannose, galactose and fucose into MOPC 104E mouse plasma-cell tumour suspensions suggest a stepwise addition of carbohydrate residues to immunoglobulin M (IgM) during the process of secretion. Mannose and glucosamine residues are added at an early stage, whereas galactose and fucose are added just before, or at the time that, IgM leaves the cell. Free light chains secreted in excess by the same tumour cells incubated with mannose, galactose or fucose contained barely detectable amounts of radioactivity.
Digestion of an exogenous protein by rat yolksac cultured in vitro
Authors: Williams K. E. , Lloyd J. B. , Davies M. , Beck F.
Published: August 2015

Yolk-sacs were removed from 17.5-day pregnant rats injected 2–5h previously with 125I-labelled bovine serum albumin. The specific activities of acid phosphatase and acid proteinase, and the specific radioactivities (trichloroacetic acid-insoluble and trichloroacetic acid-soluble) were measured in subcellular fractions prepared by homogenization and differential centrifugation. The conversion of acid-insoluble into acid-soluble radioactivity within cultured tissue was followed and the nature of the liberated products was investigated by gel chromatography. The results are consistent with the protein entering lysosomes and being digested there. The radiolabel was released chiefly as free iodotyrosine.
Metabolic control mechanisms in mammalian systems. Involvement of adenosine 3′:5′-cyclic monophosphate in androgen action
Authors: Singhal Radhey L. , Parulekar M. R. , Vijayvargiya R. , Robison G. Alan
Published: August 2015

1. The ability of exogenously administered cyclic AMP (adenosine 3′:5′-monophosphate) to exert andromimetic action on certain carbohydrate-metabolizing enzymes was investigated in the rat prostate gland and seminal vesicles. 2. Cyclic AMP, when injected concurrently with theophylline, produced marked increases in hexokinase, phosphofructokinase, glyceraldehyde phosphate dehydrogenase, pyruvate kinase, and two hexose monophosphate-shunt enzymes, as well as α-glycerophosphate dehydrogenase activity in accessory sexual tissues of castrated rats. The 6-N,2′-O-dibutyryl analogue of cyclic AMP caused increases of enzyme activity that were greater than those induced by the parent compound. 3. Time-course studies demonstrated that, whereas significant increases in the activities of most enzymes occurred within 4h after the injection of cyclic AMP, maximal increases were attained at 16–24h. 4. Increase in the activity of the various prostatic and vesicular enzymes was dependent on the dose of cyclic AMP; in most instances, 2.5mg of the cyclic nucleotide/rat was sufficient to elicit a statistically significant response. 5. Administration of cyclic AMP and theophylline also produced stimulation of enzyme activities in secondary sexual tissues of immature rats. 6. Cyclic AMP and theophylline did not affect significantly any of the enzymes studied in hepatic tissue. 7. Stimulation of various carbohydrate-metabolizing enzymes in the prostate gland and seminal vesicles by cyclic AMP was independent of adrenal function. 8. Concurrent treatment with actinomycin or cycloheximide prevented the cyclic AMP- and theophylline-induced increases in enzyme activities in both castrated and adrenalectomized–castrated animals. 9. Administration of a single dose of testosterone propionate (5.0mg/100g) to castrated rats caused a significant increase in cyclic AMP concentration in both accessory sexual tissues. 10. In addition, treatment with theophylline potentiated the effects of a submaximal dose of testosterone (1.0mg/100g) on all those prostatic and seminal-vesicular enzymes that are increased by exogenous cyclic AMP. 11. The evidence indicates that cyclic AMP may be involved in triggering the known metabolic actions of androgens on secondary sexual tissues of the rat.
Further studies on the glycerol teichoic acid of walls of Staphylococcus lactis I3. Location of the phosphodiester groups and their susceptibility to hydrolysis with alkali
Authors: Archibald A. R. , Baddiley J. , Heckels J. E. , Heptinstall S.
Published: August 2015

1. The teichoic acid from walls of Staphylococcus lactis I3 is readily degraded in dilute alkali. 2. Degradation proceeds by selective hydrolysis of that phosphodiester group attached to an alcoholic hydroxyl group of the N-acetylglucosamine and gives a repeating unit in high yield. 3. Further studies on a different repeating unit isolated by partial acid hydrolysis have shown that the glycerol diphosphate is attached to the 4-hydroxyl group of the N-acetylglucosamine and not to the 3-hydroxyl group as was proposed earlier. 4. The susceptibility towards hydrolysis by alkali of other structural types of teichoic acid has been examined and found to vary markedly according to their structure.
Metabolism of m-tert.-butylphenyl N-methylcarbamate in insects and mice
Authors: Douch P. G. C. , Smith J. N.
Published: August 2015

The metabolism of m-tert.-butylphenyl N-methylcarbamate was studied in mice and five species of insects. Both the tert.-butyl group and the N-methyl group were hydroxylated. The major phenolic metabolite was m-(β-hydroxy-tert.-butyl)phenol, which was identified by mass spectroscopy. Significant amounts of dihydroxy compounds were formed at a constant rate from the start of the enzymic oxidation process. The considerable species variation in the yields of the different types of oxidation products suggests that N-demethylation and oxidation of the tert.-butyl groups were catalysed by different enzymes. A microsomal NADPH-dependent enzyme also catalysed the splitting of the ester link in the insecticide.
The metabolism of 3,5-di-tert.-butylphenyl N-methylcarbamate in insects and by mouse liver enzymes
Authors: Douch P. G. C. , Smith J. N.
Published: August 2015

The oxidation of 3,5-di-tert.-butylphenyl N-methylcarbamate (Butacarb) has been studied in the flies Musca domestica and Lucilia sericata, grass grubs Costelytra zealandica and the mouse. In all species eleven oxidation products, which were formed by hydroxylation of the tert.-butyl groups and the N-methyl group, were detected.
Biochemical adaptation in rat liver in response to marginal oxygen toxicity
Authors: Gorman R. R. , Jordan J. P. , Simmons J. B. , Clarkson D. P.
Published: August 2015

1. Hepatic glucose 6-phosphate dehydrogenase activity was increased in rats exposed to 5lb/in2 (equivalent to 27000ft), 100% O2 when compared with control animals in a 14.7lb/in2 (sea level), air environment. Glyceraldehyde 3-phosphate dehydrogenase, isocitrate dehydrogenase, and succinate dehydrogenase were not affected by the 5lb/in2, 100% O2 environment. 2. Animals exposed to the hyperoxic environment consumed food, expired CO2 and gained weight at the same rate as normoxic control animals. Additionally, blood glucose and liver glycogen concentrations were unchanged in the hyperoxic animals. The only readily apparent physiological difference in the hyperoxic animals was a decreased haematocrit. 3. The increase in glucose 6-phosphate dehydrogenase was eliminated by the injection of actinomycin D or cycloheximide. 4. Expiration of 14CO2 from [1-14C]glucose was approximately the same in hyperoxic and normoxic rats. However, 14CO2 expiration from [6-14C]glucose was markedly decreased in the animals exposed to the hyperoxic environment. 5. Calculations of the relative importance of the pentose phosphate pathway versus the tricarboxylic acid cycle plus glycolysis indicated that the livers from animals in the 5lb/in2, 100% O2 environment metabolized twice as much carbohydrate by way of the pentose phosphate pathway as did those from the sea-level air control animals. 6. In livers of rats exposed to 5lb/in2, 100% O2 the concentrations of pyruvate, citrate and 2-oxoglutarate were increased, that of isocitrate was slightly elevated, whereas the concentrations of succinate, fumarate and malate were decreased. 7. An inactivation of both tricarboxylic acid cycle lipoate-containing dehydrogenases, pyruvate and 2-oxoglutarate, under hyperoxic conditions is proposed. 8. The adaptive significance of the induction of glucose 6-phosphate dehydrogenase and the resultant production of NADPH under hyperoxic conditions is discussed.
Characterization of two cell-wall polysaccharides from Fusicoccum amygdali
Authors: Obaidah M. A. , Buck K. W.
Published: August 2015

1. The nature of two polysaccharides (s020 values 6S and 2S respectively in 1m-sodium hydroxide), comprising a fragment (fraction BB, [α]D +236° in 1m-sodium hydroxide), previously isolated from cell walls of Fusicoccum amygdali, has been investigated. 2. Both the major (2S) and minor (6S) components were affected by incubation with α-amylase. The 6S polysaccharide was also attacked by exo-β-(1→3)-glucanase, which is evidence that it contained both α-(1→4)- and β-(1→3)-glucopyranose linkages. By fractionation of the products of α-amylase-treated fraction BB it was possible to obtain a water-insoluble polysaccharide, fraction P ([α]D +290° in 1m-sodium hydroxide, 67% of fraction BB) and a water-soluble polysaccharide, fraction Q ([α]D +16° in 1m-sodium hydroxide, 11% of fraction BB), both of which sedimented as single boundaries with s020 values (in 1m-sodium hydroxide) of 1.7S and 4.6S respectively. 3. Evidence from periodate oxidation, methylation analysis, i.r. spectroscopy and partial acid hydrolysis showed that fraction P consisted of linear chains of α-(1→3)-glucopyranose units with blocks of one or two α-(1→4)-glucopyranose units interspersed at intervals along the main chain. The 2S polysaccharide, from which fraction P is derived, evidently also contains longer blocks of α-(1→4)-glucopyranose units, that are susceptible to α-amylase action. 4. Fraction Q consisted of glucose (88%) with small amounts of galactose, mannose and rhamnose. Evidence from digestion with exo- and endo-β-(1→3)-glucanases, periodate oxidation and methylation analysis suggests that fraction Q consists of a branched galactomannorhamnan core, to which is attached a β-(1→3)-, β-(1→6)-glucan. In the cell wall, chains of α-(1→4)-linked glucopyranose units are linked to fraction Q to form the 6S component of fraction BB.
Ribonucleic acid–deoxyribonucleic acid hybridization in aqueous solutions and in solutions containing formamide
Authors: Gillespie S. , Gillespie D.
Published: August 2015

Hybridization in 6×SSC (SSC, 0.15m-sodium chloride–0.015m-sodium citrate) at 66°C was compared with hybridization in formamide−6×SSC (1:1, v/v) at 35°C. As expected, the RNA hybridization potential was labile in the former system and stable in the latter. DNA retention by filters was poor in the formamide system, but could be improved. Several other properties of the hybridization reaction were explored and it was concluded that the formamide system is generally superior.
Pyruvatephosphate dikinase from Bacteroides symbiosus
Authors: Reeves Richard E.
Published: August 2015

1. An improved method is given for preparation of pyruvate, phosphate dikinase from Bacteroides symbiosus. 2. The bacterial enzyme is stable, free from interfering enzyme activities, and does not require thiol compounds to maintain stability during storage or assay. 3. New direct assays of enzyme activity are based on acid evolution or consumption as measured at constant pH in a pH-stat. 4. The optimum rate of reaction in the direction of pyruvate formation occurs at about pH6.4; in the direction of phosphoenolpyruvate formation, it is at pH7.2–7.8. 5. Newly determined substrate Km values for the enzyme are: AMP, 3.5×10−6m; ATP, 1×10−4m; pyruvate, 8×10−5m; Pi, 6×10−4m. 6. K+ may substitute for NH4+ in activating the reaction catalysed by the B. symbiosus enzyme. 7. In the direction of pyruvate formation the bivalent metal ion requirement of the enzyme is fulfilled by salts of nickel, manganese, magnesium and cobalt. In the other direction only magnesium salts were effective. 8. The nucleotide specificity of the enzyme is strictly limited to the adenine nucleotides. CTP and ITP strongly inhibit the reaction in the direction of phosphoenolpyruvate formation.
The inactivation of phenylalanine hydroxylase by 2-amino-4-hydroxy-6,7-dimethyltetrahydropteridine and the aerobic oxidation of the latter. The effects of catalase, dithiothreitol and reduced nicotinamide–adenine dinucleotide
Authors: Jakubovič A. , Woolf L. I. , Chan-Henry E.
Published: August 2015

1. Phenylalanine hydroxylase is inhibited by its cofactor, 6,7-dimethyltetrahydropterin. The rate of inactivation, which is irreversible, increases with the concentration of cofactor. 2. Catalase, in sufficient amount relative to cofactor, prevents this inactivation. More tyrosine is formed in the presence of added catalase. 3. Dithiothreitol in the presence of liver extract also prevents inactivation of the enzyme by the cofactor and stimulates hydroxylation of phenylalanine, probably by protecting the cofactor from oxidation and regenerating it from a dihydropterin reaction product. Dithiothreitol restores linearity of rate at very low enzyme concentrations. 4. Dimethyltetrahydropterin is unstable when the solution is exposed to air but is stabilized by dithiothreitol the aerobic oxidation of which is greatly accelerated by dimethyltetrahydropterin. 5. NADH together with liver extract stabilizes the cofactor but not phenylalanine hydroxylase. 6. It is suggested that either hydrogen peroxide or an organic peroxide formed by oxidation in air of the cofactor is the substance attacking phenylalanine hydroxylase, dithiothreitol and cofactor.
Human senile cataractous lens protease. Isolation and some chemical characteristics
Authors: Swanson Arnold A. , Nichols John T.
Published: August 2015

A proteolytic enzyme was isolated from human senile cataractous lens by anion-exchange and gel-filtration chromatography. Sedimentation and zone-electrophoretic experiments indicated a high degree of homogeneity for the enzyme. A molecular weight of 27000 was calculated from measurements of sedimentation velocity and diffusion coefficient. Chelating agents decreased activity which could be restored by addition of certain bivalent metal ions. Di-isopropyl phosphorofluoridate and phenylmethanesulphonyl fluoride inhibit the proteolytic activities. Optimum rates of hydrolysis were observed at pH5.2.
Studies on the mechanism and regulation of C-4 demethylation in cholesterol biosynthesis. The role of adenosine 3′:5′-cyclic monophosphate
Authors: Bloxham D. P. , Wilton D. C. , Akhtar M.
Published: August 2015

1. An assay for demethylation has been developed based on the release of tritium from 4,4-dimethyl[3α-3H]cholest-7-en-3β-ol (II). 2. The maximum release of 3H from 3α-3H-labelled compound (II) in a rat liver microsomal preparation occurs in the presence of NADPH and NAD+ under aerobic conditions. 3. Incubation of 3α-3H-labelled compound (II) with NADPH under aerobic conditions leads to the formation of a 3α-3H-labelled C-4 carboxylic acid. This compound undergoes dehydrogenation on subsequent anaerobic incubation with NAD+. 4. The 3H released from the steroid was located in [4-3H]nicotinamide and the medium. Incubation with synthetic [4-3H2]NADH gave a similar result. 5. In the presence of glutamate dehydrogenase and α-oxoglutarate part of the 3H released from the steroid was transferred to glutamate. 6. A series of 3-oxo steroids were reduced equally well by [4-3H2]NADH and [4-3H2]NADPH. The reduction of 5α-cholest-7-en-3-one was shown to use the 4B H atom from the nucleotide. 7. 3′:5′-Cyclic AMP was shown to be a competitive inhibitor of the 3β-hydroxy dehydrogenase enzyme in the demethylation reaction.
Cell-free synthesis of tryptophanase from Escherichia coli. Use of ribonucleic acid isolated from induced cells and a comparison of the product from a system employing ribosomes with that from one employing ribosomes and exogenous ribonucleic acid
Authors: Parish J. H. , Bashar S. A. M. Khairul , Brown N. L. , Brown Marjorie
Published: August 2015

1. Polyribosomes and RNA were isolated from cultures in which tryptophanase (EC 4.2.1.–) was induced. The polyribosomes were incubated under conditions of protein synthesis, in the presence of a radioactive amino acid and a post-ribosomal supernatant fraction obtained from repressed cells. The RNA preparations were incubated under conditions of protein synthesis in the presence of a radioactive amino acid and a supernatant fraction containing ribosomes from repressed cells. 2. The system was characterized and the synthesis of a radioactive protein with the same chromatographic properties as tryptophanase was demonstrated. This synthesis was shown to be time-dependent and required the presence of RNA from induced cultures, ribosomes and an energy supply; it was inhibited by chloramphenicol. 3. The maximum activity for the synthesis of this protein was found to be associated with 23S rRNA isolated from sucrose gradients. 4. The N-terminal amino acid of tryptophanase was labelled in the protein synthesized in this system but not in the protein synthesized by polyribosomes (without added RNA). Conversely, the C-terminal amino acid of tryptophanase was labelled in the polyribosome system but not in the RNA-containing system. 5. Tryptic digests of protein labelled in vitro were compared with those of tryptophanase. No labelled tryptic peptides were identified other than tryptophanase tryptic peptides. An analysis of the results implied that in the polyribosome system almost the complete tryptophanase subunit chain was labelled but that in the RNA-containing system these chains were incompletely synthesized. 6. Sucrose-gradient analysis of protein synthesized in the RNA-containing system suggested that it cannot be converted into structures with the same sedimentation properties as native tryptophanase. 7. The significance of these results for the assay of tryptophanase mRNA and for an understanding of the control of the translation of this mRNA in vivo is discussed.
The effect of insulin on free amino acid pools and protein synthesis in rat skeletal muscle in vitro
Authors: Hider R. C. , Fern E. B. , London D. R.
Published: August 2015

1. The effects of insulin in vitro on tissue pools and incorporation into protein of glycine and leucine in the extensor digitorum longus muscle of the rat are reported. 2. It was found that insulin decreased the lag period before the establishment of a linear rate of incorporation of radioactive glycine into protein. 3. The hormone increased the size of the free intracellular glycine pool. No such effect was found for leucine. The accumulation of radioactive glycine in the intracellular fluid compartment was increased. The content of radioactive leucine in the intracellular compartment was decreased. 4. Insulin decreased the specific radioactivity of both glycine and leucine in the extracellular fluid. 5. The hormone also decreased protein catabolism. 6. The effect on protein synthesis was not caused by an increase in the specific radioactivity of the extracellular pool but was possibly related to increased amino acid concentrations in this pool, which could in turn have affected the aggregation of ribosomes.
The streptokinase–human plasminogen activator complex. Composition and identity of a subcomponent with activator activity
Authors: Tomar Russell H. , Taylor Fletcher B.
Published: August 2015

Reactions between purified plasminogen and streptokinase, labelled with131I and125I respectively, were investigated by polyacrylamide-gel discontinuous electrophoresis. A molecular complex consisting of both131I-labelled plasminogen and125I-labelled streptokinase migrated between plasminogen and streptokinase. This complex contained bovine plasminogen activator activity. The relative quantities of131I-labelled plasminogen and125I-labelled streptokinase in this complex were markedly affected by reaction conditions. A fragment that retained 50% or more of the parent activator activity was released from the complex after exposure to mercaptoethanol. This subcomponent had an estimated molecular weight of 70000, and contained both131I-labelled plasminogen and125I-labelled streptokinase.
Excretion in dog bile of glucose and xylose conjugates of bilirubin
Authors: Fevery J. , Van Hees G. P. , Leroy P. , Compernolle F. , Heirwegh K. P. M.
Published: August 2015

1. T.l.c. with neutral solvent systems of ethyl anthranilate azopigments derived from bile of man, dog and rat revealed pronounced species variation. The less polar components (α-group) could be separated conveniently by development with chloroform–methanol (17:3, v/v). 2. The azopigment material derived from gallbladder bile of dog contained about 10% of azobilirubin β-d-monoxyloside (azopigment α2) and 30% of azobilirubin β-d-monoglucoside (azopigment α3). The sugar moieties were identified by t.l.c. with acidic, neutral and basic solvent systems and by anion-exchange column chromatography of their boric acid complexes. Treatment of the purified azopigments with ammonia vapour led to the formation of the amide of azobilirubin, indicating that both pigments are ester glycosides. The β-d configuration was demonstrated by enzymic studies with emulsin (an adequate source of β-glucosidase activity) and with Mylase-P (an adequate source of β-glucosidase and β-xylosidase activities). 3. Hydrolysis studies with model substrates and with the α2- and α3-azopigments suggested that in Mylase-P the β-glucosidase and β-xylosidase activities reside in separate enzymes. 4. Compared with the accepted conjugation with glucuronic acid as a major route of detoxication in mammals, the detection of large amounts of xylose and glucose conjugates of bilirubin in dog bile suggests that the underlying biosynthetic pathways may be important alternative routes of detoxication.
The stability of rat liver ribonucleic acid in vivo after methylation with methyl methanesulphonate or dimethylnitrosamine
Authors: McElhone Marilyn J. , O'Connor P. J. , Craig A. W.
Published: August 2015

1. RNA was isolated from rat liver at selected times after the intraperitoneal injection of either [14C]methyl methanesulphonate (50mg/kg) or [14C]dimethylnitrosamine (2mg/kg). These doses were chosen to minimize effects due to toxicity. 2. Two methods of extraction and purification of RNA were used and an analysis of the radioactivity present was made by column chromatography of acid hydrolysates of the purified RNA. 3. The extent of methylation of guanine, the principal site of alkylation in rat liver RNA, was determined at times up to 14 days after injection. Although dimethylnitrosamine is a potent liver carcinogen and methyl methanesulphonate is not carcinogenic to rat liver, the rate of disappearance of 7-methylguanine from RNA was similar for both compounds, with a half-life of about 3.5 days. 4. An estimate of the biological half-life of rRNA was made by using [3H]orotic acid. A half-life of 5 days was obtained and this was not affected by injecting animals with unlabelled methyl methanesulphonate at the same dosage of 50mg/kg used in the studies of RNA methylation. 5. After administration of labelled orotic acid, reutilization of labelled RNA degradation products probably results in an overestimation of the biological half-life for rRNA. It is suggested that non-toxic doses of methylating agents such as methyl methanesulphonate and dimethylnitrosamine may prove to be a more effective way of accurately estimating the biological turnover of RNA species.
The action of mono- and di-functional sulphur mustards on the ribonucleic acid-containing bacteriophage μ2
Authors: Shooter K. V. , Edwards P. A. , Lawley P. D.
Published: August 2015

Bacteriophage μ2 is inactivated by both mono- and di-functional sulphur mustards at relatively low extents of alkylation. No degradation of alkylated RNA was detected. Cross-linking of RNA to protein was observed with the difunctional agent, but this reaction was only a minor contribution to the inactivation. Analyses of the reaction products in bacteriophage RNA showed that, at the mean lethal doses, more than one mono-alkylation of guanine had occurred but the sum total of other types of RNA alkylation was close to a single event. The results therefore suggest that inactivation results from the mono-alkylation of adenine or cytosine. In experiments with the difunctional agent cross-linking of RNA bases or of RNA to protein also prevented replication, the existence of these reactions accounting for the greater sensitivity of the bacteriophage to this agent.
The kinetic properties of human placental choline acetyltransferase
Authors: Morris D. , Maneckjee A. , Hebb Catherine
Published: August 2015

1. Michaelis constants for human placental choline acetyltransferase were shown to be dependent on the concentration of the second substrate present. The primary plots indicate a sequential rather than a Ping Pong mechanism and are of the same type with 300mm- and 500mm-sodium chloride. 2. Similar results have been obtained with rabbit brain choline acetyltransferase. 3. Product inhibition of the forward reaction has been studied. CoA inhibits competitively with respect to acetyl-CoA and non-competitively with respect to choline. Acetylcholine inhibits competitively with respect to choline and non-competitively with respect to acetyl-CoA. No inhibition is given by acetylcholine when the enzyme is saturated with choline. 4. It is concluded that human placental choline acetyltransferase has an ordered mechanism of the Theorell–Chance type.
Deiodination of l-thyroxine and its activity on the oxidation in vitro of reduced nicotinamide–adenine dinucleotide by peroxidase plus hydrogen peroxide
Authors: Jolin Trinidad , De Escobar Gabriella Morreale
Published: August 2015

When l-thyroxine activates the oxidation of NADH by peroxidase+H2O2, little removal of phenolic-ring iodine atoms becomes apparent until most of the NADH has been oxidized, after which it increases markedly. This extensive deiodination is accompanied by loss of the ability of thyroxine to catalyse the oxidation of NADH by peroxidase+H2O2. The slight deiodination observed before the appearance of extensive deiodination is somewhat higher when the effect of thyroxine on NADH oxidation is greater, and lower when thyroxine has exerted a slighter effect. ICN (but not I2 or thyronine) catalyses NADH oxidation, in both the presence and the absence of peroxidase+H2O2: thyroxine+peroxidase+H2O2 are thus comparable with ICN alone in their effects on NADH oxidation. The obvious conclusion from the above observation, namely that the active moiety is the halogen liberated from thyroxine (or ICN) is, however, not directly supported by some of the results obtained by measuring the degree of deiodination of thyroxine in the system. In an attempt to reconcile some apparently contradictory conclusions, it is suggested that, when thyroxine activates oxidation of NADH by peroxidase+H2O2, the diphenyl ether structure is undergoing cyclic deiodination and iodination. This would be accompanied by the maintenance in the reaction medium of an oxidized form of iodine, similar to that liberated by ICN, which would be the actual active moiety, until the NADH concentration becomes so low that the diphenyl ether structure is ruptured oxidatively. An alternative explanation is that thyroxine is oxidized to a form that either oxidizes NADH or loses iodine in competing reactions.
Formation of bilirubin glucoside
Authors: Wong K P
Published: August 2015

1. Rat liver microsomal preparation can effect the transglucosylation from UDP-glucose to bilirubin in the presence of Mg2+. 2. Other nucleotides, namely CDP-glucose, ADP-glucose and GDP-glucose, were not active as glucosyl donors. 3. Only trace amounts of galactose, galacturonic acid and N-acetylglucosamine were conjugated to bilirubin when their respective UDP derivatives were used in the reaction mixture. 4. The azobilirubin glucosides produced by coupling with p-diazobenzenesulphonic acid and diazotized ethyl anthranilic acid were separable from the corresponding azobilirubin glucuronides by t.l.c. 5. The glucoside was, however, hydrolysed by both β-glucosidase and various preparations of β-glucuronidase; azobilirubin and glucose were liberated in the process. 6. Kinetic studies showed that the effects of pH and Mg2+ on the two conjugating systems were similar. 7. The specific activities of hepatic bilirubin UDP-glucosyltransferase, expressed as μg of bilirubin ‘equivalents’ conjugated/h per mg of protein, are respectively 1.7 and 2.4 for male and female rats. 8. The Km values for bilirubin and UDP-glucose are 5.7×10-5m and 1.6×10-3m respectively. 9. The glucoside and glucuronide conjugations of bilirubin are discussed in relation to the availability of the conjugating agents and aglycone in the liver.
3β-Hydroxy steroid dehydrogenase activity in the mitochondria of rat adrenal homogenates
Authors: Basch R S , Finegold M J
Published: August 2015

The activity of 3β-hydroxy steroid dehydrogenase (EC 1.1.1.51) in the mitochondrial fraction of rat adrenal homogenates was approx. 31% of the total activity recovered after differential centrifugation and washing of the particulate fractions. Some 45% of the total activity was found in the microsomal fraction. The activity was assayed by a radioisotopic method devised in this laboratory for the purpose of studying small quantities of tissue and cell fractions. Satisfactory separation of the two fractions was demonstrated by electron microscopy of the pellets and by comparative recoveries of RNA, steroid 21-hydroxylase and cytochrome c oxidase in the various compartments. Analyses of the kinetics of the enzyme activity in the two fractions revealed no significant differences in apparent Km for pregnenolone, dehydroepiandrosterone or NAD+, but demonstrated a distinct difference in the Km for NADP+. pH optima and susceptibility to cyanoketone inhibition were similar in both fractions.
The effect of diffusible acids on potassium ion uptake by yeast
Authors: Ryan H. , Ryan J. P. , O'Connor W. H.
Published: August 2015

1. When yeast oxidizes ethanol at different pH values the uptake of K+ corresponds closely to the amount of acetate accumulated at each pH value. 2. The addition of semicarbazide to the suspension buffered at pH4.75 inhibited both the K+ uptake and the acetate accumulation by about 50%. 3. The addition of either acetate or propionate to the suspensions markedly increased the K+ uptake. 4. The addition of acetate to the suspensions lowered the intracellular pH of the yeast from a resting value of pH5.80 to 5.56. 5. The ratio of the initial rate of K+ uptake to O2 consumption was 0.77. This ratio was increased to 1.77 in the presence of 10mmol of propionate/l.
The deoxyribonucleic acid modification enzyme of bacteriophage P1. Purification and properties
Authors: Brockes Jeremy P. , Brown Paul R. , Murray K.
Published: August 2015

The bacteriophage P1 modification enzyme, assayed by the specific methylation of unmodified bacteriophage 82 DNA, has been purified 500-fold from a bacteriophage P1 lysogen of Escherichia coli. The enzyme catalyses the incorporation of approximately 20–24 methyl groups per bacteriophage 82 DNA molecule. The sole product of methylation is 6-methylaminopurine. Methylation of unmodified bacteriophage DNA confers protection against a challenge by purified bacteriophage P1 restriction enzyme. The pH optimum is 6.0–6.25: the apparent Km for S-adenosyl-l-methionine is 5×10−6m.
Lipid intermediates in the biosynthesis of the wall teichoic acid in Staphylococcus lactis I3
Authors: Hussey Helen , Baddiley J.
Published: August 2015

1. Particulate enzyme systems have been prepared from Staphylococcus lactis I3 which effect the synthesis of wall teichoic acid (a polymer containing a repeating unit in which d-glycerol 1-phosphate is attached to the 4-position on N-acetylglucosamine 1-phosphate) from the nucleotide precursors CDP-glycerol and UDP-N-acetylglucosamine. By using nucleotides labelled with 32P and 14C it has been shown that the synthesis proceeds via lipid intermediates. 2. Two intermediates have been found. In one of these N-acetylglucosamine 1-phosphate is present, whereas in the other the repeating unit of the teichoic acid occurs. 3. The simultaneous formation of the teichoic acid, a poly-(N-acetylglucosamine 1-phosphate) and an unidentified lipid, together with the poor ability of most particulate systems to synthesize polymer and the instability of the lipid intermediates themselves, have interfered with pulse-labelling experiments. Nevertheless, the biosynthetic sequence has been elucidated. It is concluded that the intermediates are derivatives of undecaprenol phosphate.
The peroxidase-catalysed oxidation of a chalcone and its possible physiological significance
Authors: Rathmell W. G. , Bendall D. S.
Published: August 2015

1. Crystalline horseradish peroxidase catalysed the oxidation of 2′,4,4′-trihydroxychalcone (isoliquiritigenin) in the presence of trace amounts of hydrogen peroxide under aerobic conditions. One atom of oxygen was consumed for each molecule of substrate. 2. The reaction course comprised a lag phase and a linear phase. The optimum pH for the linear phase of the reaction was about 7.5. The length of the lag phase decreased with increasing pH. It is suggested that the chalcone anion is the actual substrate for the reaction. 3. No evidence for the production of reducing free radicals or perhydroxyl radicals during the reaction could be found. 4. 4′,7-Dihydroxyflavonol and 4′,6-dihydroxyaurone were isolated from the reaction mixture. The immediate products of the reaction may have included 3,4′,7-trihydroxyflavanone and 4′,6-dihydroxy-2-(α-hydroxybenzyl)coumaran-one, which can be readily converted non-enzymically into the flavonol and aurone respectively. 5. A similar reaction was catalysed by cell-free extracts of hypocotyls of Phaseolus vulgaris. 6. The physiological significance of the reaction is discussed in terms of a possible free-radical mechanism. An analogy may exist between flavonoid biosynthesis and lignin formation.
The subcellular distribution of rat liver l-alanine–glyoxylate aminotransferase in relation to a pathway for glucose formation involving glyoxylate
Authors: Rowsell E. V. , Snell K. , Carnie J. A. , Rowsell Kathleen V.
Published: August 2015

1. The distribution of l-alanine–glyoxylate aminotransferase activity between subcellular fractions prepared from rat liver homogenates was investigated. The greater part of the homogenate activity (about 80%) was recovered in the `total-particles' fraction sedimented by high-speed centrifugation and the remainder in the cytosol fraction. 2. Subfractionation of the particles by differential sedimentation and on sucrose density gradients revealed a specific association between the aminotransferase and the mitochondrial enzymes glutamate dehydrogenase and rhodanese. 3. The aminotransferase activities in the cytosol and the mitochondria are due to isoenzymes. The solubilized mitochondrial enzyme has a pH optimum of 8.6, an apparent Km of 0.24mm with respect to glyoxylate and is inhibited by glyoxylate at concentrations above 5mm. The cytosol aminotransferase shows no distinct pH optimum (over the range 7.0–9.0) and has an apparent Km of 1.11mm with respect to glyoxylate; there is no evidence of inhibition by glyoxylate. 4. The mitochondrial location of the bulk of the rat liver l-alanine–glyoxylate aminotransferase activity is discussed in relation to a pathway for gluconeogenesis involving glyoxylate.
Phagocytosis and leucocyte enzymes in protein–calorie malnutrition
Authors: Selvaraj Ratnam J. , Bhat K. Seetharam
Published: August 2015

1. Enzymes pertinent to bactericidal activities of leucocytes were assayed in children suffering from protein–calorie malnutrition. 2. Leucocytes obtained from malnourished and control children contained similar activities for glucose 6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase. Granule-bound NADPH oxidase activity was low in leucocytes isolated from malnourished patients and failed to show the phagocytic stimulation which is normally seen in control leucocytes. Further, leucocytes obtained from malnourished patients did not release the acid phosphatase from lysosomes during phagocytosis, unlike those from controls. 3. Treatment of the malnourishment with a diet high in calories and protein resulted in significant increase in the activities of glucose 6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase and NADPH oxidase and in releasing the acid phosphatase from the lysosomes into the supernatant fraction during phagocytosis. 4. The significance of these enzyme changes are discussed in relation to the increased susceptibility of these patients to infection.
Further studies on the lipids of corynebacteria. The mannolipids of Corynebacterium aquaticum
Authors: Khuller G. K. , Brennan P. J.
Published: August 2015

1. The major free lipids of Corynebacterium aquaticum were characterized as dimannosyl diglyceride, monomannophosphoinositide and phosphatidylethanolamine. Bisphosphatidylglycerol and phosphatidylglycerol were also tentatively identified. 2. We regard this as the only well-documented case of an organism containing monomannophosphoinositide to the exclusion of dimannophosphoinositides and the higher homologues. 3. The co-existence of the two mannolipids in one organism is a distinctive feature. So also is the presence of phosphatidylethanolamine in a corynebacterium. 4. The monomannophosphoinositide apparently does not utilize phosphatidylinositol as a precursor, unlike the monomannophosphoinositide of Propionibacterium shermanii. CDP-diglyceride may be necessary for its synthesis.
The enzymic properties of a modified ox heart myosin adenosine triphosphatase on covalent binding to an insoluble cellulose matrix
Authors: Liu-Osheroff Phyllis , Guillory Richard John
Published: August 2015

The preparation of ox heart myosin and its partial digestion with cellulose-bound papain is described. A procedure is outlined by which heavy meromyosin subfragment 1 can be covalently bound to a cellulose ion-exchange matrix. Attachment of heavy meromyosin subfragment 1 to the insoluble matrix results in a change in the ion specificity towards ATP hydrolysis. Unlike the soluble enzyme the bound form is activated by both Ca2+ and Mg2+. Maximal activation by Ca2+ occurred at a lower concentration for the bound enzyme. Mg2+ activates at a concentration which causes near-maximal inhibition of the Ca2+-activated adenosine triphosphatase (ATPase) of the non-bound enzyme. The Mg2+-activated ATPase of the bound enzyme was in turn inhibited by the presence of Ca2+. The activation by Mg2+ resembles the characteristic enzymic action of the actin-subfragment 1 complex.
Some studies on the biosynthesis of ubiquinone, isoprenoid alcohols, squalene and sterols by marine invertebrates
Authors: Walton M. J. , Pennock J. F.
Published: August 2015

The ability of fourteen marine invertebrates to utilize [14C]mevalonate for the biosynthesis of isoprenoid compounds was investigated. Several of the animals, in particular crustaceans, bivalve molluscs, a coelenterate and a sponge, were unable to synthesize squalene and sterols, whereas gastropod molluscs, echinoderms, an annelid and a sponge could. Regardless of sterol-synthesizing ability the animals (with the exception of a sponge) always made dolichol and ubiquinone, and thus a specific block in squalene and sterol synthesis was indicated in some animals. Radioactivity accumulated in relatively large amounts in farnesol and geranylgeraniol in those animals incapable of making sterols.
The molecular weights of two forms of carbamoyl phosphate synthase from rat liver
Authors: Virden R.
Published: August 2015

1. N-Acetylglutamate-dependent carbamoyl phosphate synthase from rat liver was centrifuged in sucrose density gradients. The concentration-dependence of s was consistent with a chemical equilibrium existing between the 11S and 7.5S forms of the enzyme. 2. Under conditions favouring the 11S form, the properties of the enzyme in ultra-short-column equilibrium experiments suggest a molecular weight of 316000±42000 for the 11S form. 3. Under conditions favouring the 7.5S form, high-speed equilibrium-sedimentation measurements gave a value of 160000±10000 as the molecular weight of the 7.5S form of the enzyme.
The quantitative determination of phenylalanine hydroxylase in rat tissues. Its developmental formation in liver
Authors: McGee Margaret M. , Greengard Olga , Knox W. Eugene
Published: August 2015

A sensitive method was developed for determining the phenylalanine hydroxylase activity of crude tissue preparations in the presence of optimum concentrations of the 6,7-dimethyl-5,6,7,8-tetrahydropterin cofactor (with ascorbate or dithiothreitol to maintain its reduced state) and substrate. Tissue distribution studies showed that, in addition to the liver, the kidney also contains significant phenylalanine hydroxylase activity, one-sixth (in rats) or half (in mice) as much per g as does the liver. The liver and the kidney enzyme have similar kinetic properties; both were located in the soluble phase and were inhibited by the nucleo-mitochondrial fraction. Phenylalanine hydroxylase, like most rat liver enzymes concerned with amino acid catabolism, develops late. On the 20th day of gestation, the liver (and the kidney) is devoid of phenylalanine hydroxylase and at birth contains 20% of the adult activity. During the second postnatal week of development, when the phenylalanine hydroxylase activity was about 40% of the adult value, an injection of cortisol doubled this value. Cortisol had no significant effect on phenylalanine hydroxylase in adult liver or on phenylalanine hydroxylase in kidney at any age.
Lipid composition of membranes of aminestorage organelles
Authors: Da Prada M. , Pletscher A. , Tranzer J. P.
Published: August 2015

1. The lipid composition of the membranes from isolated 5-hydroxytryptamine-storage organelles of blood platelets of rabbits and of those from chromaffin granules of bovine adrenal medulla was compared. 2. In contrast with the membranes of the chromaffin granules, those of the 5-hydroxytryptamine organelles did not contain lysophosphatidylcholine (lysolecithin). 3. Both the cholesterol/phospholipid ratio and the relative proportions of phosphatidylethanolamine (kephalin), phosphatidylinositol and phosphatidylserine were about the same in both membranes, whereas phosphatidylcholine (lecithin) and sphingomyelin showed somewhat higher values in the membranes of the 5-hydroxytryptamine organelles. 4. In conclusion, the release of 5-hydroxytryptamine from blood platelets is probably not correlated with the presence of lysophosphatidylcholine in the membranes of the storage organelles and may thus differ from the mechanism of catecholamine release in adrenal medulla.
Oestradiol inhibits collagen breakdown in the involuting rat uterus
Authors: Ryan Janet N. , Woessner J. Frederick
Published: August 2015

1. The earlier observation (Woessner, 1969) of oestradiol inhibition of collagen breakdown is confirmed and extended. Administration of 100μg of oestradiol-17β/day to parturient rats strongly inhibits the loss of collagen from the involuting uterus. Three experiments show that this effect is due to an inhibition of collagen degradation rather than to a stimulation of collagen synthesis. 2. Uterine collagen was labelled with hydroxy[14C]-proline by the administration of [14C]proline near the end of pregnancy. By 3 days post partum, control uteri lost 83% of their collagen and 90% of their hydroxy[14C]proline. Uteri from oestradiol-treated rats lost only 50% of both total and labelled hydroxyproline, with no decrease in the specific radioactivity of the hydroxyproline. 3. Incorporation of [14C]proline into uterine collagen hydroxyproline in vivo was not affected by oestradiol treatment. 4. Urinary excretion of hydroxyproline was increased in post-partum control rats and decreased in oestradiol-treated rats. 5. An enzyme capable of cleaving 4-phenylazobenzyloxycarbonyl-l-prolyl-l-leucylglycyl- l-prolyl-d-arginine (a substrate for clostridial collagenase) increased in activity in the post-partum uterus and was unaffected by oestradiol treatment. 6. Uterine homogenates digested uterine collagen extensively at pH3.2. This digestion was unaffected by the oestradiol treatment. 7. Lysosomal fractions prepared by density-gradient centrifugation of uterine homogenates contained coincident peaks of cathepsin D activity and peptide-bound hydroxyproline. The cathepsin D and hydroxyproline contents of this peak were unaffected by oestradiol treatment.
Effects of proteins and polynucleotides on the activity of various hydrolases
Authors: Palmieri M. J. , Koldovský O.
Published: August 2015

The effect of various macromolecules on the activity of several hydrolases was studied. Dilution of partially purified acid β-galactosidase from ileal mucosa of suckling rats resulted in a decrease of specific activity. The relationship between specific activity and dilution of the enzyme suggests a dissociation of the enzyme. This could be prevented by addition of several proteins tested. However, addition of DNA to the assay mixture for acid β-galactosidase caused an inhibition. This inhibition could be prevented by addition of proteins. Other polynucleotides and tRNA also exert an inhibitory effect that is prevented by albumin, but nucleotides have no effect. This inhibition occurs maximally at a low pH (3.0–4.0); no inhibition is observed at pH5.5. A similar pH-dependent inhibition by DNA was also found with various other acid hydrolases.
The interaction of catecholamines and adrenal corticosteroids in the induction of phosphopyruvate carboxylase in rat liver and adipose tissue
Authors: Reshef Lea , Hanson R. W.
Published: August 2015

Catecholamines induced an increase in the activity of rat adipose tissue and liver phosphopyruvate carboxylases that was maintained for 48h. The response of adipose tissue phosphopyruvate carboxylase was blocked by actinomycin D, corticosteroids and propranolol, whereas corticosteroids and propranolol did not affect the liver enzyme. Cortisol phosphate, like actinomycin D, interfered only with the initiation of the increase in enzyme activity caused by noradrenaline, but not with the process of enzyme accumulation. In contrast, cycloheximide was effective in blocking enzyme induction throughout the course of the catecholamine effect. Adrenocorticotrophic hormone caused a short-term induction of adipose tissue phosphopyruvate carboxylase, which could be blocked by propranolol. Hepatic phosphopyruvate carboxylase, but not the adipose tissue enzyme, was induced by dibutyryladenosine 3′:5′-cyclic monophosphate and by glucagon. Both nicotinic acid and nicotinamide decreased the normal induction of adipose tissue phosphopyruvate carboxylase caused by starvation, but only nicotinamide increased the activity of the liver enzyme.