bluuebunny/crossref_metadata_2025_split huggingface dataset:
Biosynthesis of N‐Acetylneuraminic Acid in Morris Hepatomas
Authors: Harms Erik , KREISEL Wolfgang , Reutter Werner , MORRIS Harold P. 
Published: March 2005

The biosynthesis of N‐acetylneuraminic acid was studied in liver, neonatal liver, regenerating liver and Morris hepatomas 7777, 5123 TC, 5123 C, 5123 D, 7800, 7795, 9121 and 9618 A. The following three parameters were investigated: the biosynthesis of N‐acetylneuraminic acid from d‐[1‐14C]glucosamine and d‐N‐acetyl‐[U‐14C]mannosamine; the content of free and protein bound N‐acetylneuraminic acid and of CMP‐N‐acetylneuraminic acid; the activity of the key enzyme (UDP‐N‐acetylglucosamine‐2′‐epimerase).

The biosynthesis of N‐acetylneuraminic acid was investigated in vivo by labelling experiments using [1‐14C]glucosamine and N‐acetyl‐[U‐14C]mannosamine as precursors revealing the same metabolic route in liver and hepatoma. The biosynthesis of N‐acetylneuraminic acid deriving from [1‐14C]glucosamine in vivo is diminished to about 15% in hepatoma compared to liver, whereas the synthesis of UDP‐N‐acetylglucosamine is not altered. The uptake of N‐acetyl‐[U‐14C]mannosamine and its conversion to N‐acetylneuraminic acid in liver and hepatoma is demonstrated. The synthesis of free N‐acetylneuraminic acid deriving from N‐acetyl‐[U‐14C]‐mannosamine is not changed in hepatoma. The incorporation of this precursor into protein‐bound N‐acetylneuraminic acid of hepatoma is four‐times higher.
The content of free and protein bound N‐acetylneuraminic acid and CMP‐N‐acetylneuraminic acid have been measured in livers and hepatomas by isotope dilution technique. About 90% of total N acid content in both livers and hepatomas is protein‐bound. In Morris hepatomas the content of protein‐bound N‐acetylneuraminic acid is elevated (between 5.93 and 8.38 nmol/mg protein) compared to liver (4.50 ± 0.52 nmol/mg protein). However the content of free. N‐acetylneuraminic acid including CMP‐N‐acetylneuraminic acid is diminished in Morris hepatomas (between 108 and 164 nmol/g hepatoma in comparison to 196 ± 28 nmol/g liver), whereas the content of CMP‐N‐acetylneuraminic acid alone is not greatly changed (31.4 ± 8.5 nmol/g Morris hepatoma 9121 in comparison to 40.8 ± 10.1 nmol/g liver).
The activity of UDP‐N‐acetylglucosamine‐2′‐epimerase of all Morris hepatomas investigated was diminished to between 6% and 13% compared to that of host liver. In other fast growing liver tissue (neonatal and regenerating liver) this enzyme activity was not diminished but slightly increased showing that the decreased enzyme activity is a characteristic feature of Morris hepatomas.
Mechanism of the Exchanges Catalysed by the Oxoglutarate Translocator of Rat‐Heart Mitochondria
Authors: Sluse Francis E. , LIÉBECQ Claude , Goffart Georges 
Published: March 2005

The kinetics of the exchange reactions between internal l‐malate and external 2–oxoglutarate or malonate have been measured at 4°C in the presence of external l‐malate (external‐product inhibitor), using preparations of rat‐heart mitochondria under conditions where the oxoglutarate translocator is operating exclusively. Measurements of initial rates were made at three concentrations of the internal substrate (l‐malate), three concentrations of the external substrate (2‐oxoglutarate or malonate) and three concentrations of the external reaction product (l‐malate).The Michaelis constant for the internal malate is unaffected by the presence of the external product whereas the apparent‐maximum rate for each concentration of external substrate decreases when the concentration of external product increases. On the other hand the Michaelis constant for the external substrate, oxoglutarate or malonate, increases when the concentration of external malate increases.The external product inhibits the exchange in a competitive way when external substrate is varied and in a non‐competitive way when internal substrate is varied.The inhibition constant determined for the external malate is equal to the previously determined dissociation constant of the corresponding external‐malate · translocator complex.These results are in perfect agreement with the previously proposed mechanism (rapid‐equilibrium random bi‐bi) and a mixed dead‐end and product inhibition.
Influence of Bile Salts on Cholesterol Synthesis in the Isolated Perfused Rat Liver
Authors: Decker Karl F. A. , BARTH Christian A. , Hackenschmidt H. Jürgen , ULLMANN Harald L. , Liersch Michael E. A. 
Published: March 2005

The effect of taurocholate and cholate on hepatic cholesterol synthesis has been studied with the isolated perfused rat liver. By the infusion of bile salts at a constant rate to the portal vein, conditions in the enterohepatic circulation of taurocholate and cholatein vivowere simulated.A 1‐h infusion of sodium taurocholate, at a rate of 10mg × (100g body wt)−1× h−1, did not inhibit cholesterol synthesis, as measured by3H2O incorporation.The same results were obtained, when livers with enhanced cholesterol synthesis were studied or when bile salt infusions were extended to 2 h before starting incorporation measurements. Stimulation was achieved by feeding a 2% cholestyramine diet during 7 days to the donor animals.A 3‐h infusion of taurocholate, at the above rate, exerted no toxic effect on liver metabolism, as judged from hepatic enzyme leakage to the perfusion medium, from the rate of ketone body production and the 3‐hydroxybutyrate/acetoacetate ratio.Cholic acid in amounts equivalent to those of taurocholate did not influence cholesterol synthesis under these conditions.It is concluded, that inhibition of cholesterol synthesis by bile saltsin vivois not mediated by a short‐acting (e.g.allosteric) mechanism at the enzymatic level, but rather by a change of enzyme content.
Properties of the Elongation Factors from Escherichia coli
Authors: Chinali Gianni , PARMEGGIANI Andrea 
Published: March 2005

The interaction of elongation factor G (EF‐G) with ribosomes has been studied during or in the absence of elongation of the polypeptide chain in a system from Escherichia coli in vitro.Mutual saturation curves of ribosomes and EF‐G suggest a rapid exchange of the factor from ribosome to ribosome. In the GTPase reaction, V/2 was reached at [ribosomes]/[EF‐G] ∼ 14 and at [EF‐G]/[ribosomes] ∼ 33. In the poly(U)‐directed poly(phenylalanine) synthesis, V/2 was obtained at [ribosomes]/[EF‐G] ∼ 12 and at [EF‐G]/[ribosomes] ∼ 6.Kinetics of the exchange of EF‐G between two pure acetyl‐aminoacyl‐tRNA ·mRNA · ribosome complexes containing synthetic homo‐ or heteropolymeric mRNAs were investigated by following their respective EF‐G‐dependent incorporation of differently labelled amino acids into the corresponding polypeptides in experimental conditions in which EF‐G was either added free or already prebound to one of the complexes prior to addition of the second one.In all experiments the rates of incorporation showed that the exchange of EF‐G between the two complexes was immediate and complete. These observations indicate that EF‐G behaves as a virtually free component which interacts catalytically with ribosomes during each round of elongation of the polypeptide chain.
Reduced Rate of β‐Globin mRNA Translation in β‐Thalassemia
Authors: Conconi Francesco , DEL SENNO Laura , Borgatti Luigi 
Published: March 2005

The synthesis of α and β‐globin have been determined in normal and thalassemic erythroid cells, incubated at temperatures decreasing from 37 to 10°C.Synthesis of β‐globin by thalassemic reticulocytes decreased with temperature clearly less than α and β‐globin synthesis by normal erythroid cells, and also less than α‐globin synthesis by thalassemic reticulocytes. Consequently the excess of α‐globin synthesis, typical of β‐thalassemia, disappeared when cells where incubated at temperatures of approximately 10°C.The normalization of β‐globin synthesis, obtained in our experimental conditions, clearly suggests that the defect of β‐globin synthesis, characteristic of β‐thalassemia, does not result from a reduction in amount of β‐globin messenger, but rather from a reduction of the rate of β‐globin mBNA translation.
Immunochemical Properties of Procollagen from Dermatosparactic Calves
Authors: Timpl Rupert , WICK Georg , Furthmayr Heinz , LAPIÈRE Charles M. , Kühn Klaus 
Published: March 2005

After immunization of rabbits with dermatosparactic calf‐skin collagen a predominantly specific response to this kind of collagen and only a weak cross‐reaction with soluble skin collagen from normal calves was observed. Antibodies exclusively reacting with dermatosparactic collagen were obtained upon immunoadsorption. They displayed strong reactions with the constituent pα1‐ and pα2‐chain although both chains might not be equivalent in their antigenic structure. The serologic activity was entirely recovered on a characteristic additional peptide (molecular weight 20000) obtained from pα1‐chain after collagenase digestion. Reduction and alkylation of this peptide, but not alkylation alone, completely destroyed the antigenic activity which, on the other hand, was quite stable towards the action of various proteases. Antigenic determinants specific for the collagen α1‐chain of normal calf skin could also be demonstrated in the pα1‐chain except the one located in the N‐terminal pentapeptide sequence.In indirect immunofluorescence tests antibodies specific for dermatosparatic collagen were shown to react with collagen fibers of skin and the interstitial connective tissue of kidney and muscle of normal calves. No staining of glomerular basement membranes was observed. Strong cross‐reaction with human but not with rat tissues was obtained. These results suggest that the extended collagen molecules found in dermatosparactic calves are normal tissue constituents which in all probability are identical to the biosynthetic precursor, procollagen.
Electron‐Transport Chains of Escherichia coli
Authors: Haddock Bruce A. , Schairer Hans U. 
Published: March 2005

The properties of a 5‐aminolaevulinic acid synthetase‐deficient mutant of Escherichia coli K12Ymel grown aerobically or anaerobically in the presence or absence of 5‐aminolaevulinic acid are described. In the absence of 5‐aminolaevulinic acid no detectable cytochromes are produced, respiration rates are lowered, and the concentration of menaquinone in the cell is increased. The absence of 5‐aminolaevulinic acid from the growth medium has no apparent effect on the Mg2+ dependent ATPase activity or on the concentrations of ubiquinone and flavoprotein(s) in the cell.
Two methods for the reconstitution of respiration by cells grown aerobically or anaerobically in the absence of 5‐aminolaevulinic acid are described. With intact cells, reconstitution is achieved in 3 h at 37 °C and is dependent on the presence of 5‐aminolaevulinic acid and a fermentable carbon source. With cell‐free extracts, respiration can be reconstituted by the addition of haematin in 15–20 min at 37 °C provided that ATP is also present.
It is concluded that the reconstitution of respiration by both methods is achieved in the absence of protein synthesis, indicating that the apoproteins of cytochromes, involved in electron transport with oxygen as terminal electron acceptor, are synthesized and incorporated into the membrane in the absence of haem synthesis.
From an examination of the cytochromes produced during the reconstitution experiments, it is suggested that two membrane‐bound electron transport systems using oxygen as terminal electron acceptor can co‐exist in E. coli. The first involving cytochrome b556 with cytochrome o as terminal oxidase is the system normally present under aerobic growth conditions, and the second is induced under conditions when the concentrations of NADH and ADP in the cell are likely to be high and involves cytochrome b558 with cytochrome a2 as the terminal oxidase.
Sex Dependency of Steroid Retention in Rat‐Liver Slices Incubated with Testosterone
Authors: Ghraf Rüdiger , Lax E. Rodney , Hoff H.‐Georg , Schriefers Herbert 
Published: March 2005

The hepatic retention of steroids which occurs during the incubation of [4‐14C]testosterone with rat liver slices is demonstrated to be a sex‐specific phenomenon. After an incubation time of 30 min, slices from female animals can be shown to have retained significantly more [4‐14C] steroid than slices from male animals. The tissue/medium distribution ratio is 13 for the males and 30 for the females.Δ4‐5α‐Hydrogenase activity of the liver tissue can be regarded as the driving force for the retention of steroids derived from testosterone turnover. Namely, the steroid retention is positively correlated with the testosterone hydrogenation rate. The Δ4‐hydrogenating activity of the female exceeds that of the male by a factor of 1.73; this is the same factor by which the hepatic retention of the female exceeds that of the male.In contrast, there is an inverse correlation between the production rate of hydroxylated C19O2‐metabolites and the extent of retention. When compared to the female animal, slices from the male possess a higher hydroxylating activity accompanied by a much lower retention.The majority of metabolites isolated from tissue have a retention which is proportional to their production rate; however, the proportionality factor varies from metabolite to metabolite. Thus, for each individual metabolite, the slice is able to maintain a specific concentration gradient against the medium. For these metabolites the retention is sex‐specific only in so far as their production rate is sex‐specific.The retention of testosterone and androstenedione is independent of their production rate. They are found in the incubation medium of both sexes, but only in the tissue of males.
Multiple Molecular Forms of Pyruvate Kinase from Mucor rouxii
Authors: Friedenthal Myriam , Roselino Eduardo , Passeron Susana 
Published: March 2005

Pyruvate kinase from Mucor rouxii constitutes a family of three isoenzymes which can be separated in DEAE‐cellulose columns. These have been named type I, II and III. They also have different electrophoretic mobility in polyacrylamide gels. Pyruvate kinases type I, II and III were partially purified by ion‐exchange chromatography. Antisera were prepared against types I and III. The antigenic behaviour of the three isoenzymes supported the hypothesis on the hybrid nature of pyruvate kinase type II and confirmed the identity of pyruvate kinase from yeast‐like cells and type I from mycelium.The filamentous form contains the three types, none of them being constitutive and the proportion of each depends on the concentration and on the nature of the carbohydrate in the growth medium. Under gluconeogenic conditions of growth type III predominantes while in any condition in which aerobic glycosis is favoured type I is the dominant form.Aerobic or anaerobic yeast‐like cells which display a fermentative mode of hexose utilization contain exclusively the type I isoenzyme.Addition of glucose or xylose to a 17‐h‐old cultures devoid of carbohydrates leads to the reappearance of pyruvate kinase I and II activities. Experiments with cycloheximide and actinomycin D indicate that this process requires protein synthesis.Results of preincubation of mixtures of extracts under several conditions indicate that the system of three pyruvate kinases does not represent proteolytic degradation of a single enzyme or interconvertible forms of the same enzyme.
Isolation and Properties of Abrin: a Toxic Protein Inhibiting Protein Synthesis
Authors: Olsnes Sjur , Pihl Alexander 
Published: March 2005

A new procedure is reported for the purification of the toxic protein abrin and for the isolation of its two constituent peptide chains.Abrin was extracted from Semen jequiriti and purified by chromatography on a DEAE‐cellulose column and on a Sepharose‐4B column. The purified toxin behaved as a single protein on polyacrylamide‐gel electrophoresis. The LD50 dose in mice was about 70 ng indicating that the preparation was about 6‐times as toxic as the most active abrin previously described.After reduction of the toxin with 2‐mercaptoethanol in the presence of galactose the two constituent peptide chains of abrin were isolated by chromatography on a DEAE‐cellulose column.The smaller peptide, denoted the A‐chain, inhibited strongly protein synthesis in a cell‐free system from rabbit reticulocytes, whereas the larger peptide, the B‐chain, lacked this ability.Human erythrocytes pretreated with intact abrin or with B‐chain were agglutinated by an antiserum against abrin, whereas erythrocytes pretreated with A‐chain were not agglutinated under these conditions.The results indicate that the toxic action of abrin is associated with the A‐chain and that the B‐chain functions as a “carrier” moiety necessary for binding of the toxin to the cell surface.
A Study of the Template Properties of Chromatin for DNA Polymerase I and of the Effects of Ionising Radiation
Authors: Itzhaki Ruth F. , Saffhill R. 
Published: March 2005

The template activity of rat thymus chromatin for Escherichia coli DNA polymerase I has been found to amount to only 6% of that of pure DNA. Assays have been carried out on chromatin in 0.05 M NaCl, on a chromatin · polylysine complex, on the soluble and insoluble fractions of chromatin in the assay system and on DNA prepared from chromatin. The results suggest that the low activity is due only to a small extent to the insolubility of the chromatin in the assay system. Instead, it is proposed that initiation is restricted to certain “accessible’ DNA zones in chromatin.After γ‐irradiation of the chromatin, the template activity increases with increasing dose up to a certain level and then decreases. It is therefore proposed that both active and inactive binding sites for DNA polymerase are formed on irradiation. Assays on a chromatin · polylysine complex indicate that the new initiation sites occur in the “accessible” zones.
Détermination d'un indice d'affinité des stéroïdes pour les protéines sériques, fondée sur une méthode de mesure de la liaison à l'équilibre
Authors: Goertz Guy R. , Guérin Marie A. , Crépy Odette C. , Longchampt Jacques E. , Jayle Max F. 
Published: March 2005

A method has been developped to compare the competition between various unlabelled C19 and C18 steroids and [3H]testosterone for binding, during pregnancy, to human serum proteins. This method is based on equilibrium dialysis, according to the technic of Pearlman and Crépy, using Sephadex G‐25 in a batchwise fashion. Bound and unbound steroid may be measured without disturbing equilibrium and, in this way, the different rates of dissociation of complexes do not interfere with the measurement.The partition coefficients K' of unlabelled and unbound steroid between the Sephadex‐external and internal volumes have also been determined.Ten steroids have been studied by this method. Displacement of radioactive testosterone bound to serum proteins by increasing amounts of unlabelled steroids is measured. In order to get a linear relationship between the two parameters, graphs are plotted according to the method of Rodbard and Rees Midgley. Logit is plotted against the logarithm of unbound steroid in the external volume (Sue), where B = 100 × Sb*/Sb0*, Sb* being the still‐bound [3H]testosterone after addition of unlabelled steroid and Sb0* the total bound [3H] testosterone without addition of unlabelled steroid.In order to compare the relative affinities of the steroids studied, a “displacement index” is determined. It is expressed as the ratio of unlabelled testosterone (ng) to the test substance (ng) required to decrease the bound [3H]testosterone to 50% of its initial value. The results thus obtained are similar (with the exception of 5‐androstenediol) to those described by Murphy, especially for 19 nor‐5‐androstenediol, which is the only 19 nor‐steroid showing a very high affinity for pregnancy human serum proteins. The displacement index, using testosterone as reference steroid, is higher for dihydrotestosterone, 5α‐androstanediol, 4‐androstenediol and 5‐androstenediol and lower for dehydroepiandrosterone, 19 nor‐testosterone, estradiol and 16‐hydroxy‐C19 compounds.
Phenol Hydroxylase from Yeast
Authors: Neujahr Halina Y. , Gaal Andràs 
Published: March 2005

An enzyme, able to carry out an NADPH‐dependent hydroxylation of monocyclic phenols, was purified 20–30‐fold from Trichosporon cutaneum grown on phenol or resorcinol as a major carbon source. The purified enzyme was homogeneous upon analytical disc electrophoresis.The enzyme is a bright‐yellow protein with an absorption spectrum typical of flavoproteins. Its molecular weight is 148000, and its estimated FAD content is approximately 1 mole per mole enzyme. The purified enzyme has essentially the same broad substrate specificity as crude preparations. In addition to phenol it also hydroxylates the three isomeric diphenols and a number of phenol derivatives to their corresponding o‐diols. K
m values for the three substrates of the holoenzyme are all of the order 10 μM : Km (phenol) = 18 μM, Km(NADPH) = 71 μM. The absorption spectrum of the holoenzyme is modified in the presence of phenol.FAD could be resolved from the purified enzyme by preparative disc electrophoresis. Reassembly of the holoenzyme required SH‐groups.The enzyme activity is unaffected by chelators of iron and copper but it is inhibited by heavy metals. The inhibition by p‐chloromercuribenzoate is readily reversed by dithiothreitol.Among oxidizing agents, hydrogen peroxide and peroxidase depressed the enzyme activity, whereas catalase was without effect. Among reducing agents ascorbate depressed enzyme activity. Sodium dithionite and sodium borohydride bleached the enzyme with concomitant loss of activity. After reduction with dithionite, the enzyme was rapidly re‐oxidized, re‐gaining its original activity. After reduction with borohydride re‐oxidation was very slow. However, the enzyme could be re‐activated by incubation with FAD.The enzyme is very sensitive to inorganic salts, nitrogen bases and detergents. Chloride is quite deleterious whereas phosphate seems to stabilize the enzyme.
The Selective Extraction of Histone Fractions from Deoxyribonucleoprotein
Authors: Bolund Lars A. , Johns Ernest W. 
Published: March 2005

Three methods are described for quantitative depletion of calf thymus and chicken erythrocyte chromatin of various histone fractions. All methods involve dissociation of the histone fractions in question from chromatin in the presence of the ion‐exchange resin AG50W X 2 which, having strongly bound the free histones in solution, is easily removed from the chromatin. The depleted chromatins do not appear to be denatured.Method 1. In 0.45–0.65 M NaCl, 0.05 M sodium phosphate buffer pH 7.0, histone F1 is quantitatively and selectively removed from calf thymus chromatin. In 0.55–0.65 M NaCl, 0.05 M sodium phosphate buffer pH 7.0 and a limiting amount of ion‐exchange resin, histone F2C is quantitatively and selectively removed from chicken erythrocyte chromatin. Histone F1 can be quantitatively and selectively removed from chicken erythrocyte chromatin by repeated extraction with 0.5 M NaCl pH 7. By further depletion with resin in 0.55–0.65 M NaCl, 0.05 M sodium phosphate buffer pH 7.0, chicken erythrocyte chromatin can be quantitatively and selectively depleted in both histones F1 and F2C.Method 2. In 0.25–0.45M NaCl, 0.05 M sodium phosphate buffer, 4 M urea pH 7.0, histones F1, F2B and F2A2 are quantitatively and selectively removed from calf thymus chromatin.Method 3. In 0.35 M NaCl, 30% ethanol, 4 M urea pH 7.0, an almost complete removal of histones F2A1 and F2A2 from calf thymus chromatin is achieved. In 0.35 M NaCl, 300% ethanol, 5–6 M urea pH 7.0, histone F3 is removed as well.