bluuebunny/crossref_metadata_2025_split huggingface dataset:
Studies on the Structure and Function of Chick‐Oviduct Chromatin
Authors: STRÄTLING Wolf H. , VAN Nguyen T. , O'MALLEY Bert W. 
Published: March 2005

Chick oviduct chromatin was separated into a ribonucleoprotein fraction and two chromatin fractions (early and late eluting). We utilized a gentle procedure in which moderately hydrated chromatin was subjected to chromatography on a weak ionic‐exchange resin (ECTHAM‐cellulose) eluted with a combined pH‐salt gradient. Chemical analysis of the early (fraction I) and late (fraction II) eluting fractions revealed that their histones were identical and their nonhistone proteins were markedly different. Control experiments showed that these differences were not due to protein rearrangements during chromatin preparation and/or fractionation.The physical properties of fraction I and II differed in certain aspects. The aggregation response of fraction I to increasing concentrations of monovalent cations was five times lower than that of fraction II but the aggregation response to divalent cations was identical. Thermal denaturation assays of DNAs isolated from fractions I and II revealed identical derivative profiles of hyperchromicity vs temperature, thereby indicating similar base composition in the two fractions. Circular dichroism spectra of the purified DNAs isolated from both fractions showed identical B‐type conformations. However, DNA renaturation kinetics analyzed by computer technique indicated that fraction I DNA, contained less than half the amount of highly repetitive sequences as compared to either unfractionated chromatin or fraction II. Circular dichroism spectra of fraction I and II chromatins (at room temperature) showed significant differences in a wavelength region were only DNA is optically active (i.e: 255–320 nm). These results indicated that the DNA complexed to proteins in fraction II assumed a more C‐type conformation than the DNA in fraction I. The differences in the circular dichroism spectra could not be accounted for by differences in the RNAs or protein chromophores contained in fraction I and fraction II. When the circular dichroism spectra of fraction I and II were recorded at 55°C, the differences between the two fractions were abolished. These results were interpreted to mean that the differences in the DNA conformations found in fractions I and II were due to the differences in their nonhistone proteins. These proteins were effective in maintaining DNA conformation differences only when they were in their native form but not when heated to 55°C.Comparison of the sedimentation coefficients of fractions I and II with their calculated molecular weights suggested a more extended structure in fraction I as compared to a more compact structure in fraction II. Only small differences were observed between fraction I and fraction II with respect to either buoyant density analysis in a metrizamide gradient or in the number of phosphate charges accessible to polylysine.
Valyl‐tRNA, Isoleucyl‐tRNA and Tyrosyl‐tRNA Synthetase from Baker's Yeast
Authors: FREIST Wolfgang , VON DER HAAR Friedrich , FAULHAMMER Heinz , CRAMER Friedrich 
Published: March 2005

Nineteen analogs of ATP have been tested in the aminoacylation reaction of valyl‐tRNA, isoleucyl‐tRNA and tyrosyl‐tRNA synthetases from baker's yeast. Four compounds are substrates for valyl‐tRNA and two for isoleucyl‐tRNA synthetase, but there is no modified substrate for the tyrosyl‐tRNA synthetase. There is one inhibitor for valyl‐tRNA synthetase, eight compounds inhibit isoleucyl‐tRNA synthetase and two compounds inhibit tyrosyl‐tRNA synthetase. Their Km and Kt and V values have been determined.The substrate specificity shows that positions 2, 6, 7, 8, 9, 2′, and 3′ of ATP are important for catalytic action of these aminoacyl‐tRNA synthetases.
Activity and Subcellular Distribution of Protein Kinase Dependent on Adenosine 3′: 5′‐Monophosphate in Liver from Normal and Adrenalectomized Rats
Authors: ROUSSEAU Guy G. , MARTIAL Joseph , DE VISSCHER Michel 
Published: March 2005

We have examined whether glucocorticoids control the activity and (or) the subcellular distribution of protein kinase dependent on cyclic'AMP (adenosine 3′: 5′‐monophosphate), since they influence, cyclic‐AMP‐dependent responses to other hormones.Protein kinase activity was determined in rat liver homogenates and subcellular fractions, nuclear, large granular, microsomal and supernatant obtained by differential sedimentation in 0.25 M sucrose. 63% of the tissue protein kinase activity detected in absence of cyclic AMP reside in the particulate fractions. Upon addition of exogenous cyclic AMP, protein kinase activity is stimulated 1.8, 1.2, 1.2 and 4.5‐fold in nuclear, large granular, microsomal and supernatant fractions, respectively. Under these conditions, 66% of tissue activity are found in the supernatant fraction. The activity sensitive to exogenous cyclic AMP resolves into a major (84%) cytosoluble and a minor (16%) nucleomicrosomal component. The latter activity resists elution with isotonic saline and is increased in the presence of Triton X‐100.Three groups of rats were studied: control and adrenalectomized with or without cortisol treatment. In whole liver homogenates, both protein kinase activity detected in absence of exogenous cyclic AMP andsensitivity of the enzyme to cyclic AMP were comparable in all groups. Moreover, the distribution patterns of protein kinase activity among the fractions were essentially the same in all groups of animals, whether or not particles had been treated with Triton X‐100.Finally, in cell‐free experiments, glucocorticoids alone or in combination with their intracellular receptor did not modify protein kinase activity of rat liver.Thus the results reported do not support the possibility that glucocorticoids influence cyclicAMP‐dependent protein kinase in rat liver. Yet, this study provides data, not available before, on subcellular distribution of this enzyme in rat liver.
Multifunctional Enzyme, Bisphosphoglyceromutase/2,3‐Bisphosphoglycerate Phosphatase/Phosphoglyceromutase, from Human Erythrocytes
Authors: IKURA Koji , SASAKI Ryuzo , NARITA Hiroshi , SUGIMOTO Etsuro , CHIBA Hideo 
Published: March 2005

Bisphosphoglyceromutase and 2,3‐bisphosphoglycerate phosphatase activities responsible for 2,3‐bisphosphoglycerate metabolism in human red cells are displayed by the same enzyme protein which has phosphoglyceromutase activity [Sasaki, R., et al. (1975) Eur. J. Biochem. 50, 581–593]. This enzyme was subjected to chemical modification by trinitrobenzenesulfonate.The three enzyme activities were inactivated by trinitrobenzenesulfonate at the same rate. The sulfhydryl content of the enzyme was unchanged during trinitrophenylation, indicating that derivatization was through the amino group. Trinitrophenylation of about one amino group per mole of the enzyme resulted in complete loss of the three activities.Both 2,3‐bisphosphoglycerate and 1,3‐bisphosphoglycerate inhibited trinitrophenylation and effectively protected the enzyme from inactivation. Although monophosphoglycerates did not show any protective effect at concentrations which should be adequate based upon their kinetic constants, they were protective at higher concentrations.Inactivation by trinitrophenylation was an apparent first‐order reaction. The dissociation constant of the enzyme 2,3‐bisphosphoglycerate complex was determined by analyzing the first‐order reaction on the assumption that the protective effect of 2,3‐bisphosphoglycerate was due to competition with trinitrobenzenesulfonate. The dissociation constant was in good agreement with kinetic constants of 2,3‐bisphosphoglycerate in the enzyme reactions, which indicated that 2,3‐bisphosphoglycerate did indeed exert its protective effect through competition with trinitrobenzenesulfonate for an amino group of the enzyme. The protective effect of monophosphoglycerates could be rationalized with kinetic evidence that 2‐phosphoglycerate at high concentrations interacts with the 2,3‐bisphosphoglycerate binding site.These results indicate that the enzyme exhibits the three enzyme activities at a common active site at which one amino group essential for binding of bisphosphoglycerates is located. Based on the multifunctional properties of this enzyme, a possible mechanism was discussed for regulation of 2,3‐bisphosphoglycerate metabolism in human red.
Subunit Structure and Multifunctional Properties of Yeast Phosphoglyceromutase
Authors: SASAKI Ryuzo , UTSUMI Shigeru , SUGIMOTO Etsuro , CHIBA Hideo 
Published: March 2005

A new and efficient method for preparation of pure phosphoglyceromutase from baker's yeast (Saccharomyces cerevisiae) is described. Proteolytic alterations of the enzyme during extraction can be minimized by grinding the dried yeast with aluminium oxide at low temperature.
Yeast phosphoglyceromutase contains four highly similar, probably identical subunits of molecular weight 28000, a conclusion based on the following observations. Polyacrylamide gel electrophoresis containing dodecylsulphate or urea gives a single band, indicating that the enzyme is composed of four subunits similar in their molecular weight and net charge. Cyanogen bromide cleavage and, tryptic digestion of the enzyme yield the number of peptides expected for identical subunits from the amino acid composition analysis.
The purred phosphoglyceromutase preparation has bisphosphoglyceromutase activity synthesizing‐2,3‐bisphosphoglycerate from 1,3‐bisphosphoglycerate and 3‐phosphoglycerate. It has been reported that yeast phosphoglyceromutase catalyzes the hydrolysis of 2,3‐bisphosphoglycerate at the same active site which catalyzes the phosphoglyceromutase reaction [Sasaki, R. et al. (1971) Biochim. Biophys. Acta, 227, 584–594, 595–607]. Immunological studies and chemical modification experiments indicate that bisphosphoglyceromutase activity also is due to the phosphoglyceromutase protein and involves amino groups which have been shown to be essential for the other two activities.
Somatic Antigens of Shigella
Authors: DMITRIEV Boris A. , KNIREL Yuriy A. , KOCHETKOV Nikolay K. , HOFMAN Irihna L. 
Published: March 2005

The O‐specific polysaccharide obtained from the lipopolysaccharide of Shigella dysenteriae type 1 (Shigella shiga) by mild acid hydrolysis followed by fractionation on Sephadex G‐50 was found to be identical to that described by Morgan's group and was composed of l‐rhamnose, d‐galactose and N‐acetyl‐d‐glucosamine in a ratio 2:1:1. On the basis of methylation analysis data the polysaccharide was proved to be a linear chain of monosaccharide residues in pyranose forms substituted at position 3, except for that of galactose substituted at position 2.Selective cleavage, based on the N‐deacetylation reaction of the polymer, together with determination of linkage configurations by chromic anhydride oxidation showed that the O‐specific polysaccharide is built up of repeating tetrasaccharide units whose proposed structure is given below‐3)‐α‐l‐Rhap(1–3)‐α‐l‐Rhap(1–2)‐α‐d‐Galp(1–3)‐α‐d‐GlcNAcp(1‐ where Rhap = rhamnopyranose, Galp = galactopyranose, and GlcNAcp =N‐acetyl‐glucosamine.The present findings confirmed the considerations of Heidelberger on the substitution patterns of l‐rhamnose and d‐galactose residues from the results of serological studies.
The Binding of Glycoconjugates to Human‐Milk d‐Galactosyltransferase
Authors: PRIEELS Jean‐Paul , DOLMANS Marcel , SCHINDLER Melvin , SHARON Nathan 
Published: March 2005

Through the use of affinity chromatography, a homogeneous preparation of human β(1 → 4)‐d‐galactosyltransferase (the A protein of lactose synthase) was obtained. The specificity of this protein for glycoconjugates was studied in the presence and absence of human α‐lactalbumin.A kinetic analysis of the transfer of d‐galactose to N‐acetyl‐d‐glucosamine and to β(1 → 4)‐linked N‐acetylglucosamine oligomers, suggested that the active site region of the enzyme contains more than one binding site for acceptor molecules. Furthermore, experiments with N‐acetylglucosamine‐β(1 → 4)‐N‐acetylmuramic‐pentapeptide isolated from Micrococcus luteus indicated that the presence of a peptide chain does not enhance enzymic activity, as compared with the corresponding free disaccharide. Similar results were obtained using ovalbumin and the ovalbumin glycopeptide (which have similar apparent Km values for A protein) as galactose acceptors. In contrast to its ability to inhibit N‐acetyllactosamine production, α‐lactalbumin did not inhibit the transfer of d‐galactose to the N‐acetylglucosamine oligomers or the glycopeptides.Although α‐lactalbumin can switch the specificity of A protein from N‐acetyl‐d‐glucosamine to d‐glucose resulting in the production of lactose, no transfer of galactose was observed to β(1 → 4)‐linked glucose oligomers or to a collagen glycopeptide, d‐glucopyranosyl‐α(1 → 2)‐d‐galactopyranosyloxy‐β(1 → 5)‐lysine. It therefore appears that α‐lactalhumin can only modify human A protein for monosaccharide acceptors.
Continuous Measurement and Rapid Kinetics of ATP Synthesis in Rat Liver Mitochondria, Mitoplasts and Inner Membrane Vesicles Determined by Firefly‐Luciferase Luminescence
Authors: LEMASTERS John J. , HACKENBROCK Charles R. 
Published: March 2005

Oxidative phosphorylation in rat liver mitochondria, the mitoplast or inner membrane‐matrix fraction, and inverted inner membrane vesicles was studied with firefly luciferase luminescence. All preparations showed relatively high ATP/O ratios under initial reaction conditions in the presence of Mg2+, but only intact mitochondria demonstrated significant respiratory control. Mitochondria and mitoplasts, but not vesicles, catalyzed nearly complete conversion of ADP to ATP. Vesicles converted between 50% and 70% of added ADP to ATP establishing a phosphate potential of 9.8 to 10.5 kcal/mol (43.9 kJ/mol). In “loosely coupled” submitochondrial systems which do not display respiratory control, the phosphate potential appears to be a useful parameter of the integrity of the phosphorylating membrane, while ATP/O ratios may be dependent on the relative concentrations of ADP and ATP in the reaction medium.The rapid kinetics of oxidative synthesis of ATP by inverted inner membrane vesicles was investigated in stopped‐flow rapid‐mixing experiments. After an oxygen pulse to reduced vesicles, ATP synthesis commenced within 20 ms. In antimycin‐inhibited vesicles, ATP synthesis resulting from the rapid oxidation of the terminal portion of the respiratory chain commenced within 20 ms and took approximately 100 ms for half completion. Since half oxidation of the terminal portion of the respiratory chain occurs within a few milliseconds, ATP synthesis resulting from such redox reactions must occur over a significantly longer time course than the redox reactions themselves.
Purification, Subunit Structure and Partial Amino‐Acid Sequence of Anthranilate‐5‐phosphoribosylpyrophosphate Phosphoribosyltransferase from the Enteric Bacterium Serratia marcescens
Authors: LARGEN Michael , MILLS Stanley Ely , ROWE Joan , YANOFSKY Charles 
Published: March 2005

The enzyme anthranilate‐5‐phosphoribosylpyrophosphate phosphoribosyltransferase from Serratia marcescens was purified to apparent homogeneity. The purification procedure included ammonium sulfate precipitation, DEAE‐cellulose chromatography, Sephadex gel filtration and hydroxyapatite chromatography.The molecular weight of the native protein as determined on a calibrated Sephadex G‐200 column was 45000. Dodecylsulfate‐polyacrylamide gel electrophoresis in the presence of reducing agent revealed a subunit molecular weight of 43000 ± 900, suggesting that the enzyme exists as a monomer.The sequence of the amino‐terminal 38 residues revealed that three amino acids, glutamine (six residues), glutamic acid (five residues) and serine (five residues) comprised 42% of the sequence composition.
The O9 Antigen of Escherichia coli
Authors: PREHM Peter , JANN Barbara , JANN Klaus 
Published: March 2005

The lipopolysaccharide from Escherichia coli O9: K30 was isolated in about 2% yield with aqueous 45% phenol at 65 C, followed by ultracentrifugation. The polysaccharide moiety was obtained by graded hydrolysis and gel permeation chromatography. It consisted of a mannan which carried oil its reducing end the core oligosaccharide of the R1 type. The mannan contained 1 → 2 and 1 → 3 linkages in a ratio of 3:2 as determined by methylation analysis and mass spectrometry. On periodate oxidation, 58% of the mannose residues were destroyed.Degradation of oligosaccharide mixtures with α‐mannosidase from jack bean meal, as well as a specific rotation of [α]25D=+ 89° indicated that all mannosyl linkages have the α‐configuration.Smith degradation resulted in the liberation of mannosyl(1 → 3)‐mannose (bound to glyceraldehyde), as established by methylation analysis.From these results we conclude that the O9 polysaccharide of E. coli has a pentasaccharide repeating unit of α‐mannosyl(1 → 3)‐α‐mannosyl‐(1 → 2)‐α‐mannosyl‐(1 → 2)‐α‐mannosyl‐(1 → 2)‐mannose, which are joined in the polysaccharide through α‐(1 → 3)‐mannosyl linkages.
The Role of Interchain Disulphide Bridges in the Conformational Stability of Human Immunoglobulin G1 Subclass
Authors: VENYAMINOV Sergei Yu. , RAJNAVÖLGYI Éva , MFDGYESI György A. , GERGELY János , ZÁVODSZKY Péter 
Published: March 2005

The hydrogen‐deuterium exchange data of human immunoglobulin G1 (IgG 1) are interpreted by assuming fast fluctuations of the protein conformation, through which the peptide groups become exposed to the solvent. The probability of solvent exposure of peptide hydrogens reflects a rather loose conformation for native IgG in comparison with other globular proteins. The probability of solvent exposure is greater than 10‐3 for half of the peptide groups, which shows that the conformational transitions by which these groups are exposed to the solvent are accompanied by changes in standard free energy less than 17 kJ/mol (4 kcal/mol).In the range of pH 6.2–8.45, at 25 °C no gross conformational changes are reflected in the hydrogen‐deuterium exchange behaviour of the native, the reduced‐nonalkylated‐reassociated and the reduced‐S‐alkylated‐reassociated IgG 1.No difference could be detected in the conformational stability of the native and reoxidised reassociated IgG1 proteins. The lack of inter‐subunit disulphide bridges in S‐alkylated‐reassociated molecules results in all increased conformational motility. This destabilization of protein conformation affects about 90% of the peptide groups covered by the measurements, and corresponds to changes in standard free energy of 8 kJ/mol on the average.
A Comparison of Some Kinetic Properties of Soluble and Bound Lactate Dehydrogenase Isoenzymes at Different Temperatures
Authors: NITISEWOJO Poedijono , HULTIN Herbert O. 
Published: March 2005

A comparison was made of some kinetic properties of three chicken lactate dehydrogenase iso‐enzynmes (1, 3 and 5) at 4, 16, 23 and 40 °C. Assays were performed with an enzyme concentration of 0.01 μM at pH 6.0. Under the conditions of assay, lactate dehydrogenase 3 and 5 bound to the particular fraction of homogenized skeletal muscle and were evaluated in the soluble and particulate state.Binding of isoenzymes 3 and 5 to the cellular particulate fraction decreased V. This decrease was much greater for lactate dehydrogenase 5 than 3. Values of V for lactate dehydrogenase isoenzymes 1 and 3 did not follow a simple Arrhenius relationship; there was a rapid change in activity between 16 and 23 °C.The apparent Km (pyruvate) values of all isoenzymes (bound or soluble) increased with increasing temperature, changing 4—10‐fold. The apparent Km for lactate dehydrogenase 5 was greater than that for lactate dehydrogenase 3, which in turn was greater than that for lactate dehydrogenase 1.
Raffinose Metabolism in Escherichia coli K12
Authors: SCHMID Kurt , SCHMITT Rüdiger 
Published: March 2005

The utilisation by Edcherichia coli K12 of raffinose as sole carbon source depends on a new raffinose transport system, an invertase and an α‐galactosidase specified by the Raf‐plasmid D1021. The α‐galactosidase was purified to homogeneity from a mutant strain with constitutive synthesis of the enzyme, α‐Galactosidase hydrolyzes p‐nitrophenyl‐α‐d‐galactoside (Km 0.14 mM), methyl‐α‐d‐galactoside (Km 30mM), melibiose (Km, 3.2 mM) and rallinose (Km 60 mM). The enzymatic activity is strongly inhibited by Ag+, p‐chloromercuriphenyl sulfonic acid and, to a lesser extent, by iodoacetamide. Isoelectric focusing indicates the existence of one form of α‐galactosidase with an isoelectric point of 5.1. The purified enzyme has an SW,2o value of 11.7 ± 0.3S and a molecular weight of 329000 ± 4000; this value is not reduced at high dilutions. When examined by dodecylsulphate gel electrophoresis, purified α‐galactosidase yields a single subunit band of molecular weight 82000 suggesting that the intact enzyme consists of four subunits. Amino acid analysis indicates the presence of approximately 712 amino acid residues per quarter molecule including 8 half‐cystine residues. No carbohydrate moiety has been detected. High resolution electron micrographs and Markham rotation of α‐galactosidase show enzyme molecules of approximately 11 × 11 nm containing four globular subunits in a tetragonal arrangement.The plasmid‐coded α‐galactosidase differs from the homologous E. coli enzyme by substrate affinities. cofactor requirements, stability and toluene resistance. It can, therefore, be used as a marker enzyme suitable for the detection in vivo of Raf‐plasmids
The Inhibition of Isocitrate Oxidation by Palmitoyl‐l‐carnitine and Palmitoyl‐CoA in Rat Liver Mitochondria
Authors: LENARTOWICZ Ewa , WINTER Christa , KUNZ Wolfgang , WOJTCZAK Anna B. 
Published: March 2005

Palmitoyl‐l, carnitine decreases the oxidation of isocitrate in rat liver mitochondria in state 3 by 25–30% Palmitoyl‐l‐carnitine acts as an additional substrate raising the‐rate of oxidative phosphorylation, NAD reduction and ATP/ADP ratio in mitochondria.Palmitoyl‐CoA added to mitochondria oxidizing isocitrate in state 3 causes a strong inhibition of isocitrate oxidation and of oxidative phosphorylation and a considerable elevation of intramitochondrial NADH/NAD and ATP/ADP ratios. The effect of palmitoyl‐CoA is dependent on its concentration and is competitive with ADP. Carnitine restores only oxidative phosphorylation, but the oxidation of isocitrate remains inhibited. Evidence is presented that the transport of isocitrate is not affected by palmitoyl‐CoA in this case. From these results it can be concluded that the action of palmitoyl‐CoA is due to the inhibition of adenine nucleotide translocation.The kinetic studies of NAD‐dependent isocitrate dehydrogenase in the soluble fraction of sonicated mitochondria revealed that the enzyme is very sensitive towards the inhibition by NADH and only very slightly affected by ATP (Ki for NADH and ATP are 0.017 and 3.6 mM respectively). On the basis of the kinetic data the relative contribution of NADH and ATP in the inhibition of isocitrate oxidation by fatty acids was calculated.It is concluded that the inhibition of isocitrate oxidation caused by palmitoyl‐l‐carnitine and palmitoyl‐CoA is primarily due to the increased reduction of NAD, whereas the increase of ATP/ADP ratio is much less important.