bluuebunny/crossref_metadata_2025_split huggingface dataset:
Characterisation of RNA Synthesis in vitro in Neurone‐Rich, Oligodendroglial and Liver Nuclei
Authors: MIZOBE Fumio , TASHIRO Tomoko , KUROKAWA Masanori 
Published: March 2005

In neurone‐rich nuclei isolated from the adult guinea‐pig cerebral cortex, DNA‐dependent RNA polymerase B reaction is several times more active than in liver nuclei. This may be attributable to the unusual wealth of euchromatin in mature neuronal nuclei.
Nuclear RNA labelled in vitro was hydrolysed by alkali, and the number as well as the average length of nascent RNA chains were determined by measuring [3H]uridine and [3H]UMP/[3H]uridine ratio, respectively. In neurone‐rich nuclei, the number of growing RNA chains is greater, and the average chain length longer than in liver nuclie. In oligodendroglial nuclei which exhibit RNA polymerase B reaction of intermediate activity, the number of growing RNA chains is found comparable to, but the average chain length longer, than that in liver nuclei. Thus, the presence of a greater number of growing RNA chains appears to be a unique feature of neuronal nuclei.
In neuronal chromatin, mean residue ellipticity in circular dichroism spectrum in the wavelength range of 260–300 nm is significantly higher than in liver and oligodendroglial chromatins, and this is in apparent correlation with the number, rather than the average length, of growing RNA chains.
Digestion of chromatin with trypsin leads to reversion of the circular dichroism spectrum toward that of free DNA, and completely abolishes the difference in ellipticity pattern that is observed among native neurone‐rich, oligodendroglial and liver chromatins.
In liver nuclei, RNA polymerase B reaction is greatly enhanced with trypsin, which is primarily attributable to the facilitated elongation of RNA chains that is otherwise restricted. In neuronal nuclei where the steric restriction imposed on RNA synthesis is though to be intrinsically weakened, digestion with trypsin is more than necessary in disclosing the RNA polymerase B activity.
RNA synthesis in neuronal and oligodendroglial nuclei in vitro is discussed in correlation with that reported in vivo.
Histones from the Fruit Fly Ceratitis capitata
Authors: FRANCO Luis , MONTERO Francisco , NAVLET Jacinto M. , PERERA Julián , ROJO M. Carmen 
Published: March 2005

Histones from the fruit fly Ceratitis capitata have been obtained and characterized by amino acid analysis and polyacrylamide gel electrophoresis. It has been found that the five major histone groups are present in this insect.
The fractionation of histones has been achieved by using acid or organic selective extractions. Ceratitis capitata F1 exhibited a reduced electrophoretic mobility when compared with the mammalian corresponding histone. On the contrary, fly F2A1, mobility was similar to that of mammalian homologous histone. Some differences were found, however, in amino acid composition. It has been found that cysteine does not occur in fly F2A1, but this amino acid seemed to be present in the F3 histone, which aggregated under oxidizing conditions.
A strongly basic protein was isolated together with histones when pharate adult chromatin was used as starting material. Amino acid analysis of this protein revealed the presence of hydroxyproline (19%), as well as high amounts of threonine (30%) and arginine (23%). It was found that this specific protein was not a nuclear component and it seemed to be located at the pharate adult envelope; the name of purarium‐specific protein is proposed for this protein. Its strongly basic nature is judged to be the cause by which it became attached to chromatin during the isolation procedure.
Biological Properties of Lipid A from Shigella sonnei
Authors: ŁUOWSKI Czesław , ROMANOWSKA Elżbieta 
Published: March 2005

Lipid A preparations isolated from Shigella sonnei lipoplysacharides after mild acid hydrolysis were used as immunogens in rabbits. Titers of antisera to lipid A amounted to about 1:640 in passive hemolysis test. Antibodies to lipid A were purified on the column packed with lipid A coupled to Sepharose 4 B. Pure antibodies to lipid A examined in immunoelectrophoresis proved to be immunoglobulins G.The specificity of antisera to lipid A was confirmed in passive hemolysis inhibition test elaborated in this work. From the results of the examination of various products of lipid A degradation in inhibition studies of lipid A—anti‐(lipid A) system, it can be concluded that amino groups of glucosaminyl units substituted by 3‐hydroxymyristoyl residues are immunodominants of lipid A.Lipid A preparations from Shigella sonnei showed distinct anticomplementary activities which disappeared from the examined materials after removal of ester‐linked fatty acids. Shigella sonnei lipopolysaccharides, at suitable concentrations, inhibited also complement fixation. Phase II lipopolysaccharide was the best inhibitor.
Glycosphingolipids and Glycoproteins in the Wild‐Type and in a Non‐Aggregating Mutant of Dictyostelium discoideum
Authors: WILHELMS Otto‐Henning , LÜDERITZ Otto , WESTPHAL Otto , GERISCH Günther 
Published: March 2005

Fractionation of the phenol‐water extract from aggregation‐competent cells of Dictyosteiium discoideum resulted in the isolation of two main fractions containing two different antigen specificities, I and II, respectively. Antigen I was also obtained from growth‐phase cells of Dictyostelium discoideum. This antigen is a glycolipid, and antigen II is a glycoprotein, both containing the same sugar constituents but in different proportions: the ratio of N‐acetylglucosamine—fucose—mannose was 12:5:2 and 12:5:6 in antigens I and II, respectively. The lipid constituents of antigen I were identified as behenic acid and 4‐hydroxysphinanine (C18‐phytosphingosine) present in a ratio of 1:1. In addition, phosphate and ethanolamine were found. Under various conditions, antigen I dissociated into subfractions.The protein component of antigen II was rich in acidic and hydroxy amino acids. The carbohydrate moiety was linked through glucosamine units glycosidically to the hydroxy groups of the hydroxy amino acids. Antigen II strongly reacted with concanavalin A.Cells of a non‐aggregating mutant showed markedly increased agglutinability with wheat germ agglutinin, and changed serospecificity and electrophoretic mobility of antigen I. Extracts of cells from this mutant, harvested at a stage analogous to aggregation competence, yielded fractions similar in composition to antigens I and II. However, the N‐acetylglucosamine—fucose—mannose ratio in this glycolipid was 12:1:3, i.e. the content of fucose was markedly reduced. With N ‐acetyl‐β‐hexosaminidase part of the N‐acetylglucosamine was liberated from this glycolipid indicating that N‐acetylglucosamine is a terminal sugar in the mutant antigen. Various results suggest that fucose is the terminal sugar in the wild‐type glycolipid.
Preparation of Soluble and Insoluble Polysaccharide Conjugates of Peptides with a Free α‐Amino Group
Authors: JOST Rolf , YARON Arieh 
Published: March 2005

Synthetic peptides were covalently attached to Sepharose and water‐soluble dextran of high molecular weight. In these polysaccharide conjugates, the bound peptides bear a free α‐amino group. The interaction of leucine aminopeptidase with its peptide ligands Thr‐Phe‐Pro (substrate) or Thr‐(But)‐Phe‐Pro (competitive inhibitor) was strikingly different in soluble dextran conjugates and insoluble Sepharose conjugates of these ligands. The affinity of the enzyme to its inhibitor Thr(But)‐Phe‐Pro was decreased by more than 100‐fold upon insolubilization of the peptide on Sepharose. When the peptide was coupled to dextran of weight‐average molecular weight ∼ 500000, the potency of the inhibition was not impaired.Leucine aminopeptidase failed to hydrolyze the Sepharose‐bound substrate Thr‐Phe‐Pro to a measurable extent, but it released N‐terminal threonine from the soluble dextran conjugate of this peptide to an extent of at least 38% of the theory.The interference, caused by the insoluble matrix was hardly overcome, when a new type of spacer, consisting of > 80 dl‐alanyl residues, was inserted between matrix and ligand. The spacer which is several enzyme subunit diameters in length, gave rise to a small fraction (about 5%) of accessible ligand but did not improve specific interaction to a sufficient extent in order to enable specific binding of leucine aminopeptidase to inhibitor‐, or substrate‐substituted Sepharoses.Nonspecific adsorption of the enzyme was observed in an inhibitor derivative of Sepharose with ɛ‐aminohexanoic acid inserted as spacer.
The Mechanism of Inhibition of Pig‐Plasma Benzylamine Oxidase by the Copper‐Chelating Reagent Cuprizone
Authors: LINDSTRÖM Anders , PETTERSSON Gösta 
Published: March 2005

Treatment of pig‐plasma benzylamine oxidase with the copper‐chelating reagent cuprizone leads to the reversible formation of an enzyme · cuprizone complex with a dissociation constant of about 3 mM, followed by an irreversible conversion of this complex into an enzyme species exhibiting a new absorption band centered around 350 nm. The latter enzyme species shows no reactivity towards substrate or carbonyl reagents such as phenylhydrazine. One mole cuprizone per mole protein is sufficient to complete the chromophoric changes and to inactive the enzyme completely.
Cuprizone reacts with free pyridoxal phosphate under formation of a reaction product showing maximum absorption at 350 nm in the visible region, and competes with phenylhydrazine for the protein‐bound pyridoxal phosphate in benzylamine oxidase.
Cuprizone does not extract any significant amounts of copper from benzylamine oxidase, but affects electron paramagnetic resonance spectra of the enzyme and its phenylhydrazone derivative in a way suggesting a direct interaction with at least one of the two copper ions present in the protein. The latter interaction does not appear to be identical with those involved in the formation of the inactive enzyme species absorbing at 350 nm.
It is concluded that the inhibitory action of cuprizone cannot be attributed to its copper‐chelating properties, but reflects an interaction with enzyme‐bound pyridoxal phosphate through a mechanism analogous to that governing Schiff‐base or hydrazone formation between enzyme and substrates or hydrazine derivatives.
Effect of Azide on Some Spectral and Kinetic Properties of Pig‐Plasma Benzylamine Oxidase
Authors: LINDSTRÖM Anders , OLSSON Bengt , PETTERSSON Gösta 
Published: March 2005

The effect of azide on optical absorption and electron paramagnetic resonance (EPR) spectra of pig‐plasma benzylamine oxidase has been examined and correlated with the effect of azide on the catalytic activity of the enzyme.
A strong reversible interaction between azide and copper in benzylamine oxidase, corresponding to complex‐formation with a dissociation constant less than 0.2 mM, was detected by EPR techniques. This interaction is competitive with the EPR‐detectable binding of cuprizone to copper in the enzyme, but is not affected by reactions involving the active‐site pyridoxal phosphate and has no evident effect on enzyme activity.
A weaker reversible interaction between azide and protein‐bound copper, corresponding to complex formation with a dissociation constant of 40 mM, was detected spectrophotometrically and by EPR techniques. An absorption band centered at 390 nm is associated with the latter complex, the apparent stability of which was found to be unaffected by a 20‐fold variation of the oxygen concentration as well as by reduction of the enzyme with substrate.
Azide acts as an uncompetitive inhibitor of benzylamine oxidase with an inhibition constant close to 40 mM. 100 mM azide has essentially no effect on the reactivity of the active‐site pyridoxal phosphate towards phenylhydrazine or benzylamine. It is concluded that formation of the enzyme · azide complex exhibiting the 390 nm absorption band precludes reoxidation of the substrate‐reduced form of the enzyme, indicating that at least one of the two copper ions in benzylamine oxidase may have a direct catalytic function in relation to the reoxidation process.
A Conductimetric Study of Erythrocyte Lysis by Lysolecithin and Linoleic Acid
Authors: LAWRENCE Anthony J. , MOORES Geoffrey R. , STEELE James 
Published: March 2005

A linear six‐channel conductivity apparatus has been developed for rapid and accurate kinetic measurements and used to investigate the net efflux of electrolyte from erythrocytes. The pre‐lytic conductance change and a change directly associated with lysis have been measured for cells treated with lysolecithin and linoleic acid.The pre‐lytic response to lysolecithin shows three distinct concentration‐dependent phases which correlate with reported changes in morphology. A mechanism is proposed whereby in low concentration lysolecithin is confined to the outer lamella of the membrane but above a threshold level it may translocate to the inner lamella giving a temporary state of high permeability. The response to linoleic acid is more complex, but basically shows constant or increasing permeability after the initial addition without the apparent formation of transient high permeability states. The kinetic form of the response is suitable for accurate kinetic measurements and the leakage response to unsaturated fatty acids is suggested as a method for examining changes in membrane structure.
A High‐Molecular‐Weight Protein Component Required for Natural Messenger Translation in Ascites Tumor Cells
Authors: STRYCHARZ William A. , RANKI Marjut , DAHL Hans‐Henrik M. 
Published: March 2005

An IF‐M3‐deficient ascites cell‐free system is described.
An IF‐M3 species of high molecular weight has been isolated and partially purified from a high‐KCl ascites ribosomal wash fluid.
Translation of encephalomyocarditis virus RNA and globin mRNA is completely dependent upon the presence of the ascites IF‐M3, while the factor is not required with either poly(A,U,G) or poly(U) as messengers.
The sparsomycin technique for the generation of short initiation peptides has been employed to demonstrate that IF‐M3 acts at the level of chain initiation.
Rabbit reticulocyte IF‐M3 and IF‐E3, as well as rabbit liver IF‐M3, can substitute for ascites IF‐M3 in EMC‐RNA and globin mRNA translation.
The translation of EMC‐RNA in the ascites cell‐free system is, therefore, dependent upon the presence of two discrete initiation factors, a high‐molecular‐weight species, IF‐M3, and a smaller component, IFEMC.
Subunit Interactions in Enzyme Catalysis
Authors: MOUTTET Christian , FOUCHIER Francis , NARI Joannès , RICARD Jacques 
Published: March 2005

By using the principles of structural kinetics previously formulated, the effects of subunit interactions and conformation changes on transient rate of catalysis have been investigated for one‐substrate dimeric enzymes. The analytical solutions of transient rate equations and the expression of the induction time, τ have been obtained for the following models of protomer interactions: simple sequential, partially‐concerted, fully‐concerted, and exclusive allosteric.The progress curve of the reaction during the pre steady‐state phase, as well as the curve expressing the variation of τ−1 against substrate concentration, [S], have been simulated by computer analysis. When the protomers do not exhibit any interaction in the simple sequential mechanism, the plot τ−1versus [S] is a straight line and the induction time is always positive. If subunit interaction occurs but does not change during substrate binding, the plot of τ−1 against [S] is still linear but the length of the induction time depends on the value of the interaction coefficient. If this value is much lower than one, the enzyme, although Michaelian, behaves as “hysteretic” when rapidly mixed with its substrate. In the same way, the exclusive allosteric model also predicts positive z values, but the plot of τ−1 against [S] is a curve reaching a plateau for high concentrations of the substrate. The sequential models (simple, partially‐concerted, fully‐concerted) can give rise to either positive or negative induction time. Moreover, the asymptote of the curve τ−1 against [S] is oblique. It is thus possible, at least in theory, from pre‐steady‐state kinetics alone, to rule out either the exclusive allosteric or the sequential models.The above theoretical developments have been applied to the early kinetics of l‐phenylalanine ammonia‐lyase reaction. The induction time, that can be calculated from the progress curve, is negative for a large range of substrate concentrations. This result allows one to rule out, for that enzyme, the allosteric transition as a possible explanation of cooperativity. Sequential binding of substrate and subunit interactions can thus generate a burst, that is a transient activation of the reaction in the absence of any activator. It is tempting to assume this transient activation to be of regulatory significance.
Enzyme Polymorphism in Rat‐Liver Microsomes and Plasma Membranes
Authors: BLOMBERG Fred , RAFTELL Marika 
Published: March 2005

Detergent‐soluble antigens from rat liver microsomes and plasma membranes were separated by two‐dimensional immunoelectrophoresis. Enzyme activities in the immunoprecipitates were assayed by different staining reactions.Nine different microsomal esterases were detected. Six of these, including one esterase shared with plasma membranes, were inhibited by 0.1 mM difluorophosphate, but not by 1 mM eserine.Plasma membranes and microsomes each contained at least ten nucleoside diphosphatase and triphosphatase‐active antigens. Only three precipitates with these activities were detected when plasma membrane extracts were tested against anti‐microsomal sera or microsomal extracts against anti‐plasma membrane sera. Although the possible immunological identity of these antigens remains to be established these findings suggest that plasma membranes and microsomes have at most three of these nucleoside diphosphatases and triphosphatases in common.Seven microsomal nucleoside diphosphatases and triphosphatases were intimately associated with both acid phosphatase and NADH‐neotetrazolium reductase activities. These complexes also contained phospholipids as shown by incorporation experiments either in vivo, or in vitro, with [14C]‐choline or [14C]ethanolamine. Three microsomal UDPase‐active antigens did not hydrolyze ATP or ADP and were not associated with acid phosphatase or NADH‐neotetrazolium reductase activities. Furthermore, these antigens did not contain any of the labelled phospholipids.Lipid‐containing multienzyme complexes of different composition were also detected in the plasma membranes. The nucleoside diphosphatases and triphosphatases in plasma membrane extracts were associated with NADH‐neotetrazolium reductase activity and two of them also contained l‐leucyl‐β‐naphthylamidase activity. p‐Chloromercuribenzoate (0.1 mM) inhibited all NADH‐neotetrazolium reductase activity in the plasma membrane and microsomal precipitates and 10 mM tartrate all acid phosphatase activity in the microsomal precipitates, leaving the other activities intact. This indicated that the various enzyme activities present in the multienzyme complexes depended on separate catalytic sites.The plasma membrane extracts contained three and the microsomal extracts two l‐leucyl‐β‐naphthylamidase‐active antigens, respectively. The latter two and one of the plasma membrane antigens were not found to be associated with any other enzyme activities or phospholipids, in contrast to the two plasma membrane antigens mentioned above.
Enzyme Polymorphism in Rat‐Liver Microsomes and Plasma Membranes
Authors: RAFTELL Marika , BLOMBERG Fred 
Published: March 2005

Extraction of rat liver microsomes and plasma membranes with the detergents Lubrol W and sodium deoxycholate rendered extracts containing a multiplicity of electrophoretically and immunologically different enzyme‐active antigens as detected in two‐dimensional immunoelectrophoresis. To establish that this enzyme polymorphism was not merely due to an interaction of enzyme molecules with detergent micelles but reflected membrane organization, antigens solubilized by other agents like Triton X‐100, papain, phospholipase A or C were compared with those extracted by Lubrol W and deoxycholate.Different enzyme‐active antigens were differently affected by the various solubilizing agents. Thus phospholipases A and C released the same esterase‐active antigens as did Lubrol‐ deoxycholate. In contrast, phospholipid‐containing multienzyme complexes (expressing nucleoside diphosphatase and triphosphatase, NADH‐neotetrazolium reductase, acid phosphatase and L‐leucyl‐P‐naphthylamidase activities in different combinations) were not solubilized in an enzymatically active form by these agents. On the other hand, such multienzyme complexes were released in significant quantities from the microsomes but not from the plasma membranes, when a highly diluted papain solution was used for extraction. The results are assumed to reflect differences in the molecular organization of individual enzymes in the various membranes.In order to investigate the molecular identities of the antigens solubilized from the membranes with the different agents, 125I‐labelled antigens, extracted with papain or phospholipase A, were mixed with unlabelled detergent‐solubilized antigens. Autoradiography of the immunodiffusion plates showed that the enzyme‐active antigens released by papain or phospholipase A were electro‐ phoretically and immunologically similar to those solubilized by detergents.
Presence of Cytochrome c1 in Cytoplasmic “Petite” Mutants of Saccharomyces cerevisiae
Authors: CLAISSE Maurice L. , PAJOT Patrick F. 
Published: March 2005

To show the occurrence of cytochrome c1 in ϱ− petite mutants of Saccharomyces cerevisiae, in which its presence was questioned, low‐temperature absorption spectra of various ϱ+ϱ− strains have been compared. Some of these strains carry nuclear mutations producing different types of cytochrome deficiencies, thus lowering the spectral interferences on cytochrome c1 and making easier its observation. The comparison was made with whole cells and with mitochondrial preparations. All the ϱ−mutants examined present a shoulder or a peak at a wavelength corresponding to cytochrome c1 in the ϱ+ strains. This absorption band at 552 nm in ϱ− mutants was further identified by isolating a cytochrome with an α absorption maximum at this wavelength. The low‐temperature absorption spectrum of this cytochrome was compared to low‐temperature absorption spectra of cytochrome c1 isolated from a wild‐type strain, and of cytochrome c and cytochrome b2. It is similar, if not identical, to the spectrum of the wild‐type cytochrome c1, and quite distinct of that of the other cytochromes.The cytochrome c1 content, in the various ϱ+and ϱ− strains examined, is estimated with the help of a calculation procedure taking into account the spectral cross interferences of the cytochromes.In the absorption spectra of the ϱ− strains, a shoulder is recorded at about 557–558.5 nm. The nature of this shoulder is discussed.