bluuebunny/crossref_metadata_2025_split huggingface dataset:
Phosphate Transfer between 2,3‐Bisphosphoglycerate and Monophosphoglycerate Catalyzed by Bisphosphoglycerate Phosphatase
Authors: de Verdier Carl‐Henric , GROTH Torgny L. 
Published: March 2005

2,3‐Bisphosphoglycerate phosphatase from human erythrocytes was extensively purified with the use of various chromatographic procedures and electrofocusing. All detectable phosphoglyceromutase activity was removed as tested with an optical assay at pH 7.6.
At the pH optimum of the phosphatase reaction (pH 6.5) the enzyme transfers phosphate groups from 2,3‐bisphosphoglycerate to monophosphoglycerates (3‐phosphoglycerate or 2‐phosphoglycerate). The transfer is more than hundred times faster than the hydrolyzing activity.
Chloride and 2‐phosphoglycolate stimulate the hydrolyzing but not the transferring activity.
Data from seven experiments with differently labelled phosphoglycerates were analyzed in terms of rate constants defined by a mathematical model for the different transfer reactions.

With the direction of phosphate transfer expressed from 2,3‐bisphosphoglycerate to mono‐phosphoglycerate the constants were estimated to have the following values (ml · min−1·μmol−1): C‐3 → C‐2 0.006 ± 0.001, C‐2 → C‐2 0.024 ± 0.003, C‐3 → C‐3 0.009 ± 0.006 and C‐2 → C‐3 0.034 ± 0.008.The values indicate that the velocity of the transfer from C‐2 in 2,3‐bisphosphoglyerate to the vacant carbon atom in 2‐phosphoglycerate is about the same or possibly somewhat higher than the velocity of the transfer to 3‐phosphoglycerate.The transfer velocity from 2,3‐[3‐32P]bisphosphoglycerate to 3‐phosphoglycerate is much lower. Due to the experimental conditions the results do not allow determination of the velocity of the transfer from C‐3 to 2‐phosphoglycerate with any high degree of precision. The C‐3 to C‐2 transfer however, is slower than the transfer from C‐2 in 2,3‐bisphosphoglycerate.
Hormonal Interactions at the Molecular Level
Authors: Camier Maryse , ALAZARD Robert , Cohen Paul , PRADELLES Philippe , Morgat Jean‐Louis , FROMAGEOT Pierre 
Published: March 2005

Bovine neurophysins I and II have been obtained in a highly purified state using isoelectric focusing. The interactions of oxytocin and [8‐lysine]vasopressin with these proteins have been investigated by thin‐film equilibrium dialysis using highly radioactive hormones retaining their full biological activities.

Binding of both oxytocin and [8‐lysine]vasopressin to neurophysins I and II as a function of pH leads to typical “bell‐shaped” curves. The patterns obtained indicate functional differences in the binding of oxytocin to neurophysin I compared with neurophysin II. No significant change was detected in the case of [8‐lysine]vasopressin.
Scatchard plots indicate that both hormones bind neurophysins I and II with similar affinities. They demonstrate that at pH 5.7 one oxytocin molecule and two [8‐lysine]vasopressin molecules can be bound per neurophysin I or II monomer.
Ca2+ and Mg2+ had no effect on the binding process of either [8‐lysine]vasopressin or oxytocin. Iodination on the tyrosyl ring of [8‐lysine]vasopressin and oxytocin resulted in the suppression of their binding ability.
Competition experiments between oxytocin and vasopressin show that each compound inhibits the binding of the other peptide in a non‐competitive fashion.
Identification of Polysomes Synthesizing Thyroglobulin
Authors: Assart Gilbert V , DUMONT Jacques E. 
Published: March 2005

In preliminary experiments, we have shown that membrane‐bound thyroid ribosomes specifically synthesize thyroglobulin. This article presents more precise characterization of the thyroglobulin‐synthesizing polysomes.

A population of heavy membrane‐bound polysomes has been isolated from horse thyroid in the detergent‐treated pellet of centrifugation at 27000 ×g.
Immunoprecipitation experiments performed with labelled nascent peptides have revealed that this class of polysomes is specifically engaged inthyroglobulin synthesis.
The pattern of [3H]uridine incorporation into these polysomes suggest that the thyroglobulin template has a very slow turnover. This is further supported by the unique resistance of these polysomes during long‐time incubations of thyroid slices with actinomycin D.
Demonstration of Two Types of Proteins Synthesized in Isolated Rat‐Liver Mitochondria
Authors: Kadenbach Bernhard , HADVÁRY Paul 
Published: March 2005

Isolated rat‐liver mitochondria incorporate various labeled amino acids into two types of proteins. About one third of the incorporated radioactivity is soluble in chloroform—methanol (2:1, v/v), the other part is insoluble.
The incorporation into the chloroform‐methanol‐soluble proteins is sensitive to chloramphenicol and ethidium bromide, but insensitive to cycloheximide.
The rate of incorporation into the chloroform‐methanol‐soluble proteins is different from that into the insoluble proteins, depending on the time of incubation, the intactness of the mitochondria and the sucrose concentration of the incubation medium.
The possibility of the existence of two different types of protein synthetic systems in mitochondria is discussed.
5‐S RNA: Investigation of the Different Extent of Phosphorylation at 5′ Terminus
Authors: Soave Carlo , NUCCA Roberto , Sala Elide , VIOTTI Angelo , Galante Ennio 
Published: March 2005

5‐S RNA from wheat germ, homogeneous by gel filtration and polyacrylamide gel electrophoresis, yields three peaks by hydroxyapatite chromatography. Experiments are presented which firmly establish that the three peaks are constituted by 5‐S RNA molecules ending at their 5′ terminus with mono‐, di‐, or triphosphate groups. The isolation of 5‐S RNA by direct extraction of RNA from wheat embryos homogenized in different buffers and the inability of Escherichia coli alkaline phosphatase to dephosphorylate unmelted 5‐S RNA molecules suggests that the three species are not an artifact of extraction.The peculiar ability of the hydroxyapatite chromatography to fractionate 5‐S RNA molecules on the basis of the extent of their 5′‐terminal phosphorylation was used to investigate the pattern of 5′‐phosphorylation of 5‐S RNA from various organisms. The most striking feature observed is that 5‐S RNAs from prokaryotic organisms contain only 5′‐monophosphate molecules. In contrast, the presence of 5′ mono‐, di‐ and triphosphate molecules was observed in all the eukaryotic organisms investigated. The significance of this finding in relation to the biosynthesis of 5‐S RNA is discussed.
Studies on Soybean Trypsin Inhibitors
Authors: Koide Takehiko , IKENAKA Tokuji 
Published: March 2005

Soybean trypsin inhibitor (Kunitz) was divided into four peptide fragments by the combination of limited hydrolysis with trypsin at acidic pH, chemical cleavage of methionyl bonds with cyanogen bromide, and reduction and carboxymethylation of disulfides in protein or peptide. Each fragment was separated and purified by gel filtration on Sephadex G‐50, G‐75 or G‐100, and referred to as fragments A, B, C, and D from the amino‐ to the carboxyl‐terminal region of the inhibitor.Soybean trypsin inhibitor (Kunitz) consisted of 181 amino acid residues and its molecular weight was proved to be 20100. Fragments A, B, C, and D consisted of 63, 21, 30, and 67 amino acid residues, respectively. One of the two disulfide bridges in the inhibitor was involved in fragment D, and the other linked fragment A with fragment C.
Purification and Properties of an Inhibitor of the Tryptophan‐Synthase‐Inactivating Enzymes in Yeast
Authors: Ferguson A. Ross , Katsunuma Tsunehiko , Betz Heinrich , Holzer Helmut 
Published: March 2005

An inhibitor of the tryptophan‐synthase‐inactivating enzymes has been isolated from bakers' yeast and purified 27‐fold on a protein basis, by heat treatment, precipitation by trichloroacetic acid, ethanol fractionation and acid precipitation. The inhibitory activity of the purified material is not affected by incubation with RNAase, DNAase, or muramidase, but is destroyed by incubation with pronase or trypsin. Inhibitory activity depends, therefore, on the protein content of the material.The extent to which the inactivating enzymes are inhibited is proportional to inhibitor concentration. Tryptophan‐synthase inactivases I and II are both inhibited. The inhibitor appears to be specific in its action: it is much more effective than soybean trypsin inhibitor in inhibiting the inactivases, but has comparatively little effect on the esterolytic activity of trypsin.Inhibitory activity was measured in crude extracts of Saccharomyces cerevisiae growing in batch culture. In the exponential growth phase, the inhibitor was barely detectable, but its levels subsequently rose at the end of the exponential phase and in the stationary phase. The possible biological significance of these observations is discussed.
A Mutant of Escherichia coli Defective in Ribonucleosidediphosphate Reductase
Authors: Karlström H. Olle , Fuchs James A. 
Published: March 2005

We have studied the ribonucleosidediphosphate reductase of a deoxyuridine‐requiring mutant LD195 of Escherichia coli K. This enzyme was purified 50fold from the mutant and a control strain. In all steps of the purification the mutant strain yielded normal activity of the B1 subunit of the enzyme but approximately 10% of the wild‐type activity of the B2 subunit. The activity is similarly reduced for all three substrates tested CDP, UDP, and GDP.The mutation appears to affect the physical structure of the B2 subunit, since addition of high concentrations of sodium acetate can restore enzymatic activity to the partially purified B2 subunit from the mutant strain. In contrast, the same concentrations strongly inhibit the wild‐type B2 subunit.Although the mutant has a requirement of deoxyuridine at 42°C but not at 30°C (as shown in an accompanying paper), mutant protein B2 is not more thermo‐labile than wild‐type protein B2. This requirement therefore probably reflects an increased demand of deoxyribonucleotides at higher temperatures.Hydroxyurea inhibits mutant and wild‐type ribonucleosidediphosphate reductase to the same extent in vitro. In spite of this, hydroxyurea inhibits growth of the mutant strain at a 20fold lower concentration than that required for comparable inhibition of growth of the parent. This is explained by the greatly decreased protein B2 activity of the mutant.
The Kinetics of the Conversion of Chicken Pepsinogen to Chicken Pepsin
Authors: Bohak Zvi 
Published: March 2005

The kinetics of the conversion of chicken pepsinogen to chicken pepsin was studied at 25°C by following the appearance of peptic activity using a pH‐static method and by following the changes in the fluorescence emission of the protein. It was found that in pH range of 2–4 both processes were first‐order reactions, and that the change in fluorescence emission was faster than the appearance of activity. The results are interpreted in terms of a unimolecular mechanism comprising two steps. In this pH range the zymogen undergoes a rapid conformational change to form an intermediate, and the subsequent conversion of this intermediate to the active enzyme is the rate‐determining step. In this step the amino‐terminal peptide of the zymogen is split by intramolecular catalysis which requires the protonation of a group, presumably a carboxyl group, with a pK of 3.1 in the active site of the enzyme. Above pH 4 the conformational change becomes the rate‐deter mining step in the unimolecular activation process, and a bimolecular pathway in which chicken pepsinogen in its native conformation is activated by active chicken pepsin becomes apparent.It was found that the peptide fragment formed during the conversion of chicken pepsinogen to chicken pepsin, though not an inhibitor of chicken pepsin activity, can combine with chicken pepsin and protect it against denaturation at pH 10.
Adsorption of Nerve Growth Factor onto Surfaces Implications for the Assay in Tissue Culture
Authors: Pearce Frederick L. , BANTHORPE Derek V. , Cook Janet M. , VERNON Charles A. 
Published: March 2005

Apparently minor modifications to the basic method for the assay of nerve growth factor resulted in differences in measured biological activity of up to nine orders of magnitude. These differences were caused by the reversible adsorption of the material onto the apparatus used in the assay procedure. The adsorption was probably promoted by London dispersion forces since the effect persisted both over a range of pH and when glass, plastic and silicone‐coated glass apparatus were used.Under conditions in which the adsorption was minimised, the test material showed a specific activity in tissue culture of between 10−2 and 10−3 [μg ml−1. Previous reports of highly active preparations of nerve growth factor, some having specific activities of about 10−10μg ml−1, can probably be attributed to similar artefacts in the assay method.
Tyrosyl‐tRNA Synthetase of Escherichia coli B
Authors: Chousterman Suzanne , Chapeville François 
Published: March 2005

Using EDTA‐dialyzed tyrosyl‐tRNA synthetase and tRNA, which retain 0.8 and 0.25 atom of Mg2+ per molecule, respectively, tyrosine activation as well as tyrosyl‐tRNA formation were investigated in the absence of added magnesium ions. It is shown that the tyrosyladenylate · enzyme complex is still formed, but the rate of the reaction is reduced by a factor of 104. The rate of the ATP‐PPi exchange reaction, under these conditions, is reduced by a factor of 4 · 104. In the presence of pyrophosphate the predominant reaction is not the pyrophosphorolysis of the enzyme‐bound tyrosyladenylate but its hydrolysis.The rate of tyrosyl‐tRNA formation, starting with free ATP and tyrosine and in the presence of spermidine, was only 15 times lower than in the presence of Mg2+. Spermidine has no effect on the rate of the ATP‐PPi exchange reaction.
Properties of Purified Malic Enzyme in Relation to Crassulacean Acid Metabolism
Authors: Brandon Pieter C. , van Boekel‐mol Titie N. 
Published: March 2005

Malic enzyme from leaves of the Crassulacean species Bryophyllum tubiflorum was purified more than 1200‐fold by a rather simple procedure involving repeated fractionation with ammonium sulphate and gel‐filtration. A specific activity (international standards) of 68 was obtained. The molecular weight of the enzyme, measured by the gel‐filtration method, was 237000.Thiol activation is required for catalytic activity. The activity is completely dependent on divalent cations; greatest activity was obtained with Mn2+, an equimolar amount of Mg2+ was 86% as effective. Maximum activity was obtained at about pH 7.2. At 30 °C and pH 7.2 the Km for Mn2+ was 0.46 mM, that for NADP was 14 μM, and that for malate amounted to 0.37 mM. The Km for malate was temperature‐dependent: it remained constant in the temperature range from 17 to 39 °C, but below 17 °C and beyond 39 °C the affinity for malate decreased and a Km‐value of 0.94 mM was recorded. The enzyme was completely NADP‐specific and was stable at temperatures up to 55 °C; maximum activity was recorded at 50 °C.No effects of monovalent cations such as Na+, K+, Li+, or NH4+ could be recorded. The enzyme was inhibited by coenzyme A and thiamine pyrophosphate, but not by acetyl‐coenzyme A. The effects of temperature as well as the inhibition by coenzyme A and thiamine pyrophosphate fit the physiology of Crassulacean acid‐metabolism.
Fingerprinting Double‐Stranded Non‐Radioactive RNA from a Fungal Virus
Authors: Loviny Thérèse , Székely Maria 
Published: March 2005

The fingerprinting technique has been applied to double‐stranded viral RNA. Under appropriate conditions pancreatic and T1 ribonuclease digests were obtained which upon two‐dimensional paper electrophoresis yielded specific patterns, similar to those obtained from single‐stranded RNAs. The structure of some simpler nucleotides on the fingerprints have been determined. The method seems suitable to characterize and sequence all kinds of double‐stranded RNA molecules.Labelling of the virus in vivo yielded RNA preparations of rather low radioactivity. Polynucleotide kinase was used therefore to label the digest of non‐radioactive RNA for fingerprinting. The possibilities and limitations of this technique for sequencing non‐radioactive nucleic acids is discussed.
Etude de l'acide polyxanthylique et de ses complexes
Authors: Bachner Lucien , Massoulié Jean 
Published: March 2005

Poly(X) displays two kinds of thermal dissociations, corresponding to two different states of the polymer. At low temperatures, and for values of pH which are not too high, the polymer is organized in a multi‐stranded structure, in which the xanthylic residues are partially ionized, i.e.carry negative charges; this structure is called poly(Xα−)n, where α is the degree of ionization, and n the number of strands. α is a variable parameter. The thermal dissociation of this structure is cooperative, and its transition temperature (TmX) is maximum at pHmax. pHmax depends upon the saline concentration.In alkaline solutions, the xanthylic residues are totally ionized: poly(X−) shows another type of thermal transition. Even though this transition is not cooperative, it differs from purely non‐cooperative transitions in that its melting temperature is dependent upon salt concentration (ΔTm= 13 °C for a factor of 10), and the ΔH value is extremely high (109 kJ/mol, or 26 kcal/mol).Poly(X) forms a double‐stranded complex with poly(U) in which the xanthylic residues are negatively charged, poly(X−) · poly(U). With poly(I) we find two complexes. The pH range of stability of these complexes implies that the degree of ionisation of the xanthylic groups is variable, as we found in the structure poly(Xα−)n: These two‐ and three‐stranded complexes, poly(Xα−) · poly(I) and poly(Xα−) · 2 poly(I) are differently affected by ionic strength. We describe the stability diagrams (temperature, pH, ionic strenght) for all these structures, and we present possible hydrogen bonding schemes. It appears that the negative charges of the xanthylic residues help strengthen the hydrogen bonds, so that the stability of these structures depends upon this ionization (α): the balance between this factor and electrostatic repulsion would then explain the occurrence of an optimum pH value for the stability of the structure.
Studies on the Fluorescence of 1‐Anilino‐8‐naphthalenesulfonate by the Membranes of the Sarcoplasmic Reticulum
Authors: Augustin Jan , Hasselbach Wilhelm 
Published: March 2005

The quantum yield of 1‐anilino‐8‐naphthalenesulfonate associated with native and lipid‐depleted sarcoplasmic vesicles is nearly identical. The slight increase of the average apparent dissociation constant of the interaction of this dye with lipid‐depleted vesicles is accompanied by a decrease in the number of binding sites for the dye.
Oleic acid is a noncompetitive inhibitor of the anilino‐naphthalenesulfonate binding to native and lipid‐depleted sarcoplasmic vesicles.
Rising temperatures decrease the quantum yield and increase the average apparent dissociation constant.
The polarisation of the fluorescence of membrane‐bound anilino‐naphthalenesulfonate reaches relatively high values with a significant increment for lipid‐depleted sarcoplasmic vesicles. There is only a slight temperature dependence and no viscosity dependence for both native and lipid‐depleted vesicles. The polarisation is reduced by rising concentrations of anilino‐naphthalenesulfonate.
Energy transfer measurements indicate that the dye is located very closely to the membrane‐bound tryptophan, suggesting a donor‐acceptor distance of 2 nm.
Biosynthesis of Cartilage Procollagen
Authors: Dehm Peter , Prockop Darwin J. 
Published: March 2005

Cells were isolated by controlled enzymic digestion of sternal cartilages from chick embryos and the cells were studied immediately after isolation for up to 4 h in vitro. The cells isolated under the conditions employed had a high degree of viability, they remained in suspension for 2 to 4 h in vitro and during this time they synthesized and secreted collagen in the precursor form which has been called “transport form” or “procollagen”. The cells also incorporated [35S]sulfate into macromolecules which were secreted into the medium.Continuous labeling of the cartilage cells demonstrated a lag of about 33 min between the time at which the rate of synthesis of collagen [14C]hydroxyproline became linear and the time at which the secretion of collagen [14C]hydroxyproline into the medium became linear.Gel filtration and polyacrylamide‐gel electrophoresis in sodium dodecylsulfate indicated that all the secreted collagen was in the precursor form. The procollagen differed from the procollagen previously isolated from bone, tendon and cultured fibroblasts in that it apparently consisted of a single type of polypeptide chain. Isotopic studies suggested that the NH2‐terminal extension on the cartilage procollagen contained cystine and tryptophan and appeared therefore to be similar in this respect to that of other procollagens.
Characterization of Five Neurotoxins Isolated from the Venoms of Two Elapidae Snakes Naja haje and Naja nigricollis
Authors: Kopeyan Charles , van Rietschoten Jurphaas , Martinez Gérard , Rochat Hervé , Miranda François , Lissitzky Serge 
Published: March 2005

Two neurotoxins were purified from Naja nigricollis venom using equilibrium chromatography. This method was shown to produce the most resolution to purify proteins having very close physico‐chemical properties. The toxins obtained were characterized, as well as three neurotoxins previously isolated from Naja haje venom, by the determination of N‐terminal (35 to 39 residues) and C‐terminal sequences.The usefulness of N‐terminal automatic sequencing for rapid identification of proteins is discussed.
Biochemical and Immunological Characterization of Threonyl‐tRNA Synthetase of Two Borrelidin‐Resistant Mutants of Escherichia coli K12
Authors: Paetz Wilfried , Nass Gisela 
Published: March 2005

The threonyl‐tRNA synthetase of two wild‐type Escherichia coli K12 strains and of two borrelidin‐resistant mutants thereof has been purified about 100‐fold. A neutralizing antiserum has been prepared against one of the wild‐type threonyl‐tRNA synthetases. The comparative characterization of the threonyl‐tRNA synthetases of the two wild strains and the two borrelidin‐resistant mutants revealed that the threonyl‐tRNA synthetase of one borrelidin‐resistant mutant (K12B borr 2) exhibits a lowered Km value for threonine and ATP and an increased K1 value for borrelidin with respect to threonine. Furthermore the activity of the threonyl‐tRNA synthetase of this mutant k more sensitive to higher pH values, is more rapidly heat‐inactivated and shows a different shape of its antiserum‐neutralization curve than the threonyl‐tRNA synthetase of the parental strain (K12B). The threonyl‐tRNA synthetase of the other borrelidin‐resistant mutant (K12B borr 3) was investigated in the same way and it did not show any difference to the threonyl‐tRNA synthetase of the parental strain, except that the specific activity of the enzyme in crude extracts of this mutant is 5 times as high as the corresponding value in crude extracts of K12B.From these data it is concluded that borrelidin resistance in K12B borr‐2 is due to a structurally altered threonyl‐tRNA synthetase (structural mutant) whereas in K12B borr‐3 it is due to the constitutive increase of the threonyl‐tRNA synthetase level by a factor of five (regulatory mutant).The inhibition of the threonyl‐tRNA synthetase by borrelidin with respect to threonine was noncompetitive in all four cases. In order to observe a considerable heat inactivation of the threonyl‐tRNA synthetase within 10 min a temperature of 60°C was necessary. The heat‐inactivation process was not affected by the presence of threonine, but it was hastened by the presence of ATP or tRNA.