bluuebunny/crossref_metadata_2025_split huggingface dataset:
Further Evidence for the Structure of the Teichoic Acids from Bacillus stearothermophilus B65 and Bacillus subtilis var. niger WM
Authors: DE BOER Willem R. , WOUTERS Jan T. M. , ANDERSON Alistair J. , ARCHIBALD A. Ronald 
Published: March 2005

Bacillus stearothermopnilus B65 and Bacillus subtilis var, niger WM both contain teichoic acids in their walls composed of glycerol, phophate and glucose. The 13C nuclear magnetic resonance spectrum of B. stearothermophilus teichoic acid showed 13C‐31P coupling on the signals from the C‐5 and C‐6 carbon atoms of the glucose molecule and an α‐glucosidic linkage between glucose and the C‐1 atom of the glycerol moiety. These data are consistent with a poly[glucosylglycerol phospate] as the cell‐wall teichoic acid in the organism. B. subtilis var. niger WM teichoic acid was oxidized by periodate and incubated in glycine buffer at pH 10.5. This treatment did not significantly increase the phosphomonoester content (by β‐elimination of the phosphate groups) of the teichoic acid molecule (7.1 to 9.5%). which is in accordance with earlier data derived from 13C nuclear magnetic resonance spectrocopy [De Boer et al. (1976) Eur. J. Biochem. 62, 1–6], that in this organism the glucose is not an integral part of the polymer chain. Similar treatment of B. stearothermophilus B65 teichoic acid increased the phosphomonoester content of the preparation from 0.15 to 68.1%.
Electron Transport in the cni‐1 Mutant of Neurospora crassa
Authors: EDWARDS David L. , KLEIN Jerry L. , WARDEN Joseph T. 
Published: March 2005

Mitochondria from the nuclear mutant cni‐1 have no optically detectable cytochrome aa3 in early log phase growth. These mitochondria have a high level of respiration that is not inhibited by cyanide but is inhibited by salicylhydroxamic acid. They also show a substantial amount of cyanide‐sensitive respiration.
As cultures of mutant cni‐1 age, flux through the hydroxamate‐sensitive pathway decreases markedly while flux through the cytochrome chain remains constant.
Growth studies with mutant cni‐1 indicate that the cytochrome chain in this mutant is more important in supporting growth than the hydroxamate‐sensitive pathway.
Measurements of the steady‐state level of reduction of cytochrome c in mutant cni‐1 indicate that the rate‐limiting step in the cytochrome chain is at the position occupied by cytochrome oxidase.
Electron spin resonance studies with cni‐1 mitochondria show normal cytochrome oxidase signals in the g ≅ 6 region although there is little or no optically detectable cytochrome aa3.
The Interaction of Daunomycin with Polydeoxynucleotides
Authors: PHILLIPS Don R. , DiMARCO Aurelio , ZUNINO Franco 
Published: March 2005

The ability of daunomycin to bind to various DNA polymers has been studied by thermal denaturation, spectrophotometric analysis and inhibition of the polymerisation reactions catalysed by Escherichia coli DNA polymerase I and rat liver DNA polymerase α. The quantitative binding measurements revealed that the antibiotic binds tightly to all synthetic polydeoxynucleotides studied. The results demonstrated that daunomycin can bind with equal affinity to dG · dC or dA · dT base‐paired sequences. However, the number of binding sites per nucleotide for poly(dA) · poly(dT) is significantly lower than that found for poly(dA‐dT) · poly(dA‐dT), thus indicating an appreciable preference of the drug for the alternating copolymer.The inactivation of the template properties of the synthetic DNA polymers in the DNA polymerase system is consistent with their daunomycin binding ability. However, a lack of correlation was observed between the drug binding ability of different DNA polymers and the binding‐induced stabilisation of the double helix to heat denaturation.
The Steady‐State Kinetics of Yeast Phosphoglycerate Kinase
Authors: SCOPES Robert K. 
Published: March 2005

A re‐investigation of the kinetics of yeast phosphoglycerate kinase in the direction of 1,3‐bis‐phosphoglycerate formation has been carried out, covering a 1000‐fold range in substrate concentrations. A variety of improved spectrophotometric and fluorimetric assay procedures have been used.
Kinetic plots proved to be non‐linear for each variable substrate. A variety of checks have been carried out to show that this is not due to artifacts in the assay procedures or heterogeneity of the enzyme preparation.
The effects of a variety of salts on the activity of the enzyme have been examined. Most salts, especially those with multivalent anions, can cause activation of the enzyme, but inhibit at high concentration.
The salt effect is shown to be principally due to anions rather than cations, and not to ionic strength changes. Sulphate, as one of the most effective anions has been used in most comparisons.
Salt activation is steepest when the substrate concentrations are low; maximum activation has been about 5‐fold with 0.2 mM MgATP and 0.2 mM 3‐phosphoglycerate. Inhibition at the higher salt concentrations is strongest at the same substrate concentrations as when activation is steepest, indicating a link between the two effects.
The presence of 20 mM or more Na2SO4 converted non‐linear kinetic plots to linear ones. A study of the kinetics in the presence of 40 mM Na2SO4 was interpreted in terms of a random sequential binding mechanism, with sulphate acting as a competitive inhibitor.
Possible explanations for these anomalous results are discussed in terms of several mechanisms which have been shown to apply in other systems.
X‐Ray and Neutron Small‐Angle Scattering Studies of the Complex between Protein S1 and the 30‐S Ribosomal Subunit
Authors: LAUGHREA Michael , ENGELMAN Donald M. , MOORE Peter B. 
Published: March 2005

X‐ray and neutron solution scattering experiments have been done to investigate the influence of the binding of ribosomal protein S1 on the conformation of the 30‐S ribosomal subunit of Escherichia coli. The following conclusions were made.

The alterations (if any) in conformation of the non‐S1 parts of the 30‐S subunit induced by S1 binding are too small to be detected (< 0.1 nm change in radius of gyration).
The center of gravity of protein S1 bound to the 30‐S subunit is quite far from the center of gravity of the particle (≅ 7.5 nm).
The Redox Potential of Dithionite and SO−2 from Equilibrium Reactions with Flavodoxins, Methyl Viologen and Hydrogen plus Hydrogenase
Authors: MAYHEW Stephen G. 
Published: March 2005

It has been shown that redox equilibria can be formed between dithionite ion (plus SO−2) and (bi)sulphite, and the low‐potential electron carriers flavodoxin and methyl viologen. The equilibria were established either by treating the oxidized electron carriers with dithionite, or by treating flavodoxin hydroquinone or methyl viologen semiquinone with (bi)sulphite. Similar redox equilibria were established between dithionite/(bi)sulphite and hydrogen using catalytic amounts of hydrogenase in the presence of a low‐potential electron carrier. The effects of pH and temperature on the equilibria were determined.
The equilibria were analyzed to determine the redox potential of the dithionite/(bi)sulphite system. In accordance with the results of earlier kinetic studies, it was assumed that the reductant in dithionite solutions is the dissociation product SO−2. The calculated midpoint redox potential E' for the couple SO−2/HSO−3 at pH 7 and 25 °C was –0.66 V. The reductant is present largely as the dimer at concentrations of dithionite above about 10nM. Consequently, the midpoint potential, Em, of dithionite solutions becomes less negative as the concentration of dithionite is increased (ΔEm/Δlog S2O2−4= 29 mV). The theoretical potential of a solution of 1 M S2O2−4 and 2 M (bi)sulphite at pH 7 was calculated to be –0.386 V. This value is 59 mV more negative than that determined in 1911 by potentiometry, but considerably more positive than other values in the literature. The effects of pH on the equilibria showed that E' is controlled by the pK of (bi)sulphite at 6.9; the slope ΔE'/ΔpH was –59 mV below the pK and – 118 mV above the pK. The effects of temperature on the equilibria suggested that Em for dithionite changed by –1.6 mV/°C for a rise in temperature between 2 °C and 40 °C. If sodium dithionite is contaminated with small amounts of (bi)sulphite, its addition in large excess to a low potential electron carrier can cause oxidation of the carrier.
Investigation of the Interaction of Triethyltin with Rat Liver Mitochondria Using Binding Studies and Mössbauer Spectroscopy
Authors: FARROW Barry G. , DAWSON Alan P. 
Published: March 2005

The binding of triethyltin to rat liver mitochondria is measured and the results indicate the presence of a three component system. This involves triethyltin binding to two specific sites, one of high affinity, which is found to have a concentration similar to that found for the F1‐ATPase complex and one of lower affinity. The third component is attributed to partition of triethyltin into the mitochondrial membrane.
Non‐specific partition is shown by inhibition of triethyltin binding to the low affinity site with N‐ethylmaleimide and to the high affinity site by buffer pH values lower than pH 5.7. Combination of these two conditions leaves a Scatchard plot which can be interpreted as being due to triethyltin partitioned into the mitochondrial membrane.
The selective inhibition of triethyltin binding to the low affinity site by N‐ethylmaleimide preincubation indicates the presence of a thiol group in the low affinity site and the absence of N‐ethylmaleimide‐reactive thiol groups necessary for triethyltin binding in the high affinity site.
Mössbauer spectra of triethyltin bound to mitochondria also seem to be derived from the same three component system. N‐Ethylmaleimide preincubation assigns an absorption doublet to low affinity bound triethyltin and the concentration dependence of the absorption intensities assigns a second doublet to partitioned triethyltin.
Washing triethyltin pretreated mitochondria leaves an absorption doublet which is assigned to high affinity bound triethyltin. This assignment is confirmed by binding and ATPase activity studies before and after this washing procedure.
Triethyltin bound to the high and low affinity sites is found, by the ratio of the Mössbauer quadrupole splitting and the isomer shift obtained, to be four coordinate or possibly cis‐five coordinate.
The assignment of the absorption doublet, from triethyltin in the high affinity site environment, to a four coordinated triethyltin‐mitochondrial membrane complex is suggested by the investigation of some intramolecularly coordinated triorganotin compounds.
Study of Follitropin Receptors in Testis Using a Homologous System
Authors: MAGHUIN‐ROGISTER Guy , CLOSSET Jean , COMBARNOUS Yves , HENNEN Georges , DECHENNE Charles , KETELSLEGERS Jean‐Marie 
Published: March 2005

The properties of follitropin receptors in immature porcine testis were determined using highly purified porcine follitropin.

The characteristics of follitropin binding to a subcellular fraction rich in plasma membranes were studied using a 125I‐labelled follitropin with high specific activity (75–100 Ci/g) and high binding activity. The binding is dependent on time, temperature and pH.It is specific to follitropin as demonstrated by the very low binding activity of the follitropin α and β subunits and of the other glycoprotein hormones. Scatchard analysis of binding data indicated an equilibrium association constant of 2 × 1010 M−1 and a concentration of high affinity binding sites of 500 fmol/mg membrane proteins.
A sensitive radio‐ligand receptor assay was developed. Fifty percent inhibition of binding was obtained with as little as 2 ng of porcine follitropin. Ovine and bovine follitropins and pregnant mare serum gonadotropin gave binding inhibition curves parallel to that given by porcine follitropin. With equine and human follitropin, significantly different slopes were recorded.
Kinetics of dissociation of labelled follitropin from its testis receptors showed the presence of at least two compartments with fast and slow dissociation rate constants. The ratio between the sizes of the slow and fast compartments appeared dependent upon preincubation time.
A temporal correlation was observed between binding of follitropin to testis receptors and activation of membrane bound adenylate cyclase.
The Binding of Kirromycin to Elongation Factor Tu
Authors: PINGOUD Alfred , URBANKE Claus , WOLF Heinz , MAASS Guenter 
Published: March 2005

The influence of kirromycin on the elongation factor Tu (EF‐Tu) in its binary and ternary complexes was investigated. The equilibrium constant for the binding of the antibiotic to EF‐Tu · GDP and EF‐Tu · GTP was determined by circular dichroism titrations to be 4 × 106 M−1, and to EF‐Tu · GTP · aa‐tRNA by a combination of circular dichroism titrations and hydrolysis protection experiments to be 2 × 106 M−1.In the presence of kirromycin the binding of aminoacyl‐tRNAs to EF‐Tu · GTP is weakened by a factor of two. The antibiotic changes the conformation of the ternary complex in such a way that the aminoacyl moiety of the aminoacyl‐tRNA is more accessible to the non‐enzymatic hydrolysis. It is concluded that this structural alteration is responsible for the inhibitory action of the antibiotic.
Chorismate Mutase/Prephenate Dehydratase from Escherichia coli K12
Authors: GETHING Mary‐Jane , DAVIDSON Barrie E. 
Published: March 2005

The effects of phenylalanine, NaCl and pH on the conformation of chorismate mutase/prephenate dehydratase have been investigated, using measurements of far and near‐ultraviolet circular dichroic spectra and ultraviolet difference spectra. At pH 8.2 in 20 mM Tris‐Cl buffer the enzyme was found to contain 10–20% helix and 40–50%β‐structure. There was little or no change in these values on the addition of 1 mM phenylalanine (the allosteric effector) or 0.4 M NaCl or by decreasing the pH to 7.4.Both phenylalanine and NaCl caused significant changes in the conformation of the enzyme. The most prominent of these was the movement of a tryptophan residue into a more hydrophobic environment. There was also a slight perturbation of this tryptophan when the pH was decreased to 7.4. The conformational changes can explain sigmoidal kinetic behaviour observed previously [Gething et al. (1976) Eur. J. Biochem. 71, 317–325].
Evidence for Essential Histidine Residues in Tryptophanyl‐tRNA Synthetase
Authors: FAVOROVA Ol'ga O. , MADOYAN Igor A. , KISSELEV Lev L. 
Published: March 2005

Beef pancreas tryptophanyl‐tRNA synthetase (Mr 60000 × 2) reacts with diethyl pyrocarbonate at pH 6.0. A differential spectrum of modified versus native enzyme as well as the restoration of the initial spectrum after NH2OH treatment of the modified enzyme prove the formation of carbethoxy histidine residues. In 8 M urea, 22 histidine residues are carbethoxylated by diethyl pyrocarbonate in accord with the total histidine content of the enzyme. In the native enzyme only 4–5 histidine residues react at equal rate whereas the others are resistant to modification. The circular dichroism spectra of the native and modified protein are similar. The modified enzyme does not dissociate into subunits.After modification of accessible histidine residues the enzymatic activity sharply decreases both in the reaction of ATP‐[32P]pyrophosphate exchange and that of tRNA aminoacylation. The modified enzyme is almost completely reactivated after treatment with NH2OH. The rate of the enzyme inactivation in the aminoacylation reaction approaches quite closely that of carbethoxylation of the histidine residues; a complete loss of the ability of tryptophanyl‐tRNA synthetase to catalyze tryptophanyl‐tRNA synthesis is brought about by the modification of 2–3 histidine residues per dimer. The modified enzyme losses almost completely its ability to bind tryptophanyl‐tRNA or uncharged tRNA. A conclusion is made that the accessible histidine residues are involved directly or indirectly in the tRNA binding. The ATP‐[32P]pyrophosphate exchange is less affected with modification of the enzyme by diethyl pyrocarbonate than the tRNA acylation reaction. All substrates protect the tryptophanyl‐tRNA synthetase against inactivation with diethyl pyrocarbonate; the most effective protectors are tryptophan and tryptophan plus ATP‐Mg2+ for the exchange reaction. Modified peptide(s) was isolated by diagonal electrophoresis.
On the Activity of RNA Polymerase B in Lysates from Ehrlich Ascites Cells
Authors: DREYER Christine , HAUSEN Peter 
Published: March 2005

Transcription by endogenous RNA polymerase B in lysates of Ehrlich ascites cells was investigated. The enzyme exhibits two salt optima at 0.025 M and at 0.3 M (NH4)2SO4 respectively. Preincubation of the cells with the nucleoside analogue 5,6‐dichloro‐1‐β‐d‐ribofuranosylbenzimidazole results in an inactivation of the polymerase molecules active under condition of low salt. This indicates two functional states of the enzyme in vivo.Initiations of RNA chains by polymerase B do not occur in vitro as judged by the incorporation of [β‐32P]GTP. Thus the two functional states seem to be both elongating polymerase molecules.Polymerase B does not occur in the lysates in a state ready to initiate on an exogenous template, in contrast to polymerase A and C which do occur in free form. Pretreatment with dichlororibofuranosylbenzimidazole in vivo does not result in an accumulation of free polymerase B.
Effect of Cycloheximide on Maturation of Replicative Intermediates into High‐Molecular‐Weight DNA in Tetrahymena
Authors: WESTERGAARD Ole , MARCKER Kjeld A. , LEER Johan Chr. 
Published: March 2005

Replicative intermediates of discrete size (∼ 41 S) are observed in the eukaryotic organism Tetrahymena pyriformis, when the organism is grown under defined physiological conditions. The intermediates (believed to represent replicons) are synthesized and accumulated over longer periods of time (< 90 min), if the cells are treated with low concentrations of cycloheximide. Under these conditions the rate of total DNA synthesis is only slightly inhibited (< 15%), while maturation of intermediate DNA into high‐molecular‐weight DNA is completely blocked (> 98%). Cycloheximide appears to inhibit the maturation process more specifically than other protein synthesis inhibitors.Studies of the accumulated intermediates on alkaline buoyant density gradients demonstrate that initiation of new putative replicons occurs during treatment with cycloheximide.
Phosphorylation in vivo of Proteins Associated with Heterogenous Nuclear RNA in HeLa Cell Nuclei
Authors: BLANCHARD Jean‐Marie , BRUNEL Claude , JEANTEUR Philippe 
Published: March 2005

As a step towards understanding the role of nuclear ribonucleoprotein particles (hnRNA · proteins) in the nucleocytoplasmic transfer of heterogenous nuclear RNA sequences destined to become the cytoplasmic messenger RNA, the phosphorylation in vivo of the protein moiety of these hnRNA · proteins from HeLa cells has been studied.After exposure of HeLa cells to [32P]orthophosphate in vivo, purified hnRNA · proteins were found to contain radioactivity covalently bound to proteins in the form of serine and threonine phosphate esters. The association of these phosphoproteins with hnRNA was demonstrated by sedimentation analysis in sucrose gradients, isopycnic banding in cesium chloride density gradients and chromatography on oligo(dT)‐cellulose.The pattern of radioactive phosphorylated species has been analysed by electrophoresis on sodium dodecylsulphate/polyacrylamide slab gels followed by autoradiography. Although some hottrichloroacetic‐acid‐resistant 32P radioactivity was present throughout the gel, the highest amount of label was found associated with four discrete bands of molecular weights 28000, 30000, 37000 and 52000.Incorporation of 32P into hnRNA · proteins could be detected after labeling periods as short as 15min, the same species being labeled after 1 and 24‐h labeling times. Although all the above species appear to be present in hnRNA · proteins of different size classes, as resolved by sedimentation through sucrose gradients, their relative proportion is not reproducibly constant.Exposure of hnRNA · proteins to conditions of increasing ionic strength led to the ready dissociation of the 28000‐Mr species at variance with other labeled species which appeared to be more firmly bound to hnRNA than the bulk of proteins. Upon treatment with pancreatic and T1 ribonucleases, the 28000‐Mr species was also preferentially released while the 37000‐Mr one was the main phosphorylated species remaining associated with core structures containing an RNA component resistant to nucleases which is now under further characterization.
Streptomycin Uptake via an Inducible Polyamine Transport System in Escherichia coli
Authors: HÖLTJE Joachim‐Volker 
Published: March 2005

The rate of uptake of streptomycin by Escherichia coli K12 becomes greatly enhanced after 10–20 min of incubation. Maximal rate of streptomycin and of gentamicin uptake is achieved by preincubation of the cells at 37 °C for 20 min in the presence of 5 μg/ml of streptomycin, kanamycin, gentamicin or neomycin. Spectinomycin and kasugamycin failed to increase streptomycin uptake. Enhanced streptomycin uptake is absolutely energy‐dependent.The uptake of streptomycin by wild type E. coli is much repressed in the presence of glucose. A cya mutant of E. coli which cannot synthesize adenosine 3′:5′‐monophosphate (cyclic AMP) exhibits poor uptake of streptomycin. The addition of cyclic AMP however, induces the uptake of streptomycin in this mutant.Transport of spermidine is ten times increased in the presence of 5 μg/ml streptomycin. The uptake of putrescine is enhanced two‐fold under the same conditions. Increase of uptake is prevented by chloramphenicol.Streptomycin competitively inhibits the transport of spermidine and putrescine. Conversely, spermine, spermidine and putrescine inhibit the uptake of streptomycin.It is concluded from these results that interaction of streptomycin with the ribosomes results in the induction of a polyamine transport system which can be utilized by streptomycin.
Complex Formation between the Adenovirus Type 5 DNA‐Binding Protein and Single‐Stranded DNA
Authors: van der VLIET Peter C. , KEEGSTRA Wilko , JANSZ Hendrik S. 
Published: March 2005

The adenovirus‐type‐5‐coded single‐strand‐specific DNA‐binding protein was purified from infected human KB cells and characterized by sucrose gradient centrifugation and gel filtration. The protein has a sedimentation coefficient of 3.3 S and a diffusion coefficient of 4.4 × 10−7 cm/s, which corresponds to a molecular weight of 68000 for the native protein. This is close to the peptide molecular weight of 72000 and indicates that the protein is a monomer. The frictional coefficient ratio is 1.82 which suggests that the DNA‐binding protein is highly anisometric.The binding of the protein to various nucleic acids was studied by filtration through nitrocellulose filters and sucrose gradient centrifugation. The binding to single‐stranded DNA is a fast process which does not require bivalent cations or sulfhydryl groups and occurs over a broad pH range. The binding to synthetic polydeoxyribonucleotides is at least 10‐fold less efficient than that to natural single‐stranded DNA.At low protein‐to‐DNA ratios the binding to single‐stranded DNA is cooperative. A maximal protein to DNA ratio of 33 (w/w) can be obtained which corresponds to 1 protein molecule for about 7 nucleotides. Sedimentation and electron microscopic studies indicate that the DNA‐binding protein keeps single‐stranded DNA in an extended configuration. The saturated nucleoprotein complex is slightly more resistant to nuclease digestion than single‐stranded DNA.