bluuebunny/crossref_metadata_2025_split huggingface dataset:
On the Morphology of Phytohemagglutinin from Phaseolus vulgaris Seeds
Authors: Höglund Stefan , Dahlgren Karin 
Published: March 2005

A purified sample of phytohemagglutinin molecules from Phaseolus vulgaris seeds was shown to be homogenous by analytical ultracentrifugation, electrophoresis and by electron microscopy. A test for the agglutination capacity of phytohemagglutinin a” in the contrast agent used for electron microscopy indicated that the contrast agent did not interact with the phytohemagglutinin a” molecule to destroy its biological activity.The phytohemagglutinin a” molecules were distributed uniformly in the electron microscopic specimen, where they appeared as small light dots against the background. The general shape of the projection of the molecule was almost rectangular with the minimum mean dimensions: 77 Å X 43 Å. Often it was also observed that phytohemagglutinin a” seemed to be irregularly constructed, and that it might consist of two chain units, joined lengthwise. An approximation of the molecular weight of phytohemagglutinin a” from its size and partial specific volume gave a value just below 100000; this value is close to the molecular weight of the monomer of phytohemagglutinin a” of 80000 daltons which was obtained from the chemical composition.
Specific Staining for Electron Microscopy of Murein Sacculi of Bacterial Cell Walls
Authors: Formanek Helmut , Fobmanek Sibylle 
Published: March 2005

The murein sacculi of the cell walls of Spirillum serpens consist of a two‐dimensional network of polysaccharide chains covalently connected with oligopeptides. Each oligopeptide in the murein sacculi is a stainable locus which reacts with one heavy atom compound by addition of a stoichiometric amount of the staining agent to a sacculi suspension.The single free amino groups of the oligopeptides are specifically stained by stoichiometric amounts of the heavy atom compounds 3‐hydroxy‐mercuri‐4‐hydroxy‐5‐carboxy‐benzenesulfonic acid, anhydro‐3‐hydroxymercuri‐4‐toluene‐sulfonic acid, Zeise's salt, platinum blue and tri‐bromoacetaldehyde.In a two‐step staining reaction sulfur compounds are bound to the free amino‐ or carboxyl groups of the oligopeptides by reaction with propylenesulfide, N‐acetyl‐DL‐homocysteine‐thio‐lactone and S‐acetylmercaptosuccinic anhydride. These sulfur atoms then react specifically with stoichiometric amounts of 2,4,6‐tris‐acetoxymercuri‐3‐acetaminotoluene. Stained and unstained murein sacculi are observed by high resolution electron microscopy. Unstained murein sacculi can hardly the recognized in bright field and do not expose any structural details. Sacculi specifically stained show distinct contrast and reveal a network pattern in which the stain is localized as discrete particles of about 5 Å diameter in periods of about 10 Å.
Chemical Characterization of Tetanus Toxin and Toxoid
Authors: Bizzini Bernard , Blass Judith , Turpin André , Raynaud Marcel 
Published: March 2005

The amino acid analysis of tetanus toxin as purified from bacterial extracts or from culture filtrates of Plectridium tetani, was carried out. The results that have been obtained support the assumption of the identity of both types of toxin.Tetanus toxoid amino acid composition was comparatively determined. Significant decreases in the amounts of some amino acids were found. These deficits are accounted for within the reactions with formaldehyde which lead to the formation of Mannich‐like derivatives.The presence of 6 free SH‐groups on the molecule of toxin could be substantiated. Two of these groups are difficult to reveal. The existence of two disulfide bridges was established. These results are discussed, in particular, with regard to the expression of the toxicity.The presence of only one N‐terminal amino acid residue could be revealed.
Neutral Steroids in Urine during Pregnancy
Authors: Jänne Olli A. , Vihko Reijo K. 
Published: March 2005

Three 15‐hydroxylated C21 steroids, 3β,15α‐dihydroxy‐5α‐pregnan‐20‐one, 3β,15α‐dihydroxy‐5β‐pregnan‐20‐one and 5‐pregnene‐3β,15β,17α,20α‐tetrol, were identified and measured in late‐pregnancy urine by gas‐liquid chromatography and gas chromatography‐mass spectrometry. All these steroids were present in urine as monosulphates, as evidenced by their chromatographic properties on Sephadex LH‐20 chromatography and by their cleavage by solvolysis. Quantitatively, 5‐pregnene‐3β,15β,17β,20α‐tetrol was the most abundant compound among these 15‐hydroxylated C21 steroids in the urine pool studied.The precursors and origin of the steroids identified are discussed.
Quantitative Correlation between the Distribution of Anions and the pH Difference across the Mitochondrial Membrane
Authors: Palmieri Ferdinando , Quagliariello Ernesto , Klingenberg Martin 
Published: March 2005

The factors regulating the distribution of anions between the inner and outer mitochondrial phase have been investigated in the presence of respiratory inhibitors and oligomycin.

As the external pH is decreased, the ratio of the intra‐ to extramitochondrial concentration of added anions increases with acetate, malate and phosphate, and with citrate in the presence of malate. In contrast, the uptake of thiocyanate does not change significantly with the pH.
In the pH range used, the log of the anion distribution ratio is proportional to the pH difference across the mitochondrial membrane for acetate, malate and phosphate.
The anion distribution ratio is highly sensitive to pH at low external concentration and becomes largely independent of pH at high external concentration.
In double reciprocal plots of intra‐ against extramitochondrial concentration, it was found that the capacity (Am) for the uptake of phosphate, acetate, malate and succinate is independent of pH. The slopes (A1/2/Am) increase with pH due to the increase of the external concentration required for half maximal uptake.
The distribution ratio of phosphate and malate is proportional to the relative “free capacity” (α) at all the external pH tested. The dependence of the distribution ratio on the external pH is maximal for α= 1 and nil for α= 0.
The following relation holds for the dependence of the slope A1/2/Am on Δ pH: log Am/A1/2=a+bΔ pH. This relation was used to obtain the equation describing the dependence of the anion distribution on the Δ pH and on the capacity.
It is concluded that the distribution of anions, such as acetate, phosphate and Krebs cycle intermediates, at equilibrium can be explained essentially as a function of two parameters: (a) the pH difference across the mitochondrial membrane (b) the capacity of the mitochondria for anions.
The Amino Acid Sequence of Neurotoxin I of Androctonus australis Hector
Authors: Rochat Hervé , Rochat Catherine , Miranda François , Lissitzky Serge , Edman Pehr 
Published: March 2005

The complete amino acid sequence of neurotoxin I of the North African scorpion Androctonus australis Hector collected in Algeria (area of Chellala) is described. S‐Methylation of cysteine residues formed by reduction of disulphide bonds was found useful for their identification in sequence determination using the phenylisothiocyanate method.From the N‐terminal sequence (38 residues) established in the protein sequenator, and from the sequence of chymotryptic peptides the complete sequence of the protein has been deduced. The molecule consists of a single polypeptide chain cross‐linked by four disulphide bridges. An isoneurotoxin (I′) has been found in the venom of A. australis collected in Tunisia (area of Tozeur). It differs from neurotoxin I in the replacement of the valine residue in position 17 by an isoleucine residue.
The Activation of Mutant β‐Galactosidase by Specific Antibodies
Authors: Melchers Fritz , Messer Walter 
Published: March 2005

An immunosorbent was prepared of antiserum against purified wild type β‐galactosidase coupled to Sepharose. Extracts of wild type Escherichia coli K12 3300 cells and of eleven mutant cells producing proteins activatable by specific antibodies were adsorbed to this immunosorbent. The adsorbed proteins could be eluted from the immunosorbent by 8 M urea. By co‐electrophoresis in polyacrylamide gels containing 8 M urea, 0.01 M mercaptoethanol and 1% sodium dodecylsulfate in a buffer at pH 7.2 subunits of all the eleven [3]leucine‐labelled mutant proteins were found to have the same size (molecular weight 130000) within the accuracy of the method (± 15000) as subunits of [14C]leucine‐labelled wild type enzyme. After renaturation from urea solutions the molecular weights of the native forms of the eleven [3H]leucine labelled mutant proteins were compared to that of the 14C‐labelled native wild type enzyme by co‐sedimentation in sucrose gradients. The native proteins were found to be identical in size as well (molecular weights 520000—540000). Thus, activation of these eleven mutant proteins by specific antibody occurs at the four subunit structure level.Such purification on immunosorbents represents a rapid, easy and generally applicable method to be used in systems with immunologically cross‐reacting mutant proteins, where the wild type protein can be obtained in sufficiently pure form.
Energy‐Dependent Quenching of Chlorophyll a Fluorescence in Isolated Chloroplasts
Authors: Wraight Colin A. , Crofts Antony R. 
Published: March 2005

A light‐induced, energy dependent quenching of chlorophyll a fluorescence in the presence of 2,3,5,6‐tetramethyl‐p‐phenylene diamine (diamino‐durene) and 3‐(3,4‐dichlorophenyl)‐1,1‐dimethylurea is described for isolated chloroplasts, and is related to the formation of the high‐energy state of photophosphorylation.
The quenching is distinguished from the normal type, operative through the redox state of the Photosystem II quencher, by its light‐intensity dependence, reversibility in the dark and sensitivity to uncoupling agents.
The sensitivity of the quenching to uncouplers and ionophorous antibiotics closely parallels that of photophosphorylation.
A similar quenching is described for other electron transport systems in the absence of dichlorophenyl‐dimethylurea, when it is distinguished from Q‐quenching on a time‐basis.
The pH‐optimum for the uncoupler‐reversible quenching is about pH 8.0 for all electron flow systems used. This apparent discrepancy with the pH optimum for XE‐formation is discussed.
Two other types of slow quenching seen during the Hill reaction with ferricyanide are described. One is found to be uncoupler enhanced and is related to acceleration of electron transport giving rise to a lower level of reduced Q in the steady state; the other is an irreversible loss of total fluorescence yield, which is most pronounced in coupled chloroplasts, and it is suggested that it is due to photochemical destruction of bulk pigment molecules via a side reaction of the excited state of chlorophyll.
Turnover Rates and Intracellular Pool Size Distribution of Citrate Cycle Intermediates in Normal, Diabetic and Fat‐Fed Rats Estimated by Computer Analysis from Specific Activity Decay Data of 14C‐Labeled Citrate Cycle Acids
Authors: Sauer Frank , Erfle James D. , Binns Michael R. 
Published: March 2005

Specific radioactivity curves of citrate, malate, glutamate and aspartate were measured in vivo in liver from normal, fat‐fed and diabetic rats with [1‐14C]acetate as precursor at time intervals of 0.1 to 20 min. A mathematical model of 14 pools and 40 interconnecting flux rates been constructed and by computer analysis on the Univac 1108 flux rates and intra‐ and extra‐mitochondrial metabolite pool distribution calculated from the measured specific radioactivity curves. Flux rates and pool distributions were varied by using an automatic minimization routine to minimize the difference between observed and theoretical values until an acceptable fit of computer generated specific radioactivity time curves to experimentally observed specific radioactivities was obtained. Steady state kinetics were assumed, i.e. the model was fixed with no change in pool sizes over the measured time interval. The main conclusions were:

A state of non‐equilibration of isotope exists for the acetyl‐CoA and acetylcarnitine, citrate, malate, aspartate, glutamate and α‐ketoglutarate pools distributed between cytoplasmic and mitochondrial compartments.
Cytoplasmic pools of citrate, acetyl‐CoA and acetylcarnitine account for the slow rate of radioactivity decay observed for these metabolites in vivo.
Key flux rates were acetate entry into the citrate cycle of 0.36, 0.28 and 1.05 μmoles × min−1× g liver−1 in normal, fat‐fed and diabetic rats. Citrate turnover rates were 1.21, 0.37 and 1.94 μmoles × min−1× g liver−1 for normal, fat‐fed and diabetic rats. The acetate pool was 0.23 μmoles/g liver in normal, 0.44 μmoles/g liver in fat‐fed and 0.38 μmoles/g liver in diabetic rats. Metabolite pool distribution was greater in the extra‐ than intramitochondrial space: the shift was most marked in the diabetic rat where the intra‐ to extramitochondrial malate distribution was in the ratio of 1:21.5.
The Cleavage of Prolyl Peptides by Kidney Peptidases
Authors: Dehm Peter , Nordwig Arnold 
Published: March 2005

Peptides of the X‐Pro‐Y type were found to be readily cleaved by crude swine kidney homo‐genates. Using Gly‐Pro‐Hyp as a substrate, a peptidase releasing glycine was isolated and partially purified from the microsomal fraction of the homogenate. The purification steps used included treatment of the kidney microsomes with n‐butanol, and chromatography of the protein fraction thus released on DEAE‐cellulose as well as gel filtration on Sephadex G‐200. The increase in specific activity was approximately 250‐fold.The purified preparation consisted of two enzymatically active components exhibiting molecular weights of one million and two millions, respectively. There is evidence that these components are lipoproteins, i.e. membrane fragments of different size.The activity of the enzyme was optimal at pH 8 and was greatly increased in the presence of manganese ions and albumin. Since the peptidase required a prolyl residue in the second position of the substrate, the designation “X‐prolyl‐aminopeptidase” is suggested. Although the enzyme acted on substrates of different chain length, the reaction rates were greatest with peptides of the structure Gly‐Pro‐Y. Therefore, a role of X‐prolyl‐aminopeptidase in the degradation of collagen appears plausible.
Interactions entre l'oestradiol et des sites récepteurs utérins
Authors: Best‐Belpomme Martin , Friés Jeanne , Erdos Tamas 
Published: March 2005

The oestradiol‐binding protein of calf uterine cytosol forms a slowly reversible complex with oestradiol. Kinetic studies on the formation of this complex, carried out with crude or partially purified uterine cytosol preparations, indicate a reaction of the second order. The rate constant shows an inverse relationship with protein concentration. Dissociation of the complex appears to occur in two stages, both following first order reaction rate kinetics. Affinity constants calculated from association‐dissociation kinetics, decrease with increasing temperature. The affinity constant measured under equilibrium conditions shows an inverse relationship with protein concentration, and does not appear to vary at temperatures between 0° and 18°. The treatment of the data from binding studies at equilibrium suggests a cooperative effect.These findings indicate that the interaction between the oestradiol‐binding protein, and its ligand cannot be described by simple reaction schemes; work with purified material, now under progress, could simplify interpretation by eliminating ambiguities due to eventual specific or unspecific protein‐protein interactions in our present impure system.
Structural Comparison of Ovalbumins from Nine Different Species
Authors: Fothebgill Linda A. , Fothergill John E. 
Published: March 2005

The structures of nine avian ovalbumins were compared by starch gel electrophoresis, ultracentrifugation, tryptic peptide mobilities, and amino acid composition.The results showed that the ovalbumins belonged to two structurally different groups which corresponded to the two taxonomic orders studied. Relationships within the orders were also indicated.
Direct Spectrophotometric Determination of the Structure of Dinucleoside Monophosphates
Authors: Feulicoli Romano A. , Cervelli Stefano , Ipata Piero L. , Rossi Carlo A. 
Published: March 2005

The identification of the nucleoside liberated by the action of spleen or snake venom phos‐phodiesterase on dephosphorylated dinucleosidemonophosphates directly reveals whether the nucleoside participates in the phosphodiester bond with its 3′‐ or 5′‐hydroxyl. We report a sensitive direct Spectrophotometric procedure for the detection and estimation of the liberated nucleoside, based on the following specific enzyme systems: guanosine phosphorylase, guanase and xanthine oxidase for guanosine at 290 nm, adenosine deaminase for adenosine at 265 run; cytidine deaminase for cytidine at 280 nm and uridine phosphorylase for uridine at 282 nm.The structure of dinucleotides is determined in single experiments: the liberated nucleoside bearing the free 3′‐ or 5′‐hydroxyl is identified and quantitatively measured by its response to one of the ancillary systems; the remaining 5′‐ or 3′‐mononucleotide is identified as nucleoside after addition of phosphomonoesterase.The use of T1 and pancreatic ribonucleases, along with the proposed principles and techniques, in studying the sequence of trinucleotides and tetranucleotides is discussed in the Appendix.
Regulation of Pyruvate Kinase by Fructose 1,6‐diphosphate in Saccharomyces cerevisiae
Authors: Barwell Clive J. , Woodward Brian , Brunt Rodney V. 
Published: March 2005

The concentrations of glycolysis intermediates, citric acid and adenine nucleotides were measured in resting cells of Saccharomyces cerevisiae during aerobic glucose metabolism. Addition of glucose resulted in a decrease in the intracellular content of phosphoglyceric acids and phosphoenolpyruvate. As extracellular glucose became depleted the level of these intermediates increased. Thus changes in their intracellular content were regulated by factors other than the supply of extracellular substrate.Cross‐over behaviour of phosphoenolpyruvate and pryruvic acid together with the inverse relationship which existed between the concentration of phosphoenolpyruvate and the rate of the pyruvate kinase reaction, indicated that regulatory changes in the activity of pyruvate kinase occurred. These activity changes were compared with those of the intracellular content of reported in vitro effectors of the enzyme: fructose 1,6‐diphosphate, citric acid, ATP and ADP. ATP and ADP did not appear to be involved in regulating changes in the activity of the enzyme. Although citric acid may have contributed towards some of the changes in enzyme activity, fructose 1,6‐diphosphate appeared to be the primary regulator, under the conditions of these experiments.Comparison of the results with the properties reported for the isolated enzyme, suggested that, in vivo, fructose 1,6‐diphosphate affects the affinity of pyruvate kinase for phosphoenolpyruvate. This, in turn, appears to regulate the intracellular content of phosphoenolpyruvate and phosphoglyceric acids. The significance of the results is discussed in relation to both glycolysis and gluconeogenesis.
Reinigung und Kristallisation der NAD‐Glykohydrolase aus C‐Streptokokken
Authors: Fehrenbach Franz J. 
Published: March 2005

The purification and concentration of NAD‐glycohydrolase from group‐C streptococci was achieved by zone precipitation, adsorption chromatography and re‐chromatography on hydroxylapatite. Subsequently, the enzyme could be isolated as a microcristalline product showing double refraction in the polarization microscope.Analytical gel permeation chromatography, analytical gradient elution chromatography on hydroxylapatite, density gradient centrifugation, sodium dodecylsulfate–urea electrophoresis and agar gel diffusion have been used to demonstrate the homogeneity of the purified enzyem. The polyacrylamide gel electrophoresis shows the enzyme as an uniformly migrating band. Furthermore the electrophoretic mobility of NAD‐glycohydrolase corresponded approximately to that of hemoglobin. This finding provides first evidence for the existence of identical subunits of the enzyme protein. Besides the demonstration of its catalytic function and antibody specificity the crystallized enzyme has been shown to possess hemolytic activity which could be activated by thiol groups. Though a variety of efficient purification procedures were employed they failed to remove Streptolysin‐O from the enzyme. Considering analytical data and criteria of homogeneity it may be assumed that a chemical correlation exists between the crystallized enzyme from group‐C streptococci and Streptolysin‐O previously described as two different substances.
Pyruvate Dehyrogenase Component Subunit Structure of the Escherichia coli K 12 Pyruvate Dehydrogenase Complex
Authors: Vogel Otto , Henning Ulf 
Published: March 2005

The pyruvate dehydrogenase component of the Escherichia coli pyruvate dehydrogenase complex is shown to be a dimeric protein consisting of identical or very nearly identical polypeptide chains with a molecular weight 90 000 to 110 000.In reasonable agreement with data obtained by Reed and collaborators the native enzyme was found to be a species with a sedimention coefficient s°20, w= 9.8 S, a diffusion coefficent D°20, w= 4.4 × 10−7 cm2/sec, and a molecular weight of 200 000 to 220 000. Amino acid analyses and analyses of tryptic peptides showed that the minimum chemical molecular weight is around 100 000. Serine was found as an N‐terminal residue but with poor yields. One carboxyl terminal sequence…Arg‐Leu‐Ala‐CO2H per 105 daltons protein was shown by carboxypeptidase degradation.The protein could be dissociated by a variety of methods into only one species of subunit. The molecular weight of this subunit was found, by dodecylsulfate polycrylamide‐gel electrophoresis and sedimentation equilibrium in various solvents, to be 90 000 to 110 000.
Interaction of Hemoglobin with Ions
Authors: Klinger Reinhard G. , Zahn Dieter P. , Brox Dieter H. , Frunder Horst E. 
Published: March 2005

Methods are described for the rapid determination of free ATP and ATP loosely‐ and firmlybound to oxygenated hemolobin using anion exchange resin and ultrafiltration. In hemolyzates of human erythrocytes under simulated in vivo conditions about 20% of total ATP is firmly‐bound to hemoglobin with an association constant of about 1 × 109 (M−1).Depending on the total ATP concentration used, equilibrium between free and firmly‐bound ATP is reached within 0.2 and 0.5 sec after ATP addition to “stripped” hemolyzates. In equilibrium the exchange between these two fractions is very slow, not reaching readioactive equilibrium until several days after labelling the free ATP fraction. On the other hand, exchange between free and loosely‐bound ATP is very fast.In addition to the known two binding sites for ATP in oxy‐ and deoxyhemoglobin there exists a third binding site which firmly binds ATP and other anions independet of the oxygenation state of the hemoglobin molecule.
Étude du transfert enzymatique de D‐[14C]mannose á partir de GDP‐[14C]mannose, sur un accepteur endogène dans les microsomes d'Aspergillus niger
Authors: Letoublon Robert , Richard Michel , Louisot Pierre , Got René 
Published: March 2005

A glycoprotein‐mannosyltransferase, utilsing an endogenous acceptor found in Aspergillus niger is herein characterized. Data presented indicate that among the subcellulr fractions, only microsomers are active in the mannose transfer reactions.The enzyme catalising the transfer requires Mn++ or Mg++ (1–2 mM) and has an optimum pH of 6.8 and an optimum temperature of 22°.It exibits a Km of 0.2 μM for GDP‐mannose. The transfer is inhibited by ATP, CTP and also bt GTP or its structural analogue β‐γ‐methlene‐guanosine triphosphate, but AMP or GDP have only a slight inhibitory action.Triton X‐100 causes of 2‐ to 3‐fold enhancement of incorporation of mannose when a suitable detergent: protein ratio is applied.The efficiency of this enzyme‐aceptor system is discussed.