bluuebunny/crossref_metadata_2025_split huggingface dataset:
Komplexbildung von Nucleinbasen mit Cu2+
Authors: Sigel H. 
Published: March 2005

By means of UV difference spectra, the acidity constants KHHL of the N‐1‐proton in the nucleotides inonine, guanosine, uridine, and thymidine 5′‐triphosphate were measured. The same deprotonation (KHCuHL) in the respective Cu2+ complexes was also determined. The measurement were carried out with and without (Table 1) addition of NaClO4 (0.1 M). It was found that the influence of the ionic strength is great, possibly due to the formation due to Na+ complexes.The difference, ΔpKA=pKHHL–pKHCuHL∽ 2.0, for the nucleotide systems is of the same order as the respective value 2.2 of the Cu2+ guanosine system. This indicates a strong interaction between Cu2+ and the nucleic base in the Cu2+‐nucleotide complexes. From experimental indices and the analogy to the Cu2+‐ATP complex, it is concluded that macrochelates are formed.The acidity constants determined by means of UV difference spectra were confirmed by potentiometric titrations (Table 2).The Cu2+‐nucleoside triphosphates are known to form stable complexes with 2,2′‐bipyridyl. In such ternary complexes, the nucleic base can no longer coordinate to the Cu2+. The relatively small ΔpKBipy/A values (Table 2) are in agreement with this.The hydrolytic tendencies of Cu2+ bound to the different nucleotides were also estimated (Table 2). The pKHCuL(H2O)x values show that the hydrolytic tendency of the Cu2+ complexes decreases in the order CTP > ATP > TTP > UTP > ITP ∽ GTP. The influence of the coordination number of the different ligands and also other problems concerning nucleotide complexes were discussed.
Acides ribonucléiques messagers de la glande thyroïde
Authors: CARTOUZOU G. , ATTALI J.‐C. , LISSITZKY S. 
Published: March 2005

Nuclear and cytoplasmic RNA from sheep thyroid slices in vitro labeled with [3H]uridine or [32P]phosphate from 5 min up to 9 h, have been obtained by different phenol extraction procedures. Early‐labeled RNA was only extracted by phenol containing 0.1 or 1% (w/v) sodium dodecylsulfate (SDS) at 0° or 60°, whereas stable nuclear or polysomal RNA was easily extracted by pherol at 0°.
Time course of incorporation of label into subcellular particles has shown the early labeling of nuclear RNA and the transfer of radioactivity into the cytoplasm.
Rapidly‐labeled RNA extracted from nuclei by phenol‐1% (w/v) SDS and analyzed by sedimentation was shown to sediment in three discrete classes 50–30S, 22 S and 13 S, different from ribosomal RNA. After 9 h of incubation the 22 S and 13 S labeled RNAs were still present in detectable amounts in the nuclei, indicating a somewhat low turnover rate for these components.
The kinetics of incorporation of label into polysomes was studied in thyroid slices incubated from 15 min up to 5 h with [3H]uridine or [32P]phosphate. RNA was dissociated from polysomes by dodecylsulfate treatment and centrifuged on sucrose gradient. As for nuclei, the rapidlylabeled RNAs sediment in discrete size classes, 22 S and 13 S. After a long incubation time (5 h) the label was predominant in 28 S and 18 S ribosomal RNA but 28 S RNA was not completely labeled. Up to 30 min labeling a high molecular weight, early‐labeled RNA was present in purified polysomes. Its radioactivity declines after 60 min labeling.
A part of the rapidly labeled polysomal RNA can be reversibly separated from rRNP by the action of EDTA (1.25 to 5 μ moles EDTA/mg RNP). It sediments in a broad zone (12–30 S) of high specific radioactivity. Increasing Mg++ concentration results in the reassociation of rapidly‐labeled RNA to ribosomal sub‐particles.
Buoyant densities of the polysomal RNP particles obtained by EDTA treatment of 32P‐labeled polysomes and stabilized by means of formaldehyde fixation, were determined by banding in CsCI gradients. The bulk of the stable material banded at characteristic densities (d= 1.55 and 1.48), but most of the rapidly‐labeled RNA banded at d= 1.38.

The specific radioactivity of this material was the same as that of the 12–30 S RNA isolated by sucrose gradient centrifugation from the same EDTA‐treated polysomes. Preliminary double labeling experiments and computation of the RNA to protein ratio calculated from the RNA and protein density values indicate this material to be early‐labeled nucleoprotein particles with a high protein content.
Synthesis of Lens Protein in vitro
Authors: BLOEMENDAL H. , ZWEERS A. , SCHOENMAKERS J. G. G. 
Published: March 2005

Poly U‐directed stimulation of preincubated calf lens ribosomes revealed that these particles from a mammalian source have a phenylalanine incorporation ability comparable to the Escherichia coli system. A search was made for optimal conditions.After variation of the magnesium concentration during preincubation two levels of maximal incorporation were observed. The region of highest relative stimulation did not coincide with these maxima. Preincubation under conditions of optimal amino acid incorporation yields 80S monomers and to a lesser extent 120S particles. No smaller units or breakdown products were observed in the analytical ultracentrifuge. Such particles could be demonstrated after prolonged dialysis of either polysomes or 80S monomers against 0.1 mM MgCI2. Their sedimentation coefficients were 50 S and 30 S respectively but not 60 and 40 S.
Etude de la méthionyl tRNA synthétase de Escherichia coli
Authors: CASSIO D. 
Published: March 2005

Treatment of partially purified methionyl tRNA synthetase from Escherichia coli B (E. coli B) with p‐hydroxymercuribenzoate (pHMB) leads to loss of the capacity to form methionly adenylate and methionyl tRNA. Inactivation of the capacity to activate methionine could not be reversed by the addition of GSH or 2‐mercaptoethanol.
The Presence of low concentrations of l‐methioninyl adenylate (a structural analog of methionyl adenylate) during pHMB‐treatment, results in complete and selective protection of the active site responsible for the activation of methionine. Under these conditions, the addition of 2‐mercaptoethanol leads to full recovery of the capacity to charge tRNA.
The enzyme which has lost the capacity to form methionyl tRNA as a result of pHMB‐treatment in the presence of methioninyl adenylate, has the same elution characteristics as untreated enzyme when examined by gel filtration through Sephadex G‐200. It is concluded that loss of the capacity to charge tRNA is not a consequence of dissociation of the enzyme into subunits.

These results provide a clear‐cut distinction between the active site responsible for the activation of the amino acid, and the site(s) involved in the formation of aminoacyl tRNA, in the case of methionyl tRNA synthetase.
Structural and Metabolic Heterogeneity of Rat Liver Glycerophosphatides
Authors: ARVIDSON G. A. E. 
Published: March 2005

A quantitative thin‐layer chromatographic method is described for the separation and isolation of subfractions with different degrees of unsaturation of phosphatidylcholines and phosphatidylethanolamines from rat liver. In fasting male and female rats the tetraunsaturated or arachidonoyl‐containing subfraction was found to be of quantitative dominance in both phosphatidylcholines and phosphatidylethanolamines, whereas the latter contained relatively more of the hexaunsaturated fraction than the phosphatidylcholines. All the monoenoic subfractions contained more palmitic acid than stearic acid. In both the phosphatidylcholines and the phosphatidylethanolamines more than 50% of the stearic acid in each phospholipid was recovered in the tetraenoic fraction in both male and female rats. Incorporation of 32P and [1,2‐14C2]ethanolamine was found to occur at a high rate in the hexaenoic phosphatidylethanolamines. Another characteristic finding was the comparatively low metabolic reactivity of the arachidonoyl fraction in both phosphatidylcholines and phosphatidylethanolamines.The results have tentatively been interpreted to reflect differences in substrate specificity of CDPcholine: 1,2‐diglyceride cholinephosphotransferase and CDPethanolamine: 1,2‐diglyceride ethanolaminephosphotransferase towards different molecular species of l‐1,2‐diacylglycerol. The differences observed between the hexa‐and tetraunsaturated subfractions may also be related to differences in the turnover rate between polyunsaturated fatty acids of the linoleic acid type and those belonging to the linolenic acid series.
Proton Translocation Coupled to ATP Hydrolysis in Rat Liver Mitochondria
Authors: MITCHELL P. , MOYLE J. 
Published: March 2005

Glass electrode measurements of the pH of anaerobic suspensions of rat liver mitochondria during the hydrolysis of ATP to ADP and Pi by the oligomycin‐sensitive ATPase show that almost 2H+ ions are translocated outwards through the M phase of the cristae membrane per ATP molecule hydrolysed. Adenylate kinase activity is minimised by the presence of EDTA, and valinomycin must be added to the normal mitochondrial suspensions to collapse the membrane potential and permit the outwardly translocated H+ ions to diffuse practically quantitatively to the glass electrode system.Classical inhibitor studies of the overall ATPase reaction catalysed by the mitochondria indicate that the specific ATP/ADP porter is not rate‐limiting under the conditions of these experiments; and it is also shown that the overall process of ATP entry and ADP + Pi exit through the porter systems in the M phase is not coupled to net proton translocation. It is concluded that the ATPase system responsible for or coupled to proton translocation has a proton translocation quotient (→H+/P) of 2, corresponding to that of the type II ATPase of the chemiosmotic hypothesis.
Regulation of the Enzymes of the β‐Ketoadipate Pathway in Moraxella calcoacetica
Authors: Cánovas J. L. , Johnson B. F. 
Published: March 2005

Mutants of Moraxella calcoacetica which synthesize constitutively β‐ketoadipate succinyl‐CoA transferase, but in which all other enzymes of the β‐ketoadipate pathway are still normally inducible, have been isolated by selection for the ability to grow with β‐ketoadipate. The constitutively synthesized transferase was identified as transferase III. This enzyme is induced in the wild type by adipate and its physiological function is probably adipate activation. The properties of these constitutive mutants confirm that the synthesis of transferase III is regulated independently of the synthesis of enzymes that mediate reactions of the β‐ketoadipate pathway.Two phenotypically different classes of mutants which synthesize transferase II constitutively have also been isolated by the same selective procedure. Both classes of mutants synthesize constitutively the five enzymes that convert catechol to β‐ketoadipyl‐CoA. Strict coordinacy between four of these enzymes (muconate lactonizing enzyme, muconolactone isomerase, β‐ketoadipate enol‐lactone hydrolase II and β‐ketoadipate succinyl‐CoA transferase II) is maintained in one class of mutants. However in the other class this coordinacy is destroyed and muconate lactonizing enzyme is synthesized in succinate‐grown cells at a relative level much lower than those of the other three enzymes. These findings are discussed and evaluated in terms of earlier evidence concerning the regulation of the synthesis of enzymes of the catechol branch by M. calcoacetica.
The Synthesis of One‐Carbon Units from CO2 in Clostridium kluyveri
Authors: Jungermann K. , Thauer R. K. , Decker K. 
Published: March 2005

Clostridium kluyveri was grown (a) on standard media, in which alternately [14C]bicarbonate, [U‐14C], [1‐14C], and 2‐14C]acetate plus ethanol were the labeled substrates, and (b) on standard media supplemented with either formate, glycine or serine, in which bicarbonate or formate were 14C‐labeled. Adenine, guanine and methionine were isolated from the cells and degraded.(a) Carbon atoms 2 and 8 of the purines were found to be synthesized from CO2 (about 70%) and from the methyl group (about 20%) and the carboxyl group (about 10%) of acetate. The S‐methyl group of methionine was shown to be derived almost equally from CO2 (about 50%) and from the two‐carbon substrates, the methyl group of acetate accounting for about 35% and the carboxyl group for about 15%.(b) The participation of CO2 in the synthesis of the S‐methyl group of methionine remained unaffected (about 50%) in the presence of serine, while it was decreased to below 5% in the presence of formate and increased to over 80% in the presence of glycine. The participation of formate was in the order of 55%.These data demonstrate that (1) two major competitive pathways for the synthesis of onecarbon units exist in Clostridium kluyveri: C1‐units are formed predominantly by a reduction of CO2 and to a minor degree by a cleavage of serine, the usual pathway (C3‐serine = C2‐acetate) and (2) the relative rates of the two pathways are controlled by the glycine requirement of the growing organism.
Interactions between Neutral and Dibasic Amino Acids for Uptake by the Rat Intestine
Authors: Robinson J. W. L. 
Published: March 2005

Kinetic studies on the interaction between neutral and dibasic amino acids for absorption by the rat intestine have been performed. l‐Arginine uptake is stimulated by l‐citrulline, the kinetics formally revealing “uncompetitive” stimulation. l‐Isoleucine inhibits l‐arginine uptake “uncompetitively”. In the case of l‐methionine and l‐leucine, stimulation occurs at low concentrations of the neutral amino acid, but inhibition occurs at higher concentrations. The double reciprocal plots bisect in the first quadrant. Arginine inhibits the uptake of both methionine and citrulline, the kinetics being consistent with partially competitive inhibition. These kinetic situations were observed by means of steady‐state observations, but appear to be paralleled when initial velocities are determined.Efflux of arginine from a preloaded tissue is stimulated by external neutral amino acids, though not to such an extent as by arginine itself. Similarly, the efflux of neutral amino acids is stimulated to a lesser extent by external arginine than by the same or different external neutral amino acids.Intracellular methionine inhibits arginine efflux, and the efflux coefficient for arginine, determined from isotopic exchange experiments, is depressed by methionine. These results suggest that the stimulation of net arginine uptake by methionine may be explained in terms of an inhibition of arginine efflux. Intracellular citrulline and isoleucine have no effect on arginine efflux, however.Preloading with neutral amino acids produces the same effect on arginine uptake as the presence of these amino acids in the incubation medium. Although preloading with unlabelled arginine has no influence on the subsequent uptake of labelled arginine, it does tend to neutralise the isoleucine‐inhibition or the citrulline‐stimulation of arginine uptake. Methionine stimulation is, on the other hand, not affected by this preloading. This and other results suggest an intracellular locus of action of citrulline and isoleucine. Indeed these two amino acids seem to act in a similar manner but with an opposite effect on arginine uptake.
Guanosine Triphosphate Dependent Enzymic Binding of Aminoacyl Transfer Ribonucleic Acid to Yeast Ribosomes
Authors: Ayuso M. S. , Heredia C. F. 
Published: March 2005

The binding of phenylalanyl‐tRNA to purified yeast ribosomes at low magnesium concentrations is dependent upon one of the yeast supernatant transfer factors (factor A). This reaction requires GTP. The GTP analogue guanylyl methylene diphosphonate cannot be substituted for GTP but it acts as a good inhibitor. After incubation of the ribosomes with phenylalanyl‐tRNA, factor A and GTP practically no compound other than phenylalanyl‐tRNA itself is found linked to the ribosomes.In addition to this enzymic type of binding, phenylalanyl‐tRNA is also bound to yeast ribosomes in the absence of factor A and GTP provided that the magnesium concentration is increased to about 20 mM. This non‐enzymic binding is effectively inhibited by deacylated tRNA and this inhibition is prevented if factor A and GTP are included in the incubation mixtures.The binding of N‐acetylphenylalanyl‐tRNA to yeast ribosomes also requires GTP, and is dependent upon factor(s) other than the amino acid transfer factors, which are bound to the ribosomes.
Zusammensetzung der Phospholipide und deren Fettsäuremuster in Kaninchenerythrocyten und ghosts
Authors: Gercken G. , Brockmann U. 
Published: March 2005

The phospholipids of rabbit erythrocytes were separated by thin‐layer chromatography on silica gel HR and determined quantitatively by the molybdenum blue method. Prior to thinlayer chromatography phosphatidic acid and phosphatidyl inositol were concentrated by column chromatography.
The total phospholipid contents were 313 ± 27 amoles1 per erythrocyte and 289 ± 13 amoles per ghost. The proportions of the individual phospholipids of erythrocytes in moles % were 36.3 of phosphatidyl choline, 32.6 of phosphatidyl ethanolamine, 15.7 of sphingomyelin, 12.4 of phosphatidyl serine, 1.6 of phosphatidic acid, 1.3 of phosphatidyl inositol, and < 0.1 of lysophosphatidyl choline. The same distribution was obtained in ghosts with the exception of a significantly higher concentration of sphingomyelin.
The fatty acid composition of phosphatidyl choline, phosphatidyl ethanolamine, phosphatidyl serine, ceramide trisaccharide, and sphingomyelin was analysed by gas‐liquid chromatography after the lipids had been fractionated by preparative thin‐layer chromatography.
For identification of the fatty acids, their methylesters were isolated on silica gel G‐silver nitrate plates according to degree of saturation.
Typical of sphingolipids were the low concentration of linoleic acid, the absence of arachidonic acid, the presence of lignoceric acid and the very high concentration of nervonic acid.
Phosphatidyl choline and phosphatidyl ethanolamine were hydrolysed by phospholipase A (ophiophagus hannah) in order to analyse the fatty acid distribution of the two positions. In position 1 of phosphatidyl choline and phosphatidyl ethanolamine the saturated fatty acids amounted to 83.6% and 65.6%, respectively. Unsaturated fatty acids, especially oleic acid and linoleic acid, also occurred in position 1. These acids were the main components in position 2.
Structure‐Function Relationships for some Metalloalkaline Phosphatases of E. coli
Authors: Lazdunski C. , Petitclerc C. , Lazdunski M. 
Published: March 2005

The alkaline phosphatase of Escherichia coli is a zinc metalloprotein with a metal to enzyme ratio of 4. Inorganic phosphate protects the four zinc atoms against removal by EDTA. Cadmium and manganese can replace zinc on its four binding sites. Samples of phosphatase with different proportions of zinc and cadmium or of zinc and manganese have been prepared and tested for their enzymatic activity. These results together with others suggest a model which involves two functional and two structural zinc atoms. The zinc atoms necessary for the activity would have different apparent binding constants for the apoenzyme. This conclusion is in agreement with Cohen and Wilson's scheme postulated in 1966.Dissociation constants for Me2+‐apophosphatase complexes were determined (Me2+= Zn2+, Ni2+, Cd2+, Co2+, Mn2+). These constants follow the order Ni2+ > Co2+ > Zn2+ > Cd2+= Mn2+. The optical rotatory dispersion spectra of these different metallophosphatases are very similar between 25° and 55° suggesting little change in protein structure. However, only Co2+ and Zn2+ alkaline phosphatases are active enzymes.
Microsomal Marker Enzymes and their Limitations in Distinguishing the Outer Membrane of Rat Liver Mitochondria from the Microsomes
Authors: Brunner G. , Bygrave F. L. 
Published: March 2005

Repeated washing of rat liver mitochondria with sucrose medium removes all of the cyto‐chrome P‐450 while low but significant amounts of glucose 6‐phosphatase and NADPH cyto‐chrome c reductase are still retained. At the same time monoamine oxidase and succinate cyto‐chrome c reductase activities increase slightly. Cytochrome P‐450 and glucose 6‐phosphatase are completely retained by the microsomes even after five washings with sucrose medium.The specific activities of glucose 6‐phosphatase and of NADPH cytochrome c reductase in the outer membrane of rat liver mitochondria subfractionated by “swelling‐shrinking” are about 4‐ to 8‐fold greater than in the intact mitochondria and can range from about 20–60% of the value found for microsomes. The cytochrome P‐450 content of the outer membrane fraction is less than 3 % of that of the microsomes.From a consideration of these data it is concluded that NADPH cytochrome c reductase and glucose 6‐phosphatase are intimately associated with the outer membrane of rat liver mitochondria whereas cytochrome P‐450 is not. The limitations of these enzymes as microsomal “markers” is discussed.
Pyruvate Dehydrogenase, Substrate Specificity and Product Inhibition
Authors: Bremmer J. 
Published: March 2005

Studies with intact mitochondria and with soluble pyruvate dehydrogenase indicate that pyruvate and α‐ketobutyrate are oxidized by the same enzyme (pyruvate dehydrogenase), while α‐ketovalerate is oxidized by a different enzyme.
Pyruvate and α‐ketobutyrate have about the same affinity for the enzyme, but pyruvate is oxidized at a much higher rate.
Acetyl‐CoA and propionyl‐CoA both behave as competitive inhibitors to CoA. The Ki for both is slightly higher than the Km for CoA. Accordingly the enzyme is only moderately inhibited by a high acetyl‐CoA/CoA ratio.
NADH behaves mainly as a competitive inhibitor to NAD. The Ki is significantly lower than the Km for NAD. Accordingly the enzyme is strongly inhibited by a high NADH/NAD ratio.
The significance of these properties of the enzyme for the regulation of the activity of pyruvate dehydrogenase in vivo is discussed.
An Analysis of the Polyoma Virus DNA‐RNA Hybridization Reaction in vitro
Authors: Hudson J. B. 
Published: March 2005

A study was made of the parameters involved in the hybridization reaction between Polyoma virus DNA and its complementary RNA, with a view to establishing conditions suitable for routine measurement of the viral specific RNA synthesized in vivo. In general the reaction obeyed the same principles as those determined for the Escherichia coli DNA‐RNA system. An interesting result was the finding that the neutralized “random cyclic‐coil” form of Polyoma DNA had as good a hybridization capacity as the heat‐denatured naturally occurring double‐stranded circular form, under certain conditions; an observation which has important relevance to the structure of the cyclic‐coil DNA. An analysis was also made of the “competition hybridization” reaction, leading to the development of conditions minimizing the non‐specific effect displayed by various types of RNA.Under the conditions derived for optimal assay, the level of viral RNA synthesis was found to be several times greater than previously reported values.
Fractionation in a Density Gradient of some Glycoproteins from Human Ovarian Cyst Fluids
Authors: Dunstone J. R. 
Published: March 2005

Equilibrium density gradient sedimentation in caesium chloride has been used as an effective method of separating the components of sulphite‐reduced sparingly soluble glycoproteins from two human ovarian cyst fluids. An almost quantitative recovery of the carbohydrate and peptide constituents has been obtained. Some fractions had overall chemical compositions similar to those isolated previously by Dunstone and Morgan in 1965, while others contained much greater proportions of amino acids, especially of aspartic acid, glutamic acid and cystine.The results suggest that the sparingly soluble glycoproteins are large macromolecules containing serologically active carbohydrate chains, bound to specific peptide fragments which form an integral part of larger macromolecules, the insolubility of which is due to the presence of –S‐S‐bonds.The experiments emphasize the value of equilibrium density gradient sedimentation in the study of complex proteinpolysaccharide systems.