bluuebunny/crossref_metadata_2025_split huggingface dataset:
Regulation der Biosynthese der aromatischen Aminosäuren in Claviceps paspali
Authors: Lingens F. , Goebel W. , Uesseler H. 
Published: March 2005

DAHP‐synthetase, chorismate mutase, prephenate dehydrogenase, prephenate dehydratase and anthranilate synthease were identified in extracts of Claviceps paspali mycelium. We were able to separate three DAHP‐synthetases, one sensitive to l‐phenylalanine, one sensitive to l‐tyrosine and one sensitive to l‐tryptophan. The l‐tryptophan sensitive isoenzyme provided the main part of the DAHP‐synthetase activity (about 60%). The inhibition due to l‐tryptophan was not removed by l‐tyrosine nor l‐phenylalanine. Chorismic acid did not inhibit these enzymes. Chorismate mutase was inhibited by l‐tyrosine and also by l‐phenylalanine, though these inhibitions were only detected after purification of the enzyme, and even then the highly purified enzyme (ca. 100‐fold) could not be separated into isoenzymes. The inhibition of the purified enzyme by l‐tyrosine and l‐phenylalanine could be removed entirely by l‐tryptophan.Prephenate dehydratease was inhibited only by l‐phenylalanine and it could not be activated by l‐tyrosine nor l‐tryptophan.Prephenate dehydrogenase showed an inhibiting effect by l‐tyrosin only after purification.Anthranilate synthetase was not inhibited by l‐tryptophan even after purification.Anthranilate synthetase was not inhibited by l‐tryptophan even after purification. With the simultaneous additions of l‐tyrosine, l‐phenylalanine and l‐tryptophan (in equal concentrations of 1 × 10−3 M) no repression effects could be produced.
The Solubilization of the Cytoplasmic Membrane of Bacillus subtilis by Sodium Dodecyl Sulphate
Authors: Bishop D. G. , Rutberg L. , Samuelsson B. 
Published: March 2005

Cytoplasmie membranes of Bacillus subtilis have been solubilized by brief exposure to the anionic detergent, sodium dodecyl sulphate. Complete solubilization was obtained when the relative proportation of detergent to membrane protein was approximately 1:2.The solubilized membrane preparation can be completely freed of detergent by gel filtration. The membrane components do not reaggregate, and membrane phospholipid remains associated with the protein during solubilization and subsequent removal of the detergent.Model experiments showing that enzymes can be selectively solubilized by varying the concentration of sodium dodecyl sulphate, indicate that the solubilization of the membrane preparation by the detergent is not a random disaggregation.
Spectrométrie de masse de glycolipides
Authors: Adam A. , Senn M. , Vilkas E. , Lederer E. 
Published: March 2005

This paper describes mass spectrometric studies of the peracetylated derivatives of synthetic and natural 6,6′‐diesters of trehalose containing acyl radicals varying from C22 to C90.The peracetylated synthetic 6,6′‐dieicosnoyl trehalose (I) gave a molecular ion at m/e 1238. The synthetic 2,3,4,2′,3′,4′‐hexaacetyl‐6,6′‐di(α‐eicosanyl‐β‐acetoxy tetracosanoyl)‐trehalose (IIb) showed no molecular ion but a peak (M‐60) at m/e 1910 (Fig. 1). The cleavage of the glycosidic bond leads to typical oxonium ions which are at m/e 611 (A) for (I) and at m/e 977 (B2) for (IIb).The three natural trehalose 6,6′‐diesters examined are the “cord factors” of a human virulent strain of Mycobacterium tuberculosis (strain Prévois), of a bovine attenuated strain (BCG) and of Mycobacterium butyricum.The mycolic acids isolated after saponification of these compounds have also been studied by mass spectrometry. Formulae III, IV, V and VI are proposed for their methyl esters.The molecular weights of the paracetylated “cord factors” are above 3000; no molecular ion could be detected. The oxonium ions, however, give significant peaks which are useful for structural determination. Oxonium ions for the glycolipid of the human virulent strain Prévois are found at m/e 1360–1528 (Fig.2). The peracetylated glycolipid of BCG contains two different series of homologous mycolic acids (acyl ions at m/e 1144–1284 and 1072–1184) and gives oxonium ions at m/e 1432–1572 for one series and at m/e 1360–1472 for the second one. The “cord factor” of M. butyricum gives homologous oxonium ions from m/e 1361 to 1445 corresponding to acyl ions from m/e 1073 to 1157.
Metabolism of trans‐3‐Hexadecenoic Acid by Chlorella vulgaris and by Lettuce Leaf
Authors: Bartels C. T. , James A. T. , Nichols B. W. 
Published: March 2005

The incubation of trans‐3‐hexadecenoic acid with cells of Chlorella vulgaris, and with lettuce leaf slices, resulted in reduction of the substrate to palmitic acid.Before this reduction was complete, the trans‐acid entered all the acyl lipid classes present in the cell.Consequently, the specific location of trans‐3‐hexadecenoic in the phosphatidyl glycerol fraction of photosynthetic tissues cannot be explained on the basis that lyso‐phosphatidyl glycerol is the only lipid capable of ‘accepting’ this acid. It is more likely that phosphatidyl glycerol or lyso‐phosphatidyl glycerol is a required substrate or cofactor for the synthesis of trans‐3‐hexadecenoic acid from palmitic acid.
A Method for Extracting Purified DNA or Protein‐DNA Complex from Escherichia coli
Authors: Favre J. , Pettijohn D. E. 
Published: March 2005

A new technique for the extraction of DNA or protein‐DNA complex from bacteria has been developed, using multiple extractions in the polyethyleneglycol‐dextran two‐phase system. The procedure has several advantages over the usual methods for DNA extraction: purified DNA contains less than 0.05% RNA or 1 μmole arginine per mmole DNA phosphate; denatured and partially denatured DNA fragments are excluded; DNA precipitation and ribonuclease treatments are unnecessary; the method is fast, less than 8 hours being required for the complete procedure. The purified protein‐DNA complex contains about 0.5 mg protein/mg DNA and significant amounts of RNA polymerase activity.A small amount of denatured DNA was separated from the double‐stranded DNA during the extraction. The size of this denaturated fraction depended upon the temperature and conditions of shear during lysis. Small amounts (0.2 to 3%) were found even at 30° under conditions where only inadvertant shears were introduced, while at higher temperatures with stirring the fraction was much larger. When purified T4 DNA was fragmented by shear to about the molecular weight of the final bacterial DNA fragments, denaturation of the T4 DNA was also detected. These results suggest that a shear‐induced denaturation of DNA can occur and that the fragments of denatured DNA found in bacterial lysates are produced at least in part by shear and may therefore have no direct biological significance.
Kinetics of Glycerol Kinases from Mammalian Liver and Candida mycoderma
Authors: Grunnet N. , Lundquist F. 
Published: March 2005

Glycerol kinase (ATP: glycerol phosphotransferase, EC 2.7.1.30) has been purified from rat, beef and human liver. Glycerol kinase from Candida mycoderma was also investigated.The kinetics with regard to glycerol are of the Michaelis‐type. The Lineweaver‐Burk plots with regard to ATP are non‐linear. The data obtained provide evidence for the ATP‐Mg2− complex as the true substrate and indicate that free ATP is a positive modifier for the enzymes.Inhibition experiments showed no effect of ethanol, acetaldehyde, acetate, d,l‐glycerol‐3‐phosphate, fructose‐1,6‐diphosphate or NADH upon the reaction velocities. ADP and AMP are strong inhibitors of the enzyme.The inhibition by AMP has been examined. It is concluded that AMP is a negative modifier for glycerol kinase. The inhibition by AMP is counteracted by free ATP. The interaction coefficient for AMP is about one.The kinetic properties of the glycerol kinases examined are to some extent similar to those of phosphoglycerate kinase. The possible physiological significance of the inhibition by AMP is discussed.
Specificity of the Constitutive Hexose Transport in Yeast
Authors: Heredia C. F. , Sols A. , Fuente G. Dela 
Published: March 2005

A method has been developed based on the osmotic sensitivity of protoplasts, that permits turbidimetrical estimation of the penetration of non‐metabolizable compounds into yeast protoplasts. Using this method a wide variety of sugars and related compounds have been tested as presumptive substrates for the constitutive hexose transport system in baker's yeast. An integration of the results obtained by this method with those obtained by other approaches has led to the establishment of the structural requirements that a compound has to meet in order to be transported. The basic structural requirement seems to be met by a pyranose ring for glucose and a furanose ring for fructose. With compounds that can be regarded as structurally related to d‐glucopyranose, there is a broad tolerance for modifications at carbons 1 and 2, and somewhat less so for modifications at carbon 3. Similar requirements, except for carbon 2, apply to compounds that can be regarded as structurally related to D‐fructofuranose.Physical diffusion of sugars, and related compounds, through the cell membrane of baker's yeast is very low, so that they only can enter into the cell at measurable rates by means of specific transport devices. The factor by which the entrance of glucose is increased over its physical diffusion, at concentrations in the millimolar range, is of the order of 106.
A Comprehensive Fractionation Procedure for Avian Erythrocyte Histones
Authors: Vidali G. , Neelin J. M. 
Published: March 2005

A fractionation procedure for avian erythrocyte histones was devised to give the six known, reproducible histone fractions from a single nuclear preparation, with limited manipulation and little wastage. The proposed method includes selective, two‐step acid extraction, followed by cation‐exchange chromatography and exclusion chromatography. The fractions were identified by amino acid analysis and by starch gel electrophoresis. The method is flexible in scale and permits a continuous balance of compositions and activities in metabolic experiments.Factors contributing to the relative extractability of histones from chromatin in acid were considered. The essential role of this step was the separation of erythrocyte‐specific, serine‐rich histone V, and arginine‐rich histones III, IV, which otherwise would be eluted together from Amberlite CG‐50 by guanidinium chloride. Two other arginine‐rich components, more easily eluted in chromatograms of other tissues, were absent from the usual extracts of mature erythrocytes. However, similar components were produced from histones III, IV by treatment with β‐mercaptoethanol.The conditions for good resolution of sub‐fractions of moderately lysine‐rich histones and of arginine‐rich histones by exclusion chromatography on Bio‐Gel P‐60 and P‐30 were examined. Reversible adsorption appeáred to be more significant than pore size of the gel; the sulphates of arginine‐rich histones were resolved better than the chlorides.
Thioredoxin: 4. Amino Acid Sequence of Peptide B
Authors: Holmgren A. 
Published: March 2005

Tryptic and chymotryptic digestion of peptide B, the N‐terminal fragment obtained by cyanogen bromide cleavage of thioredoxin, provided the information required to determine the contiguity of the peptic fragments studied previously. The complete sequence of the 37 residues of peptide B could thus be established. Peptide B contains the active center of thioredoxin. Its amino acid sequence was found to be: . This sequence supports earlier conclusions drawn from fluorescence studies of thioredoxin, which indicated a close relationship between one or both tryptophan residues and the disulfide bridge of the active site.Peptide maps of the tryptic digests of carboxymethylated thioredoxin and of the two cyanogen bromide fragments are described. Amide nitrogen analyses of thioredoxin and the two cyanogen bromide fragments demonstrate that all seven amide groups of thioredoxin are localized in the C‐terminal fragment (peptide A).
The Proflavin‐Induced Increase in the Catalytic Activity of Ficin
Authors: Hollaway M. R. 
Published: March 2005

The dye proflavin (3,6 diaminocaridine) binds to a single site on the ficin molecule (EC 3.4.4.12).
The value of the dissociation constant (Kp) for the enzyme‐dye complex was 3.5 ± 1.8 × 10−4 M measured spectrophotometrically and 1.55 ± 0.61 × 10−4 by equilibrium dialysis.
Interaction with proflavin increased the catalytic constant (kcat) for the ficin‐catalysed hydrolysis of benzoyl‐L‐arginine ethyl ester although the value of KM app was unaltered.
Dye‐binding did not significantly affect the value of kcat for the ficin‐catalysed hydrolysis of p‐nitrophenyl hippurate, a substrate for which acylation is not rate‐limiting in catalysis.
The dependence of initial velocity of benzoyl‐l‐arginine ethyl ester hydrolysis on dye concentration enabled the computation of a value of Kp= 1.61 × 10−4 M assuming a kinetic model involving formation of a ternary dye‐enzyme‐substrate complex in which substrate is more rapidly hydrolysed than in the binary enzyme‐substrate complex.
Dye‐binding leads to an enhanced nucleophilicity of the essential thiol group of ficin.
It is concluded that deacylation is not the rate‐limiting step in the ficin‐catalysed hydrolysis of benzoyl‐L‐arginine ethyl ester.
Possible explanations of the proflavin‐induced acceleration of benzoyl L‐arginie ethyl ester hydrolysis by ficin are discussed, including consideration of an allosteric modification mechanism.
Studies on the Interaction of F‐Actin with Tropomyosin
Authors: Drabikowski W. , Nowak E. 
Published: March 2005

The interaction of tropomyosin with F‐actin was studied under various conditions. The maximum ratio of tropomyosin bound to actin was found by ultracentrifugation to be 1:2, by weight. Viscometric study showed, however, no further increase of viscosity due to formation of the actin‐tropomyosin complex when the above ratio exceeded 1:5. Higher ionic strength caused dissociation of the complex.The comparison of the rate of polymerization of tropomyosin free actin under various salt conditions with that in the presence of tropomyosin showed that the latter partially modified some properties of actin.The pattern of polymerization of actin extracted at room temperature (RT‐actin) resembled that of the mixtures of actin with tropomyosin, although some differences were observed indicating tighter binding in the case of RT‐actin, possibly due to the presence of troponin.The results obtained are discussed from the point of view of the possible localization of tropomyosin in thin flaments.
Initiation de chaînes de RNA par la RNA polymérase in vitro
Authors: Sentenac A. , Ruet A. , Fromageot P. 
Published: March 2005

In the presence of cordycepin triphosphate (3′‐deoxyadenosine triphosphate), an analogue of ATP, RNA polymerase synthesizes very short RNA chains in vitro. These chains, labelled on the first nucleotide with [γ32P]GTP, are acid soluble but are retained on nitrocellulose filters. After alkaline hydrolysis, chromatography on Dowex 1 (CI−)shows that the 32P‐labelled molecules behave like guanosine tetraphosphate.
When filtered on Sephadex G‐75 the oligonucleotides synthesized are associated with a structure of high molecular weight, which is probably the complex DNA‐enzyme, and remain so after ribonucleases or deoxyribonuclease treatment.
The number of nascent oligonucleotides measured by incorporation of [γ32P]GTP, represents only 50% of the chains initiated by guanosine in the control. This observation, together with the fact that nascent oligonucleotides are not sensitive to pancreatic or T1 ribonuclease, shows that they are involved in a structure which determines their stability. This structure could be an Enzyme‐(DNA‐RNA hybrid) complex at the growing point of the RNA chains.
The kinetics of initiation of RNA are the same whether chain propagation is prevented or not.
The number of RNA chains initiated per phage T7 DNA molecule has been measured with [γ32P]ATP and [γ32P]GTP. It increase linearly with the amount of RNA polymerase added although the amount of RNA made tends towards a maximum. On the other hand, the number of nascent oligonucleotides made does not increase with an excess of enzyme, which shows the existence of a finite number of initiation sites. These results are thought to indicate that initiation sites are reutilized during normal RNA synthesis.
The number of chains initiated by adenine and guanine per phage T7 DNA at the point where RNA synthesis, as a function of RNA polymerase concentration, departs from proportionality, is around 50.
Excess of α‐Globin Synthesis in Homozygous β‐Thalassemia Its Cytoplasmic Molecular Forms
Authors: Conconi F. , Bargellesi A. , Pontremoli S. 
Published: March 2005

The cytoplasmic distribution of the excess newly synthesized α‐globin, present in the red cell lysate of β‐thalassemic subjects has been investigated.The labelled globins present in the cytoplasm, as well as in purified Hb A and Hb F, were separated by CM‐cellulose chromatography, and the ratios 3Hα/3Hβ and 3Hα/3Hγ were determined.The results obtained show that an intracellular exchange occurs between non radioactive α‐chains of existing hemoglobins and the newly synthesized ones. Since the amount of labelled α‐globin chains which is found in combination with Hb A or Hb F does not account for all the newly synthesized α‐globin, it is concluded that part of this α‐globin must exist in a free form. A series of experimental data support the conclusion that this “free fraction” is probably composed of non heminated α‐globin molecules.
La Tyrosyl‐tRNA synthétase de Escherichia coli
Authors: Chousterman S. , Sonino F. , Stone N. , Chapeville F. 
Published: March 2005

Partially purified tyrosyl‐tRNA synthetase from Escherichia coli B was incubated either in the presence of [14C]tyrosine and ATP or [14C]tyrosyl‐tRNA and AMP, or [3H]tyrosinyl‐AMP, a structural analogue of tyrosyl‐AMP. After filtration on a Sephadex G‐75 column, [14C]tyrosyl‐AMP: Enzyme and [3H]tyrosinyl‐AMP: Enzyme complexes were isolated. The [14C]tyrosine is transferred directly from [14C]tyrosyl‐AMP: Enzyme to the tRNA; no such transfer was observed with the [3H]tyrosinyl‐AMP:Enzyme. The stability of the tyrosyl‐AMP:Enzyme complex was examined under different conditions: (a) its half‐life at pH 7.0 is 150 minutes at 0°, 35 minutes at 20°, and 5 minutes at 37°; this instability is due to the non‐enzymatic hydrolysis of the anhydride bond; (b) its stability is affected neither by the presence of homologous or heterologous tRNAs which do not accept tyrosine, nor by the presence of ATP and/or tyrosine; (c) when the preformed complex is incubated with tyrosinyl‐AMP in conditions where this analogue inhibits completely tyrosyl‐AMP: Enzyme formation from ATP and tyrosine, tyrosyl‐AMP is not exchanged with tyrosinyl‐AMP. The stability of tyrosinyl‐AMP: Enzyme was also studied: after incubation for 30 minutes at 37°, or for several days at 4°, 90% of the [3H]tyrosinyl‐AMP was found associated with the enzyme after gel filtration. The complex remains unaffected when incubated in the presence of inorganic pyrophosphate, ATP or tRNA, but in the presence of tyrosinyl‐AMP, tyrosinol and ATP or tyrosine and ATP, only 20% of [3H]tyrosinyl‐AMP remained bound to the enzyme.These results show that tyrosyl‐AMP does not dissociate from the enzyme while tyrosinyl‐AMP does. It follows that, despite the great similarity of tyrosyl‐AMP and tyrosinyl‐AMP, the differences between the anhydride and the ester bond seem to influence decisively the stability of the complexes formed with the enzyme.
Oxidative Metabolism in Cells Isolated from Brown Adipose Tissue
Authors: Prusiner S. B. , Cannon B. , Lindberg O. 
Published: March 2005

Brown fat cells exhibit a classical mitochondrial electron transport system.
The respiration of brown fat cells can be stimulated by nor‐epinephrine and this hormonally mediated respiration can be mimicked by oleate.
Both nor‐epinephrine and oleate‐stimulated respiration can be blocked by rotenone, antimycin, azide and malonate, and the antimycin block can be by‐passed by tetramethyl‐p‐phenylenediamine.
α‐Glycerol phosphate and succinate were found to stimulate respiration.
The potent α‐glycerol phosphate oxidase cannot quantitatively contribute to the respiration evoked during catecholamine induced thermogenesis.
Stereoselective Alkylation with Asymmetric Reagents
Authors: Eisele B. , Wallenfels K. 
Published: March 2005

Yeast alcohol dehydrogenase is inactivated at different rates by the antipodes of α‐iodopro‐pionic acid and its amide; the d(+) antipodes react faster. The influence of coenzyme, substrate and pH on the reaction velocity were investigated. Under all conditions second order kinetics have been observed. Activation parameters for the alkylation of the SH groups with the different antipodes were calculated from the temperature dependence of the reaction rates. Large differences in the activation enthalpies (ΔΔH= 6.8 kcal/mole and 2.3 kcal/mole, respectively) were found for the reaction of yeast alcohol dehydrogenase with the different antipodes of α‐iodopropionic acid and α‐iodopropionamide. These were compensated for by differences in activation entropy of —18.4 and —5.6 entropy units, respectively.These results are interpreted by assuming highly dissymmetric surroundings of the reacting SH groups favouring the d(+) antipodes in the transition state of the alkylation reaction. The negative activation entropy indicates that this favourable transition state may be attained only by restriction of the degrees of freedom.The relationship between the stereoselective chemical reactivity and the stereospecificity of the enzyme is discussed.