bluuebunny/crossref_metadata_2025_split huggingface dataset:
Partition of Proteins in Two‐Phase Systems Containing Charged Poly(ethylene glycol)
Authors: Johansson Göte , Hartman Annika , Albertsson Per‐Åke 
Published: March 2005

Proteins have been partitioned in dextran–poly(ethylene glycol)–water biphasic systems containing positively charged poly(ethylene glycol). The effect of pH, buffer concentration, polymer concentration, protein concentration and temperature on the partition has been investigated. Characteristic extraction profiles are obtained when the percentage of protein in the upper phase is plotted versus pH. By keeping the salt concentration low, very steep extraction profiles can be obtained. Thus, within an interval of one pH unit, a protein is transferred from one phase to the other. This pH‐interval is located close to the isoelectric point of the protein.The possibility of resolving the components in a natural enzyme mixture, yeast lysate, by counter‐current distribution in this type of biphasic system has been investigated. According to activity measurements, several glycolytic enzymes are resolved by this procedure. Good separation is achieved. For some of the enzymes the activity is split into more than one peak in the counter‐current distribution.
The Interaction of Glutamate Dehydrogenase and Malate Dehydrogenase with Phospholipid Membranes
Authors: Dodd George H. 
Published: March 2005

The interaction between sonicated dispersions of purified phospholipids, corresponding to those found in the inner mitochondrial membrane, and the enzymes was studied by kinetic and spectroscopic methods.With glutamate dehydrogenase the charge type of the lipid head group determines the extent of lipid‐protein complexing. Phosphatidylcholine did not significantly alter the activity of the enzyme at lipid concentrations up to 1 mM, and the nuclear magnetic resonance (NMR) spectrum of the lipid (1%, w/v) was not altered in the presence of enzyme (up to 20% w/v). Phosphatidylethanolamine did not affect the activity in the oxidation of glutamate, but interfered with the assay in the reverse direction, possibly owing to Schiff‐base formation with 2‐oxoglutarate. The negatively charged cardiolipin, which is found uniquely in the inner mitochondrial membrane, strongly and reversibly inhibited the enzyme (50% inhibition with 0.1–20 μM lipid, depending on conditions). Another anionic phospholipid, phosphatidylserine (which does not occur in mitochondria), also inhibited the enzyme, but less effectively (50% inhibition at about 8 μM). The lipid‐enzyme interaction decreases with increasing ionic strength, indicating an electrostatic contribution to the binding. NMR studies suggest that there is also an apolar interaction involving the lipid chains.The studies with malate dehydrogenase concentrated on detecting possible stabilization of this enzyme through interaction with lipids. A clear‐cut stabilization was found with lysophosphatidylcholine. Under conditions such that the enzyme alone lost 50% on the initial activity in about 2 h, lysophosphatidylcholine gave a slight (20%) activation and this activity remained constant for several hours. NMR studies indicated that, as with the related enzyme 3‐hydroxybutyrate dehydrogenase, both the polar choline head group and the hydrocarbon chains are involved in the binding of the lipid to the protein.These specific lipid · protein complexes are probably examples of allotopic interactions. This form of complexing may explain aspects of the behaviour of these enzymes in vivo.
Deoxyribonucleoside‐Triphosphate Pools and DNA Synthesis in Synchronized Hamster Cells
Authors: Skoog K. Lambert , Nordenskjöld Bo A. , Bjursell K. Gunnar 
Published: March 2005

Cellular pools of all four deoxyribonucleoside triphosphates were determined in synchronized Chinese hamster cells obtained by mitotic selection or by the addition of complete medium to isoleucine‐starved cells. Using either technique the degree of synchrony is high and a maximum of more than 80% of the cells incorporated labeled thymidine during a short pulse period.Cells not involved in DNA synthesis contained measurable pools of deoxyribonucleoside triphosphates, but the pools expanded when the cells entered S phase. At all time points the pool of dGTP was the largest while the dGTP pool was the smallest amounting to 1–5% of the size of the dCTP pool. The size of the dCTP pool was closely correlated to the rate of DNA synthesis while the other pools reached their highest values after DNA synthesis.
Uptake and Reduction of Radioactive Folate by Everted Sacs of Rat Small Intestine
Authors: Selhub Jacob , Brin Hanna , Grossowicz Nathan 
Published: March 2005

Uptake of pteroylglutamie acid ([2‐14C]PteGlu) by everted sacs of rat small intestine was studied. Under conditions of zero initial concentration gradient, a net transport of PteGlu from the mucosal to the serosal side was found at initial concentrations not exceeding 7 μM.Using a simplified chromatographic procedure for estimation of reduced folates it was found that part of the transported PteGlu was converted to reduced forms, tentatively identified as tetrahydropteroylglutamic acid (H4PteGlu) and its methyl and formyl derivatives. PteGlu was reduced throughout the small intestine. The reduction was time and concentration dependent with an apparent Km of 1.55 μM.Low concentrations of methotrexate (23 μM) inhibited the reduction of PteGlu but did not affect its transport; at a much higher concentration (3 μM) methotrexate decreased the accumulation of folates in the tissue without significantly affecting the vitamin concentration in the serosal solution.
Evidence for Metabolic Compartmentation in Escherichia coli
Authors: Macnab Robert , Moses Vivian , Mowbray John 
Published: March 2005

The incorporation into amino acids in protein of 14C from (a) glycerol and maltose, and (b) galactose and lactose, alone and in competition, has been measured in Escherichia coli during aerobic growth.Glycerol and maltose contributed carbon equally to growth and to amino acids whose precursors are tricarboxylic acid cycle intermediates, but unequally to amino acids derived from glycolysis or the pentose phosphate pathway. Glycolytic compartmentalisation has been suggested and the possible nature of this is briefly discussed.Lactose and galactose complemented one another as growth substrates. Free galactose, and galactose from lactose hydrolysis, contributed unequally to some amino acids, as did the glucose and galactose moieties when lactose was the sole substrate. A simple compartmented model of the cell has been adduced to explain these results. These data are in accord with an earlier suggestion of metabolic channelling of the oxidative carbohydrate pathways in E. coli.
Transport of Lactate and Succinate by Membrane Vesicles of Escherichia coli, Bacillus subtilis and a Pseudomonas Species
Authors: Matin Abdul , Konings Wilhelmus N. 
Published: March 2005

Transport of d‐lactate, l‐lactate and succinate in isolated membrane vesicles of Escherichia coli, Bacillus subtilis and a fresh‐water Pseudomonas species occurs actively by specific transport systems which are linked to the electron transport chain. These substrates are not transported by the membrane vesicles unless an electron donor (reduced phenazine methosulfate, NADH, d‐lactate, l‐lactate, or succinate) is present in the reaction mixture; in the presence of an electron donor, the substrates or their oxidized forms are concentrated several‐fold inside the membrane vesicles; and inhibitors of the electron transport chain markedly inhibit this electron‐donor mediated transport. In addition, transport of pyruvate in membrane vesicles of E. coli, but not in those of B. subtilis and the Pseudomonas sp. is stimulated by reduced phenazine methosulfate.
Activity of the membrane‐bound lactate and succinate dehydrogenases is not essential for the transport of lactate and succinate, since inhibitors of these activities have little effect on reduced phenazine methosulfate‐ or NADH‐energized transport of these substrates; and the Km values for oxidation of these substrates by the membrane vesicles are 30‐ to 80‐fold higher than the Km values for their transport.
d‐Lactate and l‐lactate cause strong inhibition of the transport of each other in the three organisms, suggesting that the two substrates share a common transport system. In general, transport of these substrates is not inhibited by dicarboxylic acids. In contrast, transport of succinate is strongly inhibited by other dicarboxylic acids in the three organisms, suggesting that these substrates may share a common transport system. The transport systems for lactate and succinate are repressed by glucose.
A Study of Several Bonds Hypersensitive to Proteases in a Complex Flavohemoenzyme, Yeast Cytochrome b2
Authors: Naslin Lillian , Spyridakis Athanase , Labeyrie Françoise 
Published: March 2005

A detailed study of the polypeptide fragments formed, and of their evolution during the first stages of the trypsinolysis of two homologous active cytochrome b2 molecules, was carried out. These cytochromes are complex flavoheme‐l‐lactate dehydrogenases extracted from different yeasts and having different molar activities.Our purpose was to gather information concerning the bonds, which, in the native proteins, are the most sensitive to trypsin (termed here protease‐hypersensitive bonds). The products resulting from the fist hydrolytic events were thus analysed using disc‐electrophoresis in sodium dodecylsulfate according to the method of Shapiro et al. (1967). Electrophoretic patterns were related to the molecular weight and to the amount of each intermediate species as a function of the extent of reaction. Schemes of the series of events, (including several rate constants) were derived from this set of data. A tentative map of the homologous loci of protease‐hypersensitive bonds along the chains of the two cytochrome b2 molecules is proposed. Considering that such bonds which are very reactive towards trypsin are certainly located on flexible and thus loosely structured regions at the surface of the molecule, this map gives a first insight into the overall secondary and tertiary structure of the chain which includes globules of high internal stability.An important finding was that the relative reactivities of the various protease‐hypersensitive bonds identified, depend on conditions such as ionic strength and the binding of a ligand which probably changes the conformation. The protein transitions are therefore possibly described in terms of modulation of the loci of these bonds and of their reactivity.
Covalent Binding of Lipid to Protein
Authors: Hantke Klaus , Braun Volkmar 
Published: March 2005

A new structure of a lipid and its covalent linkage to a protein (murein‐lipoprotein of the Escherichia coli outer membrane) is described. Glycerylcysteine (S‐(propane‐2′,3′‐diol)‐3‐thio‐2‐aminopropanic acid) at the N‐terminal end of the polypeptide chain is the attachment site of two ester‐bound fatty acids. An additional fatty acid is bound as amide to the N‐terminal group. The diglyceride residue on the cysteine could be derived from the phospholipid pathway since the fatty acid composition is very similar to that of the phospholipids from the same cells. In contrast, 65% of the amide‐linked fatty acid is palmitate. The main fatty acids are palmitic acid (53%), cis‐vaccenic acid (20.7%), 9,10‐methylene‐hexadecanoic acid (10.6%) and palmitoleic acid (9.4%).The structure of glycerylcysteine was established by chemical degradation of the compound containing the fatty acids, by incorporation studies of [35S]sulfate, [35S]cysteine, [14C]cystine, [methyl‐3H]methionine and [2‐3H]glycerol and by chemical synthesis. Glycerylcysteine was isolated as a constituent of the peptide (Ser)‐Ser‐Am‐Ala‐Lys. The C‐terminal part of the lipopeptide sequence overlaps with the N‐terminal end of the known sequence of the polypeptide chain. Its N‐terminal position was confirmed by isolation of the N‐terminal polypeptide fragment (position 1–31) after cleavage with cyanogen bromide. This contained the lipid.
The Free Lipids of Brucella melitensis and Bordetella pertussis
Authors: Thiele Otto Wolfgang , Schwinn Gottfried 
Published: March 2005

Free lipids were extracted from Brucella melitensis and Bordetella pertussis and their compositions compared with one another. The bacteria contained 1.6% and 8.1% free lipids by dry weight of the respective species.The major component of the neutral lipids of Bruc. melitensis was coenzyme Q10 (0.6 μmol/g bacterial dry weight). Smaller amounts of 1,2‐ and 1,3‐diglycerides and of fatty acyl monoesters of ethanediol and of unknown α‐glycols have been found among the neutral lipids of Bruc. melitensis. The major component of the neutral lipids of Bord. pertussis was a fraction of unesterified fatty acids. Moreover, coenzyme Q8 (0.2 μmol/g bacterial dry weight) was identified in Bord. pertussis. Almost all saponifiable lipids of Bruc. melitensis contained considerable amounts of a C19‐cyclopropane acid (lactobacillic acid) whereas no cyclopropane acid was found among the unesterified fatty acids of Bord. pertussis.From the polar lipid fraction of Bruc. Melitensis diphosphatidylglycerol (cardiolipin), phosphatidylglycerol, phosphatidylethanolamine (cephalin), phosphatidylcholin (lecithin) and phosphatidylserine were isolated; the phosphatidylethanolamine fraction also contained phosphatidyl‐N‐methylethanolamine and phosphatidyl‐N, N‐dimethylethanolamme. Phosphatidylcholine was the major component (37.6% of all phospholipids) in Bruc. melitensis. Determination of positional distribution of the component fatty acids revealed lactobacillic acid to be preferentially located in the number 2‐positions of all phospholipids, while the 1‐positions were preferentially occupied by palmitic acid and in most phospholipids, also by hexadecenoic acid.From Bord. Pertussis, diphosphatidylglycerol, phosphatidylethanolamine, and phosphatidylserine were isolated, but no phosphatidylcholine, no partially methylated phosphatidylethanolamine, and no phosphatidylinositol were found. Palmitic acid was located more frequently at the number 1‐positions, hexadecenoic acid more frequently at the number 2‐positions of the phospholipids. Diphosphatidylglycerol and phosphatidylethanolamine probably contained minor amounts of a C19‐cyclopropane acid.An ornithine‐containing lipid was isolated from Bruc. melitensis and Bord. pertussis. Besides ornithine these lipids contained ethanediol and fatty acids. A similar finding is recorded in Bruc. abortus. Ethanediol appeared to be ester linked to ornithine and to a fatty acid. Another fatty acid is probably linked to one of the amino groups of ornithine by an amide bond. In Bruc. melitensis and Bruc. abortus lactobacillic acid is predominantly ester linked, while palmitic and stearic acids are predominantly amide linked. There is no lactobacillic acid at all in the ornithine lipid of Bord. pertussis. No hydroxy fatty acid has been found in the ornithine lipids of any strain.From the lipid patterns of Bruc. melitensis, of Bruc. abortus studied previously, and of Bord. pertussis we conclude that it is probably not justified to include Brucellae and Bordetellae in the one natural family of Brucellaceae.
Carbon‐13 Nuclear‐Magnetic‐Resonance and Infrared‐Absorption Spectroscopy of Valinomycin and Its Alkali‐Ion Complexes
Authors: Grell Ernst , Funck Theodor , Sautee Hanspeter 
Published: March 2005

The 13C nuclear magnetic resonance spectra of the membrane‐active antibiotic valinomycin and its alkali‐ion complexes are interpreted. From the 13C chemical shifts and infrared absorptions of the highly stable K+, Rb+ and Cs+ complexes in methanol it is concluded that the ligand conformations of these complexes must be similar. The essential features are the coordination of all ester carbonyl groups with the cation in addition to the formation of intramolecular hydrogen bonds between the amide groups. In contrast to these results the weaker Na+ complex of the antibiotic exhibits different spectral properties which are more similar to those of the uncomplexed antibiotic in polar solvents thus indicating the existence of a considerably different ligand conformation in this complex. The structure of the Na+ complex can be characterized by a reduced number of coordinated ester carbonyl groups and intramolecular hydrogen bonds between the amide groups as compared to the complexes with larger cations.The conformational rearrangements occurring during the complexation of cations are compared to those observed for the uncomplexed valinomycin in solvent systems of different polarities. Evidence is presented that the interaction of the uncomplexed antibiotic with polar solvent molecules leads to conformational rearrangements which can be characterized by a step‐wise opening of a closed structure stabilized by intramolecular hydrogen bond formation.
Some Evidence on the Site of Action of Plastocyanin in the Photosynthetic Electron‐Transport Chain between Cytochromes ƒ and P700
Authors: PlesniČAR Marijana , Bendall Derek S. 
Published: March 2005

The effect of plastocyanin on the light‐induced steady‐state changes of cytochrome ƒ in chloroplast particles that were depleted of plastocyanin has been studied. It was observed that the extent of cytochrome ƒ photooxidation in sonic particles in the steady‐state during a short exposure to long‐wave red light was approximately 30% of that in chloroplasts. The addition of a catalytic amount of plastocyanin to the sonic particles stimulated cytochrome ƒ photooxidation to 80% of its steady‐state value in untreated chloroplasts. The reduction of cytochrome ƒ by short‐wave red light was not inhibited by sonic treatment of chloroplasts.The rate of cytochrome ƒ photooxidation in the presence of a catalytic amount of detergent‐treated chloroplasts was proportional to the amount of plastocyanin added.The enhancement of cytochrome ƒ photooxidation either in sonic particles or in digitonin‐treated particles after an addition of plastocyanin was decreased in the presence of the copper‐chelating agent sodium diethyldithiocarbamate. Plastocyanin preincubated with sodium diethyldithiocarbamate showed less ability to stimulate cytochrome c photooxidation by detergent‐treated chloroplasts.
Structures of Fucose‐Containing Glycolipids with H and B Blood‐Group Activity and of Sialic Acid and Glucosamine‐Containing Glycolipid of Human‐Erythrocyte Membrane
Authors: Kościelak Jerzy , Piasek Andrzej , Górniak Halina , Gardas Andrzej , Gregor Anita 
Published: March 2005

Fucose‐containing glycolipids with H and B blood group activity as well as sialic acid and glucosamine‐containing glycolipid were isolated from human erythrocytes. The sequences and linkages of carbohydrate residues in these glycolipids were determined by partial acid hydrolysis, periodate oxidation and methylation studies. Anomeric linkage were ascribed to those materials on the basis of their optical rotations, infrared spectra, susceptibility to enzymes and immunological activities.The structures of glycolipid H‐I and H‐II were respectively fucopyranosyl‐α‐(1 → 2)‐galacto‐pyranosyl‐β‐(1 → 4)‐2‐acetamido‐2‐deoxy‐glucopyranosyl‐β‐(1 → 3)‐galactopyranosyl‐β‐(1 → 4)‐glucopyranosyl‐β‐(1 → 1)‐ceramide; fucopyranosyl‐α‐(1 → 2)‐galactopyranosyl‐β‐(1 → 4)‐2‐acetamido‐2‐deoxy‐glucopyranosyl‐β‐(1 → 3)‐galactopyranosyl‐β‐(1 → 4)‐2‐acetamido‐2‐deoxy‐glucopyranosyl‐β‐(1 → 3)‐galactopyranosyl‐β‐(1 → 4)‐glucopyranosyl‐β‐(1 → 1)‐ceramide. The structure of glycolipid B‐I was galactopyranosyl‐α‐(1 → 3)‐[fucopyranosyl‐α‐(1 → 2)]‐galactopyranosyl‐β‐(1 → 4)‐2‐acetamido‐2‐deoxy‐glucopyranosyl‐β‐(1 → 3)‐galactopyranosyl‐β‐(1 → 4)‐glucopyranosyl‐β‐(1 → 1)‐ceramide. The structure of sialic acid and glucosamine‐containing glycolipid was N‐acetyl‐neuraminosyl‐α‐(2 → 3)‐galactopyranosyl‐β‐(1 → 4)‐2‐acetamido‐2‐deoxy‐glucopyranosyl‐β‐(1 → 3)‐galactopyranosyl‐β‐(1 → 4)‐glucopyranosyl‐β‐(1 → 1)‐ceramide. The exclusive presence of 4′‐galactosyl‐N‐acetylglucosamine in four erythrocyte glycolipids makes it likely that the (1 → 3) linkage between galactose and N‐acetyl‐glucosamine residues is missing from all membrane glycolipids which comprise these two sugars. Therefore the formation of Lea and Leb active structures is not possible in membrane glycolipids and all Lewis activity of erythrocytes must be derived from glycolipids absorbed from blood plasma.
Sites Along the Electron‐Transport Chain Controlled by the Energy‐Conversion System in Chloroplasts
Authors: Eisenbach Michael , Carmeli Chanoch 
Published: March 2005

The light‐induced absorbance changes of electron transport carriers were measured in intact (class I) chloroplasts which catalyzed oxygen evolution with phosphoglyceric acid or CO2 as electron acceptors. In these chloroplasts the reduction of cytochrome b563, induced by light at 720 nm, increased in the presence of ADP or in the presence of uncouplers such as gramicidin and NH4Cl.
In the presence of ADP there was a decrease in the light‐induced reduction of C550 and in the oxidation of cytochrome f. The effect of ADP was augmented when phosphoglyceric acid was used as an electron acceptor in class I chloroplasts. A decrease in the photoreduction of C550 was also observed in the presence of the uncoupler gramicidin in class I and class II, with benzylviologen as an electron acceptor, and in class II chloroplasts with ferricyanide.
The uncoupler gramicidin caused an increase in the light‐induced reduction of cytochrome b569. Based on these effects of ADP or uncoupler on the steady state of the light‐induced redox level of the carriers, we proposed sites along the electron transport chain in which the rate of electron flow is controlled by the energy conversion system.
Occurrence of 2‐Amino‐2‐deoxy‐hexuronic Acids as Constituents of Vibrio parahaemolyticus K15 Antigen
Authors: Torii Mitsuo , Sakakibara Keiko , Kuroda Koichi 
Published: March 2005

Acidic capsular polysaccharide of Vibrio parahaemolyticus A55 (05: K15) was esterified with ethylene oxide and then reduced with sodium borohydride. Hydrolysis of the reduced material gave two hexosamines identical with mannosamine (2‐amino‐2‐deoxy‐mannose) and gulosamine (2‐amino‐2‐deoxy‐gulose) on paper and ion‐exchange chromatograms. This fact suggested that the K15 antigen contained mannosaminuronic acid (2‐amino‐2‐deoxy‐mannuronic acid) and gulosaminuronic acid (2‐amino‐2‐deoxy‐guluronic acid).
Direct acid hydrolysis of the antigen gave two 2‐amino‐2‐deoxy‐hexuronic acids. These two were identified as mannosaminuronic and gulosaminouronic acids by paper chromatography and paper electrophoresis, being compared with synthesized standard samples.
Titration of acid form of the polysaccharide indicated that about 91.5% of the polysaccharide was composed of the N‐acetyl‐hexosaminuronic acids.