bluuebunny/crossref_metadata_2025_split huggingface dataset:
A Study on the Biosynthesis of Protein A in Staphylococcus aureus
Authors: MOVITZ Jan 
Published: March 2005

Protein A is a cell wall constituent of Staphylococcus aureus. After being synthesized on the ribosomes, the protein A molecules are linked to the peptidoglycan. This is a very rapid reaction which is completed within approximately 1 min; it is suggested that the molecules leave the ribosomes and are then directly incorporated into the peptidoglycan without passing a soluble cytoplasmic stage.The synthesis of protein A de novo and peptidoglycan take place independently of each other. Also, the incorporation of protein A into the cell wall proceeds when the synthesis of peptidoglycan has been completely inhibited by vancomycin, i.e. the synthesis of peptidoglycan is not essential for the incorporation of protein A. It is therefore concluded that protein A can be linked to pre‐existing peptidoglycan chains.
Heterogeneity of Erythrocyte Catalase II
Authors: AEBI Hugo , WYSS Sonja R. , SCHERZ Bernhard , SKVARIL František 
Published: March 2005

Normal human erythrocyte catalase, when isolated by the method of Mörikofer et al. [Eur. J. Biochem. 11 (1969) 49–57], is heterogenous in two respects: (a) there are alternative molecular forms; (b) besides the active tetramer, enzyme dissociation products (dimer, monomer) are present.
By means of exclusion chromatography on Sephadex G‐150 it was shown that catalase preparations contain dimer and monomer in small amounts (∼ 4%). Upon storage, the yield in subunits can be considerably enhanced (up to 15%).
Residual catalase activity in the blood of an individual homozygous for Swiss Type Acatalasemia (1–2% of normal level) consists of a catalase variant, which is unstable, but of approximately normal specific activity. In this mutant, the tendency of dissociation into subunits is greatly enhanced. Upon purification, little tetramer but appreciable amounts of the inactive dimer are obtained.
The antigenic properties of these molecular species were investigated using anti‐catalase and anti‐catalase‐subunit immunoglobulins G. There is complete antigenic identity between the normal and the variant catalase; the same is true for the normal and variant dimer. Different antigenic properties of dimer and monomer, however, offer evidence for the existence of hidden determinants in the subunits.
Human Glutathione Reductase: Purification of the Crystalline Enzyme from Erythorocytes
Authors: WORTHINGTON David J. , ROSEMEYER Michael A. 
Published: March 2005

Glutathione reductase from human erythrocytes has been purified 40 000‐fold in 10 steps with an overall yield of 36%.The procedure included bulk separations on DEAE‐Sephadex and CM‐Sephadex, gel‐filtration on Sephadex G‐200, and salt fractionation with ammonium sulphate. The purified enzyme was crystallised from aqueous solution.The final preparation had a specific activity of 240 units/mg. The purification procedure was completed in 4 weeks, and gave 40 mg enzyme from 35 1 blood. The schedule for glutathione reductase allowed the concurrent isolation of glucose‐6‐phosphate dehydrogenase and 6‐phosphogluconate dehydrogenase.In solutions of thiols, the glutathione reductase preparation was homogeneous, as indicated by gel‐filtration, ultracentrifugation and electrophoresis. In the absence of thiols, the enzyme showed a tendency to form aggregates. Some of the physical properties of the enzyme are discussed in relation to previous difficulties encountered in its isolation.
Isolation and Characterization of Oligosaccharides from Canine Submaxillary Mucin
Authors: LOMBART Christian G. , WINZLER Richard J. 
Published: March 2005

Pure mucin isolated from canine submaxillary glands was treated with alkaline borohydride and products of degradation were fractionated by a procedure that included ion‐exchange chromatography, gel filtration, high‐voltage electrophoresis and paper chromatography.Seven acidic, reduced oligosaccharides were isolated being divided into two distinct types: one, containing sialic acid and no glucosamine or sulfate (type A) and the other containing glucosamine and sulfate but no sialic acid (type B).The most complete oligosaccharide of type A designated component (I) was a tetrasaccharide α‐l‐fucopyranosyl‐(1 → 2)‐d‐galactopyranosyl‐[N‐acetylneuraminyl‐(2 → 6)]‐2‐acetamido‐2‐deoxy‐d‐galactitol. The most complex oligosaccharide of type B designated component (V) has a hexasaccharide with the following structure: α‐l‐fucopyranosyl‐(1 → 2)‐β‐d‐galactopyranosyl‐(1 → 3, 4, or 6)‐β‐2‐acetamido‐2‐deoxy‐d‐glucopyranosyl 3 or 4 sulfate‐(1 → 6)‐α‐l‐fucopyranosyl‐(1 → 2)]‐β‐d‐galactopyranosyl‐(1 → 3) 2‐acetamido‐2‐deoxy‐d‐galactitol. All the other acidic isolated oligosaccharides were derived from these two main components.No oligosaccharide containing both sialic acid and glucosamine sulfate could be demonstrated. Based on these structural features, a possible biosynthetic pathway leading to the two different oligosaccharide chains is discussed.Variations in ratios of sugars observed in canine submaxillary mucin and especially the reciprocal relationship between sialic acid and fucose noted earlier (1962) by Dische, Pallavicini, Kavasaki, Smirnow, Cizek and Chien may reflect variations in the proportions of these two types of oligosaccharides.
Direct Fluorometric Determination of Liver Alcohol Dehydrogenase with Berberine
Authors: KOVÁŘ Jan 
Published: March 2005

A fluorometric method for the determination of the concentration of horse liver alcohol dehydrogenase is described. It is based on the formation of a strongly fluorescent complex between the enzyme and berberine. The relative increase of fluorescence after the addition of the enzyme into a solution of berberine (at fixed concentration) is a linear function of the enzyme concentration.The results are easy to obtain, nearly temperature independent and as reliable as those of the commonly used activity measurement.The principle of this method can also be used for the estimation of other proteins forming strongly fluorescent complexes with slightly fluorescent probes.
Regulatory Behavior of Monomeric Enzymes
Authors: RICARD Jacques , MEUNIER Jean‐Claude , BUC Jean 
Published: March 2005

A new concept, that of a mnemonical transition, is proposed to explain the departure from Michaelian behavior of monomeric enzymes following ordered reaction mechanisms. The concept integrates three simple ideas: the free enzyme occurs under two conformational states in equilibrium; the collision of any of these forms with the first substrate induces the same third new configuration required for proper substrate binding; the collision of only one of these enzyme forms with the last product stabilizes that form without any new conformational change. This whole set of definitions is equivalent to assuming that the free enzyme which is released after catalysis, is in a conformation different from the initial one. The enzyme can be said to “recall” for a while the configuration stabilized by the last product before relapsing to the initial conformation. The non‐hyperbolic behavior is thus the consequence of the cooperation of two different conformations of the free enzyme to the overall reaction process.The reciprocal steady‐state rate equations have been established and thoroughly discussed both for one‐substrate, one‐product and two‐substrate, two‐product mnemonical enzymes. The departure from Michaelian behavior does not appear as a consequence of a slow conformational transition, but is defined in a simpler way by the relative values of the activation free energies of conformation changes required for substrate binding on the two enzyme forms.A two‐substrate, two‐product enzyme following an ordered reaction mechanism and exhibiting the mnemonical transition has a very distinctive kinetic behavior. The curvature of the primary plots is observed with regard to the first substrate only, and is independant of the concentration of the second substrate as well as that of the first product. The enzyme is not inhibited by an excess of the substrate and the primary plots are either concave up or down. The slopes and the intercepts of the straight lines obtained in double reciprocal plots with the second substrate should give, when these are replotted against the reciprocal of the first substrate concentration, a straight line and a curve, respectively. The cooperation of the enzyme conformations to the overall reaction process can be either positive or negative. Since the reciprocal plots cannot exhibit an extreme, the extent of that cooperation can be measured by the numerical value of the second derivative of the reciprocal rate equation. The extent of cooperation between the free enzyme forms is highly controlled by the concentration of the last product. If the cooperation was already negative, the product strengthens that cooperation. If, on the other hand, the cooperation was positive, the product decreases or even reverses that cooperation.A very general property of one‐sited mnemonical enzymes is that cooperation between enzyme forms is only kinetic and does not appear in the substrate binding isotherms.
Quantitation of Ovalbumin mRNA in Hen and Chick Oviduct by Hybridization to Complementary DNA
Authors: COX Ronald F. , HAINES Michael E. , EMTAGE J. Spencer 
Published: March 2005

DNA complementary to ovalbumin mRNA was synthesized using reverse transcriptase from avian myelobastosis virus and ovalbumin mRNA, purified 60–70‐fold with respect to polysomal RNA, as template.
The quantitative distribution of ovalbumin mRNA sequences in hen and chick oviduct was determined by hybridization with ovalbumin complementary DNA (cDNA). In hen, we calculate that there are approximately 36000 ovalbumin mRNA molecules per oviduct tubular gland cell. The polysome fraction contains 93% of ovalbumin mRNA; nuclear and non‐polysomal cytoplasmic RNA fractions contain 5% and 2% respectively. In the nucleus, the majority of sequences exist as the 16‐S “translatable” form when analyzed on formamide gradients. High‐molecular‐weight RNA species (> 30 S) were detected, and contained less than 2% of total nuclear mRNA sequences.
The appearance of ovalbumin mRNA sequences in total, nuclear and polysomal RNA fractions from chick oviduct was measured in response to estradiol. Prior to hormone treatment, low levels of sequences were present in all fractions, approximately one mRNA molecule per gland cell being detected in nuclei. Up to 2 h after treatment a small increase in ovalbumin mRNA concentration was observed in nuclear and, to a lesser extent, in polysomal RNA. Between 3 and 6 h ovalbumin mRNA accumulates rapidly in the polysome fraction and after 9 h the rate of appearance of this mRNA is approximately 7 molecules per min per gland cell.
Our results indicate that the induction by estradiol of ovalbumin synthesis in oviduct is mediated either by the synthesis of new ovalbumin gene transcripts or the stabilization of ovalbumin gene products.
Yeast Phenylalanyl‐tRNA Synthetase
Authors: FASIOLO Franco , EBEL Jean‐Pierre 
Published: March 2005

Phenylalanyl‐tRNA synthetase from yeast forms an enzyme · phenylalanyl adenylate complex, which has been isolated from the reaction mixture by gel filtration. The study of this complex leads to the conclusion that there are two binding sites for phenylalanyl adenylate per molecule of enzyme.No complex between enzyme and ATP could be characterised. Mg2+ is required for complex formation, but it can be replaced by monovalent cations in the transfer of phenylalanine from the complex to tRNAPhe. The kinetics of the transfer reaction show evidence that the rate‐limiting step is not the release of enzyme‐bound Phe‐tRNAPhe, but the acylation of tRNAPhe after the ternary complex phenylalanyl adenylate · enzyme · tRNAPhe has been formed.
The Distribution of Histones and Nonhistone Proteins in the ECTHAM‐Cellulose Fractions of Chromatin from Several Tissues
Authors: REECK Gerald R. , SIMPSON Robert T. , SOBER Herbert A. 
Published: March 2005

Chromatins from rabbit kidney and brain, calf liver, HeLa cells, and duck erythrocytes have been subjected to chromatography on ECTHAM‐cellulose, which has previously been shown to fractionate rabbit liver chromatin into low and high‐melting nucleoprotein species [Reeck et al. (1972) Proc. Natl Acad. Sci. U.S.A. 69, 2317–2321].Except for duck erythrocyte chromatin, the late‐eluted low‐melting chromatin fractions were greatly depleted in histone F1, as we reported previously for rabbit‐liver chromatin. No differences were found in the proteins of the ECTHAM‐cellulose fractions of duck‐erythrocyte chromatin, which, in contrast to all other chromatins examined, is known to be inactive in RNA synthesis. Differences in the nonhistone proteins of the early and late‐eluted chromatin fractions have been observed which appear to be unique for chromatin from each source.
Patulin Biosynthesis: The Role of Mixed‐Function Oxidases in the Hydroxylation of m‐Cresol
Authors: MURPHY Gillian , VOGEL Günter , KRIPPAHL Günther , LYNEN Feodor 
Published: March 2005

Two hydroxylation reactions of m‐cresol, an intermediate in the secondary biosynthesis of the antibiotic patulin, have been characterised in cell‐free preparations of Penicillium patulum. The ability to convert m‐cresol to 2,5‐dihydroxytoluene and m‐hydroxybenzyl alcohol is located in separable fractions of the 105000 × g microsomal pellet. The activities had Km values for m‐cresol of 80 μM and 5 μM, respectively. Both reactions require molecular oxygen and NADPH for activity. Inhibition of m‐cresol hydroxylases by carbon monoxide is reversible by light with a photochemical action spectrum maximal at 450 nm. Carbon monoxide difference spectra of mycel extracts show the presence of absorbing pigments at 422 and 450–455 nm.The enzymes are not appreciably inhibited by cyanide but are inhibited by cytochrome c. m‐Cresol hydroxylation therefore appears to be catalysed by typical mixed‐function oxidases involving an NADPH‐dependent reductase and a pigment like cytochrome P‐450.In the light of studies on the utilisation of radioactively labelled intermediates, the methyl hydroxylation of m‐cresol to m‐hydroxybenzyl alcohol probably represents an important reaction in the sequence leading to patulin formation. Ring hydroxylation of m‐cresol to 2,5‐dihydroxytoluene appears to be a side reaction functional at higher concentrations of m‐cresol.
Control of Ornithine Carbamoyltransferase Activity by Arginase in Bacillus subtilis
Authors: ISSALY Inda M. , ISSALY Abel S. 
Published: March 2005

Partially purified Bacillus subtilis ornithine carbamoyltransferase is cooperatively inhibited by excess of ornithine. The inhibition is decreased by lowering of pH from 9.2 to 7.6 without altering the affinity of the enzyme for the neutral species of ornithine which seems to be its true substrate.Arginine and lysine partially reverse this inhibition without affecting the catalytic activity of the enzyme. 2‐Aminobutyrate, an inhibitor of the enzyme, competitive with respect to ornithine, is not able to replace ornithine in this excess‐substrate type of inhibition. Zn2+ is also an inhibitor of ornithine carbamoyltransferase, with competitive action towards ornithine and its inhibition is added to the excess ornithine inhibition. These data suggest that ornithine carbamoyltransferase has a regulatory site for ornithine, distinct from the catalytic one.Partially purified arginase, prepared from the same strain, increases the inhibition of ornithine carbamoyltransferase by excess of ornithine when arginine is present. An association of the two enzymes is suggested by molecular sieving. These data postulate that ornithine carbamoyltransferase is regulated by arginase under the control of arginine and ornithine. The other substrate, carbamoyl phosphate, is not involved in this regulatory mechanism.Lysine and Zn2+ inhibit arginase competitively with respect to arginine without altering the effect of this enzyme on the inhibition of ornithine carbamoyltransferase. 2‐Aminobutyrate inhibits arginase non‐competitively with respect to arginine and prevents the inhibition of arginase towards ornithine carbamoyltransferase. These results suggest that arginase possesses a regulatory site for arginine, distinct from the catalytic one, which is also involved in the control mechanism.
Utilization of the Carbon and Hydrogen Atoms of Ethanol in the Biosynthesis of Steroids and Bile Acids
Authors: CRONHOLM Tomas , BURLINGAME Alma L. , SJÖVALL Jan 
Published: March 2005

[1‐2H2]Ethanol, [2‐2H3]ethanol and [1‐13C, 1‐2H2]ethanol were administered to bile fistula rats under conditions ensuring a maximal rate of oxidation for about 24 h. Cholesterol, cholic acid, chenodeoxycholic acid and 3β,11β,21‐trihydroxy‐5α‐pregnan‐20‐one were isolated from bile collected at intervals during the experiment. The fraction of molecules containing different numbers of heavy isotope atoms was measured by gas chromatography–mass spectrometry–computer techniques. The results were used to determine the origin of the cholesterol pools used as precursors for the different compounds isolated from bile.The bile acids were found to be formed from a cholesterol pool, the turnover of which was about 5 times more rapid than that of the cholesterol pool used for biliary excretion. The cholesterol pools used for biliary excretion and as corticosterone precursor seemed to be in equilibrium.The acetyl groups derived from ethanol were introduced into the steroids without exchange of hydrogens at C‐2, and the acetyl‐CoA pool used as precursor did not exchange with acetyl groups in e.g. citrate or fatty acids. A difference in incorporation of [2‐2H3]acetyl groups and [1‐13C]acetyl groups was interpreted to be due to an isotope effect. Ethanol contributed about 45% of the acetyl‐CoA pool used as precursor of cholesterol and bile acids.Deuterium from [1‐2H2]ethanol was incorporated in many positions of all the steroids. If it is assumed that the incorporation is mediated by NADPH, the deuterium excess of the hydrogens introduced was about 10 atoms%. The contribution via different pathways of an important part of the hydrogen and carbon atoms in cholesterol, bile acids and corticosteroids provides possible mechanisms by which ethanol may affect the biosynthesis and metabolism of steroids.
Studies on Phenylalanine‐Specific Transfer Ribonucleic Acid from Chick Embryos
Authors: WITTIG Burghardt , REUTER Stephanie , GOTTSCHLING Hubert 
Published: March 2005

The phenylalanine‐specific tRNAPhe from 13‐day‐old chick embryo muscle was purified. Its fluorescent and hydrophobic Y+ base was excised by mild acid treatment and characterized chromatographically and by its ultraviolet absorption and fluorescent spectra. The compound obtained after removal of the Y+ base from chick embryo tRNAPhe was characterized.

Chick embryo tRNAPhe looses by the removal of the Y+ base its phenylalanine acceptor capacity.
In this state it has a reduced sedimentation coefficient of 2.8 S, a low Tm value (22°C), only rudimentary hyperchromicity, but unchanged electrophoretic properties.
The compound obtained after removal of the Y+ base from chick embryo tRNAPhe regains normal melting behaviour and phenylalanine acceptor capacity when exposed to special renaturation conditions.
The Km, of the renatured form is 0.25 × 10−6 M and V= 220 pmol × min−1× ml−1, as compared to Km, = 0.11 × 10−6 M and V= 370 pmol × min−1× ml−1 for the native chick embryo tRNAPhe.
Precursors of Cytochrome Oxidase in Cytochrome‐Oxidase‐Deficient Cells of Neurospora crassa
Authors: WERNER Sigurd , SCHWAB Andreas J. , NEUPERT Walter 
Published: March 2005

Three different cell types of Neurospora crassa deficient in cytochrome oxidase were studied: the nuclear mutant cni‐1, the cytoplasmic mutant mi‐1 and copper‐depleted wild‐type cells.

The enzyme‐deficient cells have retained a functioning mitochondrial protein synthesis. It accounted for 12–16% of the total protein synthesis of the cell. However, the analysis of mitochondrial translation products by gel electrophoresis revealed that different amounts of individual membrane proteins were synthesized. Especially mutant cni‐1 produced large amounts of a small molecular weight translation product, which is barely detectable in wild‐type.
Mitochondrial preparations of cytochrome‐oxidase‐deficient cells were examined for precursors of cytochrome oxidase. The presence of polypeptide components of cytochrome oxidase in the mitochondria was established with specific antibodies. On the other hand, no significant amounts of heme a could be extracted.
Radioactively labelled components of cytochrome oxidase were isolated by immunoprecipitation and analysed by gel electrophoresis. All three cell types contained the enzyme components 4–7, which are translated on cytoplasmic ribosomes. The mitochondrially synthesized components 1–3 were present in mi‐1 mutant and in copper‐depleted wild‐type cells. In contrast, components 2 and 3 were not detectable in the nuclear mutant cni‐1. Both relative and absolute amounts of these polypeptides in the enzyme‐deficient cells were quite different from those in wild‐type cells.
The components of cytochrome oxidase found in the enzyme‐deficient cells were tightly associated with the mitochondrial membranes.
Processes, which affect and may control the production of enzyme precursors or their assembly to a functional cytochrome oxidase are discussed.
Structure and Synthesis of a Lipid‐Containing Bacteriophage
Authors: SCHÄFER Rolf , HINNEN Rosmarie , FRANKLIN Richard M. 
Published: March 2005

The lipid‐containing bacteriophage PM2 has four distinct structural proteins, as shown by polyacrylamide gel electrophoresis in the presence of sodium dodecylsulfate or electrophoresis on cellulose acetate strips in the presence of 6–8 M urea. The molecular weights of these four polypeptides determined in several polyacrylamide gel systems are: I = 43000, II = 26000–27000, III = 12500, IV = 4700. The isoelectric points of the four viral proteins were determined on disrupted virus particles by electrophoresis on cellulose acetate strips in the presence of 6 M urea at various pH values. The mobilities were extrapolated to zero mobility, a method preferable to isoelectric focusing where the denatured viral proteins aggregate and precipitate near the isoelectric point. The method is well suited for the determination of the isoelectric point of proteins in the native or only partially denatured state. Whereas proteins I, III and IV have isoelectric points slightly under neutrality (pH 6.2, 5.8 and 5.5, respectively), the isoelectric point of protein II is around 12.3. Calcium ions are tightly bound to protein II. After removal of the calcium ions in the presence of 6 M guanidine hydrochloride and EDTA, the pI of protein II is 9. The isoelectric point of the intact virus, determined on cellulose strips, is around 7.3. Protein I, the spike protein, is the only structural polypeptide which is soluble in aqueous solutions around pH 7. All of the proteins are soluble in 6–8 M urea or 6 M guanidine hydrochloride at pH 7–8.5. Proteins III and IV, the nucleocapsid core proteins, have the solubility properties of proteolipids or highly lipophilic polypeptides as shown by their solubility in nonionic detergents and by their distribution in a Bligh and Dyer fractionation. Neither glycolipids nor glyco‐proteins are present in the virus. The distribution of phospholipids has been determined by chemical modification of the outer lamella or outer and inner lamellae of the bacteriophage lipid bilayer. The viral lipid bilayer is asymmetric; most of the phosphatidylglycerol residues form the outer lipid layer, most of the phosphatidylethanolamine residues the inner lipid layer.A complete model of the virus, including the assembly process, is discussed at the end of this report.