bluuebunny/crossref_metadata_2025_split huggingface dataset:
The Effect of Ether Anaesthesia Stress on Cholesterol‐Side‐Chain Cleavage and Cytochrome P450 in Rat‐Adrenal Mitochondria
Authors: Simpson Evan R. , Jefcoate Colin R. , Boyd George S. , Brownie Alexander C. 
Published: March 2005

Cholesterol side‐chain cleavage has been studied in intact adrenal mitochondria from rats subjected to stress by ether anaesthesia as a means of raising the plasma adrenocorticotrophin levels. Control rats were either injected with cycloheximide or kept in a quiescent state. After initiation of cholesterol side‐chain cleavage by addition of isocitrate, pregnenolone formation from endogenous cholesterol in intact rat‐adrenal mitochondria follows a biphasic time‐course of formation with an initial rapid phase lasting 3–5 min followed by a much slower rate of formation.Pregnenolone formation from [4‐14C]cholesterol is linear if the tracer substrate is added to the mitochondria together with isocitrate. Preincubation of the mitochondria with [4‐14C]cholesterol prior to addition of isocitrate results in a biphasic [4‐14C]pregnenolone formation; the rate of the initial rapid phase depending on the duration of preincubation.The effect of stress is to increase 2–3 times the rate of pregnenolone formation in the initial phase compared to the rates observed in mitochondria from quiescent or cycloheximide‐treated rats.Depletion of cholesterol from adrenal mitochondria follows a similar pattern to pregnenolone formation, but the initial cholesterol content of the mitochondria is not affected by the pretreatment.On the basis of these results it is suggested that only a fraction of the total mitochondrial cholesterol is readily available for cholesterol side‐chain cleavage and that this pool is increased by stress.The cytochrome P450 type II spectral changes induced by addition of pregnenolone and isocitrate to the intact rat adrenal mitochondria have also been studied. The effect of stress was to increase pregnenolone‐induced difference spectrum in adrenal mitochondrial two‐ to three‐fold compared with cycloheximide‐treated animals. Similar results were found for the isocitrate‐induced type II difference spectrum.The spectral changes are interpreted to mean that stress causes an increase in the cholesterol complex of side‐chain cleavage cytochrome P450. Metabolism of the remainder of the cholesterol in the mitochondria is limited by the rate of transport, or binding, to this reactive centre. It is proposed that the acute effect of stress, mediated by adrenocorticotrophin, is to increase the proportion of mitochondrial cholesterol in the readily available form perhaps by an increase in the rate of transport or binding of cholesterol to sites from which it can be readily metabolised.
Chemical and Immunological Properties of Reduced and Alkylated Polypeptide Chains of Bovine Fibrinogen
Authors: Gollwitzer Rotraut , Timpl Rupert , Becker Udo , Furthmayr Heinz 
Published: March 2005

The α, β and γ chains were isolated from reduced and carboxymethylated bovine fibrinogen by chromatography on CM‐cellulose. Electrophoretically pure polypeptide chains could be obtained as judged by three different methods. The chains were soluble in buffers at or above pH 8 but exhibited non‐covalent aggregation. The molecular weights of the α, β and γ chains were estimated in dodecylsulfate‐polyacrylamide gel electrophoresis as 62000, 58000 and 48000, respectively. Each chain differed considerably from the other in amino acid composition as well as in the fragment pattern obtained after CNBr treatment. The occurrence of unresolved α/β material suggested microheterogeneity of the individual chains. Sulfitolysis yielded, at least for the α chain, less pure products.Between 20% and 54% of the antibodies in rabbit antisera to native fibrinogen precipitated with the α and/or β chain. Absorption studies, gel precipitation and hemagglutination‐inhibition suggested a considerable antigenic homology of both chains. They could be clearly distinguished from the γ chain which reacted poorly in precipitation test but considerably better in passive hemagglutination. Cyanogen bromide cleavage did not destroy the serologic activity of the chains. These findings indicated that a considerable number of the antigenic determinants on fibrinogen are not dependent on intact conformation. However, antibodies specifically reacting with the native molecule occur as well.
Intermediates in the Reaction Catalyzed by Glutamate Dehydrogenase
Authors: Antonini Eraldo , Chiancone Emilia , Yielding K. Lemone , Shafer Jules A. 
Published: March 2005

By studying the glutamate dehydrogenase catalyzed oxidation of glutamic acid under conditions where the concentration of enzyme exceeded that of NAD, it was possible to evaluate directly the first‐order rate constant (0.73 sec−1) for the decay of an intermediate associated with the burst in absorbance, which is seen at 340 nm in the pre‐steady state phase when the NAD concentration exceeded that of enzyme. A study of the wavelength dependence of the absorbance vs time plots when [enzyme] > [NAD] indicated that the intermediate associated with the burst is isosbestic with the ternary abortive complex E · Glu · NADH. This information made it possible to estimate the total amount of NADH (π) formed in the initial rapid phase when [NAD] > [enzyme].A study of the dependence of the pre‐steady‐state kinetics on [NAD] revealed that between 27 and 1000 μM NAD, π was invariant at 0.4 μmol NADH formed in the burst phase per polypeptide chain. However, the velocity of the phase immediately following the burst phase increased between 27 and 1000 μM NAD, and the pseudo‐first‐order rate constant for the burst phase increased between 30 and 300 μM NAD and decreased between 300 and 1000 μM NAD. These results were taken to mean that all the NAD sites on glutamate dehydrogenase are not catalytically equivalent and that binding of NAD to sites which are not catalytically active causes a decrease in the rate of the burst phase and an increase in the rate of the phase following the burst.Below 1000 μM glutamate, a pronounced lag in the onset of the burst phase is seen. This lag suggests that at low glutamate concentration binding of NAD and/or glutamate to the enzyme becomes slower than reduction of E · Glu · NAD.
Studies on the Biosynthesis of the Branched‐Chain Sugars from the Quinocycline Complex
Authors: Matern Ulrich , Grisebach Hans 
Published: March 2005

The incorporation of [2‐14C]pyruvate and [1‐14C]acetate into the 2,6‐dideoxy‐4‐C‐[1′‐hydroxy‐ethyl]‐l‐xylo‐hexopyranose (compound I) moiety of quinocycline A and into the 2,6‐dideoxy‐4‐C‐[1′‐oxoethyl]‐l‐xylo‐hexopyranose (compound II) portion of quinocycline B produced by Streptomyces aureofaciens was investigated. The methylglycosides of compounds I and II were subjected to periodate oxidation to yield, respectively, acetaldehyde and acetic acid from the branch carbon atoms. Acetaldehyde was oxidized to acetic acid, which was then degraded by Schmidt‐degradation. With [2‐14C]pyruvate 87 to 90% of the radioactivity incorporated into compound I or II was located in the C2 branch without randomization of the label. In contrast, with [1‐14C]acetate only very low activity was incorporated into the sugars and no 14C was localized in the C2 branch. These results prove that the C2 branch in compounds I and II originates from C‐2 and C‐3 of pyruvic acid.When 14C‐labelled quinocycline B was incubated for 17 h with S. aureofaciens, 13% was converted to quinocycline A. Under the same conditions only 2% of [14C]quinocycline A was converted to quinocycline B. It can therefore be concluded that quinocycline B is the precursor of quinocycline A. This result is in agreement with the assumption that hydroxyethylthiamine pyrophosphate is the immediate precursor for the oxoethyl group of the branched sugar II, which after transfer to the aglycone is reduced to compound I. Experiments with the thiamine antagonist neopyrithiamine did not show inhibition of incorporation of radioactivity from [2‐14C]‐pyruvate into the hydroxyethyl branch of compound I. However, this result does not exclude the possibility that thiamine pyrophosphate participates in the reaction.
Regulation of the Catabolic Qrnithine Carbamoyltransferase of Pseudomonas fluorescens
Authors: Stalon Victor , Ramos Fernando , Piérard André , Wiame Jean‐Marie 
Published: March 2005

The properties of the catabolic ornithine carbamoyltransferase of Pseudomonas fluorescens are compared to those of the anabolic transferase of the same organism. The comparison is also extended to a mutationally modified enzyme which, in contrast to the catabolic enzyme, is able to synthesize citrulline in vivo. All three activities are measured in the direction of citrulline synthesis.The catabolic ornithine carbamoyltransferase and the mutationally modified enzyme have an approximate molecular weight of 400000 while that of the anabolic enzyme is 100000.The catabolic transferase exhibits a number of regulatory properties which are not observed with the anabolic enzyme:

Its saturation by carbamyolphosphate is intensively cooperative. Its half saturation concentration is more than one hundred times higher than the Km of the anabolic enzymes; this explains its inability to perform an anabolic function in vivo.
Phosphate, a substrate in the physiological direction of the reaction and a low‐energy‐level indicator, acts as an activator of the enzyme; it abolishes the cooperativity of the carbamoylphosphate and increases the capacity of the enzymes.
ATP, a product of the catabolic pathway and a high‐energy‐level signal, reduces the capacity of the enzyme.
Putrescine, another product of the catabolic pathway, and its analogs, inhibit the enzyme by decreasing its apparent affinity for carbamoylphosphate.

The catabolic transferase of the mutant IRC204M1 shares all those properties with the wild‐type enzyme, but, owing to a reduced cooperativity of carbamoylphosphate, it is able to perform an anabolic function at reduced rate.Thus, the comparison of the regulatory properties of the catabolic ornithine carbamoyltransferase and of the modified enzyme provides a kinetic explanation for the inability of the catabolic enzyme to play an anabolic role and this without the need of referring to compartmentation of channeling of the anabolic and catabolic pathways.
Regulation of the Catabolic Ornithine Carbamoyltransferase of Pseudomonas fluorescens
Authors: Stalon Victor 
Published: March 2005

The catabolic ornithine carbamoyltransferase from Pseudomonas fluorescens catalyzes the phosphorolysis of citrulline leading to the formation of ornithine and carbamoylphosphate. The initial rates have been measured in the experimentally most accessible direction of the reaction, the carbamoylation of ornithine. The plots of reaction rates against carbamoylphosphate (substrate), phosphate (activator), and putrescine (inhibitor) are sigmoid shaped. The homotropic interactions between carbamoylphosphate molecules are decreased in the presence of phosphate and increased in the presence of putrescine.The allosteric properties of the enzyme are accounted for by the concerted transition theory proposed by Monod, Wyman and Changeux. The treatment of the kinetic data according to the method of Blangy, Buc and Monod suggests that the enzyme is an octamer made up of eight identical protomers which can exist in equilibrium in two conformational states which differ in their dissociation constant for the substrate, activator and inhibitor. The values of these constants for the binding of carbamoylphosphate, phosphate and putrescine, and the allosteric transition constant have been determined for the wild‐type enzyme and for the enzyme of a mutant which can use the catabolic enzyme for anabolic purpose.
Ein „Kurzschluß”‐Weg der Nicotinsäuresynthese bei Rhodotorula glutinis
Authors: Böttcher Fritz , Birnbaum Dieter , Samsonova Ida A. 
Published: March 2005

Nicotinic acid in Rhodotorula glutinis is synthesized from tryptophan via N'‐formylkynurenine, kynurenine, 3‐hydroxykynurenine and 3‐hydroxyanthranilic acid. Auxotrophic mutants, with blocked synthesis at various steps between tryptophan and 3‐hydroxyanthranilic acid, can grow with 100 μg/ml anthranilic acid (concentration in the agar‐agar). The usually required concentration of substances such as nicotinic acid or other substances after the blocked step is about 2 μg/ml. Auxotrophs with a block between 3‐hydroxyanthranilic acid and nicotinic acid are not able to grow with anthranilic acid. Therefore mutants requiring nicotinic acid with a blocked step between tryptophan and 3‐hydroxyanthranilic acid can synthesize nicotinic acid from anthranilic acid, a substance formed in the first step of tryptophan synthesis from chorismic acid, by eluding the blocked pathway.The same ability has been found in prototroph revertants with the original block of the auxotrophs but with a second mutation in the gene of anthranilate synthetase. As a result of this mutation the feedback inhibition by tryptophan is disturbed and anthranilic acid is produced in excess. When the culture medium of the prototrophs is analysed by paper chromatography after adding anthranilic acid to the growing culture, 3‐hydroxyanthranilic acid as one of the formed products can be found.Summarizing these facts it is evident, that there exists a “short‐cut”‐pathway of nicotinic acid synthesis in Rh. glutinis, which is realized by direct hydroxylation of anthranilic acid to 3‐hydroxyanthranilic acid. This “short‐cut”‐pathway however is only possible, when anthranilic acid in the cell is present in excess.
Fatty‐Acid Biosynthesis
Authors: Landriscina Clemente , Gnoni Gabriele V. , Quagliariello Ernesto 
Published: March 2005

The relationship between the two elongation syntheses of fatty acids, one present in microsomes and the other in mitochondria, and the system for synthesizing palmitate de novo has been studied in rat liver. The results indicate that microsomes are much more active than mitochondria in elongating and/or desaturating fatty acids synthesized by cytoplasmic system.
Fatty acids synthesized by cell sap in the presence of added microsomes are found principally esterified into phospholipids, this indicates that these compounds in the microsomal fraction are actively deacylated and reacylated.
ATP is required both for chain elongation and for the esterification of newly synthesized fatty acids.
Evidence has been obtained that acyl groups are elongated and/or esterified into complex lipids as acyl‐CoA and not as acyl‐ACP.
The high activity of palmityl thioesterase in the cell sap with respect to the other subcellular fractions points out the importance of this enzyme in the fatty acid synthesis de novo.
Fluorescence Studies on Substrate Binding to Seryl‐tRNA Synthetase from Yeast
Authors: Engel Günter , Heider Henning , Maelicke Alfred , von der Haar Friedrich , Cramer Friedrich 
Published: March 2005

The interactions of seryl‐tRNA synthetase with its ligands have been investigated by fluorescence measurements. Using a microscale fluorescence technique and an excitation wavelength of 303 nm the inner filter effect was suppressed.The fluorescence intensity of the seryl‐tRNA synthetase at 342 nm (maximum emission) was quenched by 33%, by adding tRNASer, whereas no effect was produced by the tRNAs which are specific for phenylalanine and tyrosine. For tRNASer an association constant of 2.1 × 106 M−1 per enzyme subunit was calculated from the fluorescence titration. The emission maximum of the synthetase did not shift when tRNA was added.Under similar conditions ATP caused only 6%, quenching of the enzyme fluorescence from which an intrinsic association constant of 105 M−1 for about 10 moles ATP per mole subunit could be derived. On the other hand, when the enzyme was saturated with tRNA, addition of ATP increased the fluorescence intensity. Performing the titration in the reverse order, the same fluorescence end point was achieved. The addition of CTP to the preformed complex between tRNASer and the synthetase did not induce any change in the fluorescence intensity.These results are discussed with respect to the binding process and binding sites at which tRNA and ATP are attached to the synthetase and the possible involvement of the tryptophan residues.
Partial Purification and Characterization of Glyoxalase I from Porcine Erythrocytes
Authors: Mannervik Bengt , Lindström Lars , Bártfai Tamás 
Published: March 2005

Glyoxalase I from porcine erythrocytes has been purified about 2000‐fold by ion‐exchange chromatography on SE‐Sephadex, CM‐cellulose and DEAE‐cellulose columns. The isoelectric point was estimated to be at about pH 5, and the molecular weight was 52000 as determined by gel filtration. The kinetics of the enzymatic reaction was studied and the Km value was determined to be 0.19 mM, which differs significantly from the Km value of the yeast glyoxalase I. The enzymatic activity is completely inhibited by the chelating agents EDTA and o‐phenanthroline, and activity is regained by addition of Mg2+, Mn2+, and Ca2+. The reactivation is time‐dependent, which indicates that a metal · enzyme complex is the catalytically active species. An apparent dissociation constant of 0.6 mM for the Mg2+· enzyme complex was determined.Notable differences between glyoxalase I from yeast and from porcine erythrocytes appear in molecular weight, kinetics, and reactivation of chelator‐inhibited enzyme.
The Binding of Steroid Hormones to Degranulated Microsomes from Rat‐Liver Endoplasmic Reticulum
Authors: Blyth Carol A. , Cooper Michael B. , Roobol Anne , Rabin Brian R. 
Published: March 2005

Smooth reticular membranes from male rat liver contain high affinity binding sites for oestradiol. Similar membranes from females contain analogous sites for testosterone.
Rough membranes from which polysomes have been removed by EDTA (degranulated membranes), contain no high affinity binding sites for either hormone.
Ethyl acetate extracts of degranulated membranes contain materials which block the high affinity binding sites on smooth membranes of the same but not the opposite sex.
These extracts promote the attachment of ribosomes to smooth membranes of the same but not the opposite sex.
The ethyl acetate extracts from male and female degranulated membranes contain substances which behave like oestradiol and testosterone, respectively.
Studies on Seed‐Oil Triglycerides
Authors: Gurr Michael I. , Blades Jon , Appleby Robert S. 
Published: March 2005

Until 10–12 days after flowering the lipid of Crumbé seeds accounts for less than 1% of the total seen weight. Most of this lipid is phospholipid and glycolipid, the fatty acid composition of which resembles that of green tissue.
After 10–12 days the oil content of the seed rises rapidly, due mainly to the synthesis of new triglyceride. Oil synthesis levels off at about 30 days when the oil comprises 35% of the weight of the seed.
The fatty acids characteristic of green tissue, C16:0, C18:2 and C18:3 decrease in proportion to the total fatty acids as the seeds mature. Erucic acid, which is not present before 6 days rises rapidly from 8 days to becomes the major seed oil fatty acid in the mature seeds (53%). Oleic acid and C22:1 increase in proportion reaching a maximum at 12 days
The triglycerides of the mature seed oil are composed of 5 major molecular species, having 2, 3, 3, 4 and 5 double bonds per molecule, respectively.
Position two is occupied entirely by C18 fatty acids with Δ9 unsaturation, namely C18:1, C18:2 and C18:3. No erucic acid occurs at position 2. The acids at position two are cleaved preferentially by the lipase from Geotrichum candidum.
Position 1 is occupied preferentially by saturated acids or erucic acid. Position 3 is occupied almost entirely by erucic acid.
Circular Dichroism of Avian‐Erythrocyte Chromatin and Ethidium Bromide Bound to Chromatin
Authors: Williams Ross E. , Lurquin Paul F. , Seligy Verner L. 
Published: March 2005

The circular dichroism of mature gander erythrocyte chromatin was compared with that of partially dehistonized chromatin and deproteinized DNA. The ellipticity at 275 nm for native chromatin was found to be three‐times less than that for DNA. The 275‐nm transition was gradually increased in magnitude upon removal of histones. Selective removal of the very lysine‐rich histone I and the erythrocyte‐specific histone V, but not histone I alone, resulted in a significant increase in transcription in vitro and ethidium bromide primary binding properties of chromatin. Concomitant with these changes a “blue shift” of the positive DNA band from 282 nm for native chromatin to 275 nm for purified DNA was observed.Optimal conditions for circular dichroism studies in ethidium bromide bound to DNA and native chromatin were determined.The circular dichroic spectra of ethidium bromide bound to native chromatin and partially deproteinized chromatin were found to be essentially the same as that exhibited by ethidium‐bromide · DNA complexes. However, the ellipticity of the dye bound to native chromatin was smaller than that when ethidium bromide was bound to DNA. The 310‐nm dye transition was increased after the removal of chromosomal proteins. Selective removal of histone I and V was found to generate the most notable increment in ellipticity.The represent results are consistent with recent interpretations on how ethidium bromide interacts with chromatin; native chromatin has fewer ethidium bromide primary binding sites than deproteinized DNA; the mode of this dye interaction in these two systems is nearly the same, if not identical.
Protein A from Staphylococcus aureus
Authors: Sjöholm Ingvar , Ekenäs Anna‐Karin , Sjöquist John 
Published: March 2005

Protein A, a cell wall protein from Staphylococcus aureus, has been chemically modified by acetylation with acetylimidazole. All the four tyrosines present in the molecule as well as 40 out of 52 lysines were acetylated. The conformation of protein A was not changed by the acetylation.The ability of protein A to react with the Fc fragment of immunoglobulins was decreased to 40%, but was restored to 60%, when acetylated protein A was O‐deacetylated with hydroxylamine. The reactivity of protein A against specific rabbit antiserum was not decreased by acetylation. The significance of unmodified tyrosines and lysines for the protein A reactivity is discussed.