bluuebunny/crossref_metadata_2025_split huggingface dataset:
Lysozyme from the Insect Ceratitis capitata Eggs
Authors: FERNANDEZ‐SOUSA José M. , GAVILANES José G. , MUNICIO Angel M. , PEREZ‐ARANDA Agustin , RODRIGUEZ Rosalia 
Published: March 2005

Lysozyme from eggs of the Dipterous Ceratitis capitata (Wiedeman) has been purified by ionexchange chromatography and gel filtration and its physicochemical properties have been investigated. This is the first insect lysozyme characterized so far and it exhibits some properties different to those described for other animal lysozymes.
Lysozyme from the insect eggs has a molecular weight of about 23200 and a sedimentation coefficient of 2.4 S. Molecular weight determination by sodium dodecylsulphate gel electrophoresis indicates that the molecule consists of a single polypeptide chain.
This lysozyme preparation shows notable stability at acidic pH values and lability at alkaline pH values. It shows a single optimum pH at about 6.5.
Chitinase/muramidase specific activity ratio is around 350 times higher for the insect lysozyme than for the hen egg‐white enzyme.
The amino acid composition shows the presence of one tryptophan residue per molecule of enzyme. This fact differentiates the lysozyme from insect eggs from other animal and plant lysozymes. From the amino acid composition, the absorption coefficient and the partial specific volume are calculated.
Glycine is the N‐terminal residue.
Proton Translocation and ATP Formation Coupled to Electron Transport from H2O to the Primary Acceptor of Photosystem 2
Authors: BEN‐HAYYIM Gozal , NEUMANN Joseph 
Published: March 2005

The rate of electron transport from H2O to silicomolybdate in the presence of 3‐(3,4‐dichlorophenyl)‐1,1‐dimethylurea (diuron) (which involves the oxygen‐evolving enzyme, the photochemistry of photosystem 2 and the primary electron acceptor of photosystem 2) is controlled by internal pH. This is based on the shift of the pH profile of the rate of electron transport upon addition of uncouplers, or by using EDTA‐treated chloroplasts. Both stimulation and inhibition of electron transport by addition of uncouplers (depending on external pH) could be observed. These effects are obtained in the diuron‐insensitive photoreductions of either silicomolybdate or ferricyanide. These experiments provide strong evidence that a proton translocating site exists in the sequence of the electron transport H2O→ Q (the primary acceptor of photosystem 2).
The photoreduction of silicomolybdate in the presence of diuron causes the formation of Δφ. The value of Δφ depends on the external pH and its maximal value was shown to be 2.4. The calculated internal pH at different external pH values was found to be rather constant, namely between 5.1‐5.2.
Electron transport from H2O to silicomolybdate (in the presence of diuron) does not support ATP formation.It is suggested that this is due to the fact that the Δφ formed is below the “threshold”Δφ required for the synthesis of ATP. By adding an additional source of energy in the form of a dark diffusion potential created in the presence of K+ and valinomycin, significant amounts of ATP are formed in this system.
myo‐Inositol Transport in Klebsiella aerogenes
Authors: DESHUSSES Jacques , REBER Guido 
Published: March 2005

scyllo‐Inositol is transported against a concentration gradient by Klebsiella aerogenes, but is not metabolized.
The apparent KT of scyllo‐inositol is 0.05 × 10−3 M while that of myo‐inositol is 0.23 × 10−3 M. The V values are respectively 32 and 12 nmol min−1 mg cells−1 (dry weight).
Both cyclitols bind to the same carrier, since typical competitive inhibition kinetics are observed.
Neither phosphoenolpyruvate phosphotransferase nor a periplasmic binding protein seems to be involved in the scyllo‐inositol transport system.
The induction of the transport system is not repressed by malate, fumarate, citrate, succinate or pyruvate; however, acetate represses the induction. The activity of the transport system is increased when the growth medium is supplemented with adenosine 3′:5′‐monophosphate.
Transport of Cyclitols by a Proton Symport in Klebsiella aerogenes
Authors: REBER Guido , MERMOD Marion , DESHUSSES Jacques 
Published: March 2005

The respiration and the ATP content of Klebsiella aerogenes in the presence of various inhibitors were compared to the transport of scyllo‐inositol. The ATPase was found to be inhibited by dicyclohexyl carbodiimide. The transport has been tested in anaerobiosis and aerobiosis. From the results obtained it is concluded that either ATP or respiration can sustain the transport activity in independent manner.
The energy derived from the respiratory chain reactions or the ATP hydrolysis results in electrogenic extrusion of protons. The electrochemical potential created drives the accumulation of scyllo‐inositol, as shown by an increase of pH of the medium on addition of the substrate to cells in anaerobiosis. With non‐induced cells no change in pH occurs, which demonstrates that proton flow is really linked to the transport. No H+/Na+ or K+ exchange is observed and the proton conductor carbonylcyanide m‐chlorophenylhydrazone abolishes the pH shift caused by substrate addition. The stoichiometry of the symport H+/cyclitol is 1 and the half‐maximum value of the pH variation as a function of the amount of scyllo‐inositol added corresponds to a concentration of scyllo‐inositol very close to the KT of influx.
Kinetic Studies on a 4‐Methoxybenzoate O‐Demethylase from Pseudomonas putida
Authors: BERNHARDT Frithjof‐Hans , NASTAINCZYK Wolfgang , SEYDEWITZ Vera 
Published: March 2005

A direct, sensitive and reliable photometric assay procedure for monitoring the activity of nonspecific 4‐methoxybenzoate O‐demethylases of microorganisms is described. The assay is based on the O‐demethylation of 3‐nitro‐4‐methoxybenzoate to the yellow‐coloured product 3‐nitro‐4‐hydroxybenzoate. Using this assay and by monitoring the oxidation rate of reduced pyridine nucleotides, the kinetic properties of a purified, reconstituted enzyme system composed of 4‐methoxybenzoate monooxygenase (O‐demethylating) and a reductase from Pseudomonas putida have been investigated.It has been found that the Km value of the monooxygenase of this enzyme system towards different substrates (i.e. tight couplers, uncouplers and partial uncouplers) rises from the extremely low value of 0.07 μM for the tight couplers to about 55 μM for the uncouplers.The effect of possible inhibitors and metal ions on the reconstituted enzyme system was investigated. The inhibition pattern was almost identical to that found for the purified reductase, only batho‐phenanthrolinedisulfonate showing a greater inhibition of the reconstituted enzyme system.The affinity of the reductase towards NADH was found to be approximately 200‐fold greater than that towards NADPH. Furthermore, the affinity of this reductase to NADH depended on the nature of the electron acceptor. The affinity to NADH was more than 10 times higher when the monooxygenase. substrate complex was used as the electron acceptor, than when cytochrome c or 2,6‐dichloroindophenol was used. These differences are discussed on the basis of enzyme‐enzyme interactions between the reductase and the monooxygenase.
Formate Hydrogenlyase System in Salmonella typhimurium LT2
Authors: CHIPPAUX Marc , PASCAL Marie‐Claire , CASSE Francine 
Published: March 2005

Isolation from Salmonella typhimurium of mutants unable to reduce benzyl viologen under anaerobic conditions has allowed the study of the factors involved in the multienzymic formate hydrogenylase system.

Depending on the affected activities, different classes of mutants were found: FHL‐A mutants have lost formate dehydrogenase 1 and formate dehydrogenase 2 activities; mutations in fdhA (117 min) or fdhB (33 min) lead to such a phenotype. FHL‐B and FHL‐C mutants have lost formate dehydrogenase 2 activity and part or all of hydrogenase activity, respectively; both phenotypes correspond to mutations in the hyd gene (approximately 90 min). FHL‐D mutants have lost only formate dehydrogenase 2 activity; fhlD gene maps at 120 min.
In some cases, mixtures of extracts from two mutants display formate dehydrogenase 2 and formate hydrogenylase activities. Restoration studies suggest the existence of one factor sensitive to growth conditions and inactivated by oxygen or heating. This factor which is present and active in FHL‐C mutants, is probably the one missing in FHL‐D mutants.
A new scheme for the formate hydrogenylase system is proposed, in which hydrogenase transfers electrons directly to benzyl viologen.
On the Optical Activity of Ionized Tyrosyl Residues in Ovine Lutropin
Authors: ASCOLI Mario , WARD Darrell N. , JIRGENSONS Bruno 
Published: March 2005

The effect of alkali on the circular dichroic (CD) spectra of ovine lutropin and its subunits has been studied. Mild alkaline pH induces the appearance of a new optically active band in the 250‐nm region of the spectra of lutropin without any detectable alteration in the secondary structure of the protein. This change is reversible and can be correlated with ionization of 2–3 exposed tyrosyl residues in the intact hormone. In a previous report from this laboratory it was concluded that the three exposed tyrosyl residues are located in the α subunit, in positions 21, 92 and 93 [Burleigh, B. D., Liu, W.‐K. and Ward, D. N. (1976) J. Biol. Chem. 251, 308–315]. Nitration of these residues lowers the pH at which the intensity of the 250‐nm band is maximal.The importance of the tyrosyl residues of lutropin α (as opposed to those of lutropin β) is also supported by the similarity of the effect of alkali on the CD spectra of lutropin and lutropin α. Further evidence for this involvement was also obtained by a comparison of the alkali‐induced changes of refolded lutropin (α+β recombinant) and the product obtained by recombination of des‐(92–96)‐lutropin α (obtained from carboxypeptidase treatment of the α‐subunit) and lutropin β. The results indicate that removal of tyrosines α92 and α93 results in a decrease of the intensity of the 235‐nm band of ovine lutropin (at pH 7.5) as well as that of the 250‐nm band observed under alkaline conditions. It is therefore concluded that the 250‐nm band observed in alkaline solutions of lutropin arises (at least partially) from the red shift produced in the short‐wavelength optically active band of tyrosines α21, α92, and α93 upon ionization.
Phosphorylation in vitro and in vivo of Ribosomal Proteins from Saccharomyces cerevisiae
Authors: HÉBERT Josiane , PIERRE Michel , LOEB Jacques E. 
Published: March 2005

Crude ribosomes from Saccharomyces cerevisiae cultures were phosphorylated in vitro when incubated in the presence of [γ‐32P]ATP. Analysis of the ribosomal proteins with two‐dimensional electrophoresis revealed that of the 29 proteins identified in the small subunit, only protein S6 was phosphorylated. Of the 37 proteins identified in the large subunit, one was highly phosphorylated (L3) and two only slightly phosphorylated (L11 and L14). The protein kinase activity associated with the ribosomes was extracted with 1 MKC1 and was not dependent on adenosine 3′:5′‐monophosphate; it preferentially phosphorylated casein and phosvitin, but was less active on histones.Structural ribosomal proteins were also phosphorylated in vivo when the yeast cultures were incubated with [32P]orthophosphate; the radioactivity resistant to hydrolysis by hot perchloric acid was incorporated into the proteins of the two subunits. Radioactive phosphoserine was found by subjecting hydrolysates of ribosomal proteins to high‐voltage electrophoresis. After two‐dimensional electrophoresis, one poorly phosphorylated protein (S10) was identified in the small subunit. In the large subunit, one protein (L3) was highly labelled, and two proteins (L11 and L24) only slightly labelled.
Binding Studies of a Spin‐Labelled Oxidized Coenzyme to Bovine‐Liver Glutamate Dehydrogenase
Authors: ZANTEMA Alt , TROMMER Wolfgang E. , WENZEL Herbert , ROBILLARD George T. 
Published: March 2005

NAD+ with a nitroxide piperidine ring linked to the NH2 group of the adenine possesses full coenzymatic activity with glutamate dehydrogenase. Electron spin resonance spectra in the presence of glutamate dehydrogenase show mixtures of free and strongly immobilized spin‐label. Binding studies in phosphate buffer demonstrate: (a) weak binary binding to the enzyme with a dissociation constant in the order of 2mM; (b) an indication for negative cooperativity or different sites for binding to enzyme. 2‐oxoglutarate, with dissociation constants in the order of 20–250 μM; (c) similar but much weaker binding to enzyme. 2‐oxoglutarate. ADP; (d) a strong positive cooperative binding to enzyme. 2‐oxoglutarate. GTP, dependent on the enzyme concentration.Binding of phosphate to the enzyme with a Kd of about 20 mM or binding of pyrophosphate or tripolyphosphate with a Kd of about 2.5 mM enhances the binding of spin‐labelled NAD+ in the presence of 2‐oxoglutarate. There is evidence that the binding sites for these phosphates coincide with phosphate binding subsites of GTP.
The Stereochemical Basis of Template Function
Authors: RACKWITZ Hans Richard , SCHEIT Karl Heinz 
Published: March 2005

The behaviour of nucleotides with thioketo‐substituted pyrimidine bases (4‐thiouracil, 2‐thiouracil and 2‐thiocytosine) or amino‐analogue purine bases (2‐aminopurine and 2,6‐diaminopurine) in transcription and translation was investigated. The experimental results obtained led to the following conclusions.

The stereochemical basis of substrate selection in transcription is the geometry of Watson‐Crick base pairs A · U (or A · T) and G · C between substrate and template bases.
The topology of the active site of Escherichia coli RNA polymerase is precisely adopted to the geometry of Watson‐Crick base pairs.
The enzyme active site discriminates between A·U (A · T) and G · C base pairs. An essential feature in this discrimination is the 6‐NH2 group of the A · U (A · T) base pair and the 2‐keto group of cytosine in the G · C base pair.
The codon properties of a nucleic acid base in messenger RNA can be predicted on the basis of its specificity in polynucleotide interactions. There seems to be no evidence for the participation of protein topological sites in the control of the specificity of codon‐anticodon interactions in translation.
Purification and Control of Bovine Adrenal Cortical Cholesterol Ester Hydrolase and Evidence for the Activation of the Enzyme by a Phosphorylation
Authors: BECKETT Geoffrey J. , BOYD George S. 
Published: March 2005

A procedure for the purification of cholesterol ester hydrolase from bovine adrenal cortical 105000 ×g supernatant is described. Preincubation of a crude enzyme extract with [γ‐32P]ATP followed by purification resulted in the isolation of a phosphorylated preparation of cholesterol ester hydrolase. The phosphorylated cholesterol ester hydrolase appeared to be composed of 4 subunits, each having a molecular weight of 41000 ± 280 only one of which may be phosphorylated. Preincubation of the crude enzyme preparation with [α‐32P]ATP followed by purification did not produce a phosphorylated preparation of cholesterol ester hydrolase.Cyclic‐AMP‐dependent protein kinase, cyclic AMP, ATP and magnesium ions were required for activation of purified cholesterol ester hydrolase in vitro and the time course of activation closely paralleled the time course of phosphorylation of the enzyme. The addition of ATP, cyclic AMP and magnesium ions to the bovine adrenal cortical 105000 ×gsupernatant produced a 2.5‐fold stimulation in cholesterol ester hydrolase activity. This stimulation was abolished if protein kinase inhibitor was added prior to the addition of ATP, cyclic AMP and magnesium ions.The addition of magnesium ions or calcium ions to a crude preparation of cholesterol ester hydrolase was found to inhibit activity; however the same additions made to a purified preparation of cholesterol ester hydrolase were not inhibitory. The decrease in cholesterol ester hydrolase activity on incubation with magnesium ions was accompanied by a loss of 32P radioactivity from the protein. Preincubation of a crude preparation of cholesterol ester hydrolase with alkaline phosphatase resulted in a deactivation of cholesterol ester hydrolase.It is suggested that bovine adrenal cortex cholesterol ester hydrolase is activated by a phosphorylation catalysed by a cyclic‐AMP‐dependent protein kinase. Deactivation of cholesterol ester hydrolase is accomplished by dephosphorylation catalysed by a phosphoprotein phosphatase, dependent on magnesium or calcium ions.
Effect of 2‐Deoxy‐d‐glucose on the Cell‐Surface Glycoproteins of Hamster Fibroblasts
Authors: HUGHES R. Colin , MEAGER Anthony , NAIRN Roderick 
Published: March 2005

Growth of baby hamster kidney (BHK) cells in medium containing 2‐deoxy‐d‐glucose is retarded in direct proportion to the 2‐deoxyglucose concentration. The severity of the effect is reduced in medium containing high relative concentrations of glucose.
2‐Deoxyglucose inhibits the incorporation of radioactivity from mannose, galactose, glucosamine, fucose and N‐acetylmannosamine precursors into acid‐insoluble cellular material. Incorporation of radioactively labelled leucine into protein is not affected by 2‐deoxyglucose.
BHK cells grown in the presence of 2‐deoxyglucose become less sensitive to the toxic action of certain plant lectins, ricin of Ricinus communis and Phaseolus vulgaris phytohaemagglutinin, which bind specifically to cell surface galactose and N‐acetyl‐galactosamine residues. By contrast, 2‐deoxyglucose increased the sensitivity of BHK cells to the weak toxicity of concanavalin A which binds to surface mannosides. Treated cells also become more agglutinable with concanavalin A.
Cell surface glycoproteins labelled by lactoperoxidase‐catalysed iodination have been examined by dodecylsulphate‐polyacrylamide gel electrophoresis. The radio‐iodinated glycoproteins prepared from cells grown in medium containing 2‐deoxyglucose migrate more rapidly than glycoproteins from cells grown in the absence of inhibitor.
On the Role of Ferredoxin and Ferredoxin‐NADP+Reductase in Cyclic Electron Transport of Spinach Chloroplasts
Authors: BÖHME Herbert 
Published: March 2005

Antibodies prepared against purified spinach ferredoxin and ferredoxin‐NADP+reductase were used as specific inhibitors of electron‐transfer reactions dependent on either ferredoxin or ferredoxin‐NADP+reductase. The possible role of both electron carriers in cyclic electron transport was checked using cytochromeb6photoreactions as indicator. It could be demonstrated that the ferredoxin antibody inhibits cytochromeb6photoreduction. Ferredoxin‐NADP+reductase, however, appears not to be involved in this pathway: reductase antibody in concentrations sufficient to completely inhibit electron transport to NADP+had no effect on cytochromeb6photoreduction.Quantitative treatment of the immunoassay data showed that osmotically shocked chloroplasts contain both bound ferredoxin and ferredoxin‐NADP+reductase in concentrations approximately equal to that of cytochromeb6.
Biophysical and Enzymological Studies upon the Interaction of trans‐Cinnamic Acid with Higher Plant Microsomal Cytochromes P‐450
Authors: RICH Peter R. , LAMB Christopher J. 
Published: March 2005

The interaction of trans‐cinnamic acid with the cytochrome P‐450 of microsomes derived from washed potato slices has been studied. The washing process increased the specific content of microsomal electron transport components and hence provided a useful material in which to study the interaction.Evidence is presented that the trans‐cinnamic acid interacts with the cytochrome P‐450, and that this interaction is analogous to ‘type I’ interactions of other cytochrome P‐450 systems. This evidence includes the formation of a ‘type I’ substrate binding spectrum, an increased rate of reduction of cytochrome P‐450 by NADPH in the presence of trans‐cinnamic acid, an increased oxygen uptake and NADPH oxidation when trans‐cinnamic acid is added to the microsomes in the presence of NADPH, and a close correlation between biophysical parameters of electron transport in the cytochrome P‐450 system and enzymological parameters of the trans‐cinnamic acid 4‐hydroxylation reaction.The investigation has been extended to cytochrome P‐450 systems of other tissues and it has been found that the trans‐cinnamic acid 4‐hydroxylation reaction cannot account for the presence of most of the cytochrome P‐450 in several tissues. This suggests that other functions of higher plant cytochrome P‐450 chains exist, and that the substrate specificity of the hemoprotein may vary in different plant tissues.
Progesterone Receptors in the Chick Oviduct.
Authors: MESTER Jan , BAULIEU Etienne‐Emile 
Published: March 2005

Exchange techniques were developed for measurement of total progesterone receptor binding sites concentration in the cytosol and nuclei of chicken oviduct.
The level of progesterone receptor in the cytosol was under both oestrogen and progesterone control. Primary stimulation by oestradiol benzoate increased the receptor concentration from approximately 10000 sites/cell to approximately 40000 sites/cell in the magnum cytosol; subsequent withdrawal from oestrogen treatment led to a decrease to approximately 14000 sites/cell after 6 weeks. After progesterone administration (3 mg/kg) to oestrogen‐stimulated, withdrawn chicken, the receptor concentration decreased to approximately 40% of the initial level within the first 4 h; afterwards the receptor level rose again and by 40 h exceeded slightly the initial one.
The nuclear levels of the receptor reached a maximum at 1 h after the progesterone injection; however, the gain of the binding sites by the nuclei (∼ 900/nucleus) did not account for their loss from the cytosol. The maximum nuclear receptor level was not influenced by the dose of progesterone within range of 1‐10 mg/kg.
The extractibility of the nuclear progesterone receptor by 0.5 M NaCl was strongly influenced by preincubation of the nuclei at 30 C; irrespective of this preincubation, approximately 12% of the nuclear receptor resisted extraction with 2 M NaCl/5 M urea.
Differences between Alcohol Dehydeogenases
Authors: JÖRNVALL Hans 
Published: March 2005

Comparisons of the primary structures of yeast and horse liver alcohol dehydrogenases reveal that the enzymes are homologous but distantly related. The overall positional identity is 25% between common regions, and several deletions/insertions occur in either enzyme, the longest apparently corresponding to 21 residues, showing that the different subunit sizes are largely explained by internal differences.Variabilities in the structural similarities can be coupled with functional requirements but not directly with whole domains in the previously known tertiary structure of the horse protein. The two most similar regions of the enzymes affect active‐site segments and the two most dissimilar regions seem to affect a loop structure without known function, and a segment participating in subunit interactions. The dissimilarities may probably be correlated with changes in zinc‐binding properties and quaternary structures.The extra region corresponding to the large internal chain‐length difference shows an apparent coincidence in sequence to a following segment of the horse enzyme, and additional elements of internal coincidences, or superficial similarities with other dehydrogenases, are noticed. These characteristics are not fully distinguishable from chance distributions but in view of the extensive species variations in alcohol dehydrogenases some evolutionary considerations may not be excluded, in which case a model relating all regions of these and associated enzymes to a common ancestor is shown to be compatible with all known observations.