bluuebunny/crossref_metadata_2025_split huggingface dataset:
Effect of Tamoxifen on Oestradiol and Progesterone‐Induced Synthesis of Ovalbumin and Conalbumin in Chick Oviduct
Authors: CATELLI Maria Grazia , BINART Nadine , ELKIK François , BAULIEU Etienne Emile 
Published: March 2005

Tamoxifen, a non‐steroidal antioestrogen, did not display any oestrogenic effects biochemically or histologically in the chick oviduct when 10 mg/kg were administered to oestrogen‐withdrawn animals each day and effects were monitored during periods ranging from a few hours to 10 days.After a single injection of oestradiol benzoate, 1 mg/kg, to oestrogen‐withdrawn animals, the relative rate of ovalbumin synthesis was significantly elevated by 3 h, and reached a maximum by ∼ 16 h. Conalbumin synthesis increased immediately after oestrogen administration, and attained a maximum by ∼ 16 h. Between 16 and 24 h there was a decrease of ∼ 50% in the rate of synthesis of the two proteins and in the levels of nuclear oestrogen receptor. Administration of tamoxifen together with oestradiol benzoate inhibited the oestrogen effect on ovalbumin and conalbumin synthesis. This effect of tamoxifen was dose‐dependent; 50% inhibition of the maximum induction of both proteins was obtained with 1 mg of tamoxifen/kg.A series of experiments indicated that tamoxifen given after oestradiol benzoate could rapidly inhibit the oestrogenic effect, and oestradiol benzoate administered subsequent to tamoxifen could overcome the antioestrogenic effect; that is, each ligand could produce its own characteristic activity, although in both cases, at the time of administration of the second drug, the level of cytoplasmic oestrogen receptor was low.When tamoxifen was administered during the lag period of ovalbumin induction, it decreased the entire pattern of ovalbumin synthesis, but when given later, the inhibitory effect was retarded. On the other hand, the inhibition by tamoxifen of oestradiol‐dependent conalbumin synthesis was virtually immediate. The basal level of conalbumin synthesis, already significant in withdrawn chickens (1%), was unaffected by tamoxifen, suggesting that endogenous oestrogens are not required to maintain this level of synthesis.Progesterone‐induced increases in ovalbumin and conalbumin synthesis were not inhibited by tamoxifen. On the contrary, the simultaneous injection of tamoxifen potentiated the progesterone‐increased synthesis of both proteins. These observations were supported by histological studies showing an increased accumulation of secretory granules in the magnum of chickens treated by tamoxifen plus progesterone as compared to progesterone alone.
Demonstration of Specific Receptors of the Rough Endoplasmic Membrane for the Signal Sequence of Carp Preproinsulin
Authors: PREHN Siegfried , TSAMALOUKAS Antonis , RAPOPORT Tom A. 
Published: March 2005

Evidence is given for the cotranslational processing in a wheat germ cell‐free system of carp preproinsulin to proinsulin by dog pancreatic rough endoplasmic membranes stripped of ribosomes by EDTA. Concomitantly with processing there is a translocation of the protein across the membrane as shown by the accessibility to proteases. Conversion of preproinsulin to proinsulin occurs post‐translationally only in the presence of detergent.
Stripped rough endoplasmic membranes bind a significant proportion of the cell‐free translation products synthesized in a wheat germ system in response to poly(A)‐rich RNA from carp islets. Among the bound material there is completed preproinsulin. The bound material is accessible to proteases. It is located on the cytoplasmic side of the membranes. Peptides lacking a signal sequence, such as globin or proinsulin, are not bound. The binding can be abolished by pretreatment of the rough membranes with proteases. Neither smooth endoplasmic membranes from dog pancreas nor erythrocyte plasma membranes from rabbits are able to bind signal peptides. The binding sites on the rough membranes can be saturated with cell‐free products coded by carp islet RNA and are then no longer able to process preproinsulin to proinsulin in a cotranslational assay. The amount of membranes necessary to bind a certain amount of cell‐free products post‐translationally corresponds roughly to the amount required for complete conversion of preproinsulin to proinsulin cotranslationally. The binding phenomenon occurs independently of the presence of ribosomes and is not sensitive to high ionic strength. A hydrophobic peptide (Ac‐Lys‐Phe‐Phe‐Gly‐Leu‐Nle‐NH2) did not compete with signal peptides.
The results show that stripped rough endoplasmic membranes possess specific proteincontaining receptors for signal sequences on their cytoplasmic side. The recognition of the signal by the endoplasmic membranes occurs without participation of the ribosomes. The latter are only required for the translocation of the nascent peptide into the lumen of the membrane.
Liver Glycogen Synthase in Rats with a Glycogen‐Storage Disorder
Authors: WATTS Colin , MALTHUS Rex S. 
Published: March 2005

1. A strain of rats with a genetically‐determined liver glycogen‐storage disorder (gsd/gsd) caused by a deficiency of liver phosphorylase kinase has a very high concentration of glycogen in the liver with a total glycogen synthase activity higher than in liver of fed normal animals, but only a small amount of the enzyme in the active form. In the livers of both starved and fed normal rats and gsd/gsd rats, there is a positive correlation between glycogen content and total glycogen synthase (r= 0.79) and an inverse correlation between glycogen content and active glycogen synthase (r= 0.86).2. Homogenates of livers from the gsd/gsd rats have lower glycogen‐synthase phosphatase activities than homogenates from fed normal rats which in turn have lower activities than those from starved rats. The addition of glycogen to homogenates of liver from starved rats reduced the glycogen‐synthase phosphatase activity.3. Dilution of homogenates of gsd/gsd rat liver produced an increase in glycogen‐synthase phosphatase activity which could be reversed by adding glycogen.4. The gsd/gsd rats were shown to have liver glycogen‐synthase phosphatase activity comparable to normal rats when the enzyme was measured in dilute homogenates from both groups with the same glycogen concentration.5. It is suggested that inhibition of glycogen‐synthase phosphatase by high glycogen concentrations is responsible for the low levels of activation of glycogen synthase in the gsd/gsd rats and that glycogen itself is probably the major factor controlling glycogen synthesis in gsd/gsd and fed normal rats.
Isomeric Incorporation of the Haem into Monomeric Haemoglobins of Chironomus thummi thummi
Authors: RIBBING Wilfried , RÜTERJANS Heinrich 
Published: March 2005

The Bohr effect in the monomeric haemoglobin component III of Chironomus thummi thummi is described on a molecular basis using various pH‐dependent proton resonances. One of the three titratable histidines changes its pK from 7.9 in the unligated state to 7.7 upon ligation with CO and up to 7.0 upon ligation with O2. It can be concluded from an analysis of the C‐2 proton resonance intensities that these shifts in pK occur in only one of the two conformational isomers of the component III, whereas in the second isomer these pK shifts are not observed. The differences between these functional properties are also derived from pH‐dependent changes of the mesoproton resonances as well as of the methyl resonances of Ile‐E11. The conformational change of the component III, which is connected with the Bohr effect, influences a variety of other resonances which could not be assigned. The pK values derived from the pH dependence of these changes agree exactly with the pK value of the Bohr proton donating group, which is supposed to be His‐G2. Also the corresponding signal intensities indicate the presence of two conformational isomers in the haemoglobin component III.Although the change in chemical shift of the His‐G2 C‐2 proton resonance is only small upon deprotonation of the imidazole ring, it has been shown with potentiometric and calorimetric methods that this histidine is deprotonated in this pH range.Considering the tertiary structure obtained from an X‐ray study of Steigemann and Weber we can readily explain the conformational changes in the Bohr active isomer of the component III. With binding of ligands dislocations of side chains at the proximal site produce a change in the interactions of His‐G2 with the carboxyl group of the C‐terminal methionine‐H22.
Synthesis of a Dicyclohexylcarbodiimide‐Binding Proteolipid by Isolated Pea Chloroplasts
Authors: DOHERTY Andrew , GRAY John C. 
Published: March 2005

1. A proteolipid (Mr 8000) has been purified to homogeneity from pea chloroplast membranes by extraction into butan‐1‐ol.2. The proteolipid from chloroplasts that had been incubated in the light with [35S]methionine or [3H]leucine gave a single radioactive band, coincident with the staining protein band, when analysed by polyacrylamide gel electrophoresis in the presence of sodium dodecylsulphate.3. In washed chloroplast membranes, or in the purified state, the proteolipid covalently bound [14C]dicyclohexylcarbodiimide. The reaction produced two new forms of the proteolipid, with decreased mobility in polyacrylamide gels containing dodecylsulphate.4. On analysis by dodecylsulphate/polyacrylamide gel electrophoresis, proteolipid purified from [35S]methionine‐labelled chloroplasts and treated with non‐radioactive dicyclohexylcarbodiimide gave a pattern of protein stain and radioactivity similar to that obtained when purified proteolipid was labelled with [14C]dicyclohexylcarbodiimide.5. Incorporation of radioactivity into the proteolipid, during incubation of chloroplasts with [35S]methionine, occurred in the light but not in the dark and was inhibited by D(–)threo‐chloramphenicol but not by cycloheximide or ribonuclease. We conclude that intact isolated chloroplasts from pea synthesise a dicyclohexylcarbodiimide‐binding proteolipid.
Comparison of the Nuclear Ribonucleoproteins Containing the Transcripts of Adenovirus‐2 and HeLa Cell DNA
Authors: GATTONI Renata , STEVENIN James , JACOB Monique 
Published: March 2005

A method was devised allowing the preparation of hnRNA‐containing ribonucleproteins (hnRNP) from HeLa cells infected with adenovirus type 2 under conditions where the extraction of viral replication complexes was minimal. Approximately 60% of the RNA from such hnRNP hybridized with adenovirus DNA. The hnRNP from infected cells had the same general characteristics as those from uninfected cells. Their size was heterogeneous (30–260 S) and depended upon that of their RNA. Their CsCl densities were identical (1.39–1.40 g/ml), indicating the same protein: RNA ratio. Their proteins were found in the same molecular weight range, between 25000 and 200000.The major proteins of hnRNP from HeLa cells were present in hnRNP from adenovirus‐infected cells. As 60% of the cellular RNA was replaced by adenovirus RNA in hnRNP, this indicated that there was not stringent specificity in the RNA‐protein interactions. The relative proportions of the proteins were identical in both cases, suggesting that the hnRNP assembly was independent of nucleotide sequences at least for the major proteins. The hnRNP from infected and uninfected cells differed by the size of their RNA, which was larger after infection, and by the presence of six additional minor polypeptides after infection. However, it cannot be excluded that the presence of these polypeptides in hnRNP resulted from non‐specific adsorption.
The Complex Formation between Escherichia coli Aminoacyl‐tRNA, Elongation Factor Tu and GTP
Authors: WAGNER Thomas , SPRINZL Mathias 
Published: March 2005

The interaction between Escherichia coli aminoacyl‐tRNAs and elongation factor Tu (EF‐Tu) · GTP was examined. Ternary complex formation with Phe‐tRNAPhe and Lys‐tRNALys was compared to that with the respective misaminoacylated Tyr‐tRNAPhe and Phe‐tRNALys. There was no pronounced difference in the efficiency of aminoacyl‐tRNA · EF‐Tu · GTP complex formation between Phe‐tRNAPhe and Tyr‐tRNAPhe. However, Phe‐tRNALys was bound preferentially to EF‐Tu · GTP as compared to Lys‐tRNALys. This was shown by the ability of EF‐Tu · GTP to prevent the hydrolysis of the aminoacyl ester linkage of the aminoacyl‐tRNA species. Furthermore, gel filtration of ternary complexes revealed that the complex formed with the misaminoacylated tRNALyS was also more stable than the one formed with the correctly aminoacylated tRNALys.Both misaminoacylated aminoacyl‐tRNA species could participate in the ribosomal peptide elongation reaction. Poly(U)‐directed synthesis of poly(Tyr) using Tyr‐tRNAPhe occurred to a comparable extent as the synthesis of poly(Phe) with Phe‐tRNAPhe. In the translation of poly(A) using native Lys‐tRNALys, poly(Lys) reached a lower level than poly(Phe) when Phe‐tRNALyS was used.It is concluded that the side‐chain of the amino acid linked to a tRNA affects the efficiency of the aminoacyl‐tRNA · EF‐Tu · GTP ternary complex formation.
Phosphorylation of Human Erythrocyte Pyruvate Kinase by Soluble Cyclic‐AMP‐Dependend Protein Kinases
Authors: MARIE Joelle , BUC Hélène , SIMON Marie‐Pierre , KAHN Axel 
Published: March 2005

Human red cell contain soluble adenosine‐3′,5′‐phosphate‐dependent protein kinases, which are able to phosphorylate the L′ subunits of erythrocyte pyruvate kinase. Efficiency and maximum level of phosphorylation are very comparable in human liver and red cells. Phosphorylation of red cell pyruvate kinase results in the same kinetic modifications as for liver enzyme, namely a shift towards a T′ allosteric state characterized by a decreased affinity for phosphoenolpyruvate and increased inhibition by the allosteric inhibitors ATP and alanine. In the course of red cell aging a small amount of partially proteolysed pyruvate kinase, devoid of the phosphorylatable site, appears; it resembles the subtilisin‐treated L′4 enzyme and accounts for less than 20% of total pyruvate kinase subunits.Endogenous phosphorylation of pyruvate kinase from erythrocytes incubated in the presence of cyclic nucleotides produces the same kinetic modifications as phosphorylation in partially purified extract; this, however, does not change glucose consumption, lactate production and glycolytic intermediate concentrations of the incubated cells.
Single‐Strand‐Specific Nuclease from the Nucleoplasm of Rye Germ Nuclei
Authors: PRZYKORSKA Anna , SZARKOWSKI Jan W. 
Published: March 2005

The localization of DNAase activity in rye germ nuclei revealed 48% of the total nuclear activity in the nucleoplasm and 4% in the chromatin.2. The endonuclease present in the nucleoplasm was purified more than 300‐fold as compared with the specific activity in homogenized nuclei by means of ammonium sulphate fractionation and CM‐cellulose and CM‐Sephadex column chromatography. The preparation gave a single band on polyacrylamide gels. Its molecular weight is 45000, pH optimum 5.4. The preparation does not exhibit activity of non‐specific phosphomonoesterase and phosphodiesterase.3. The nuclease catalyses the hydrolysis of denatured DNA and RNA to acid‐soluble 5′‐phoshate‐terminated products and cleaves the phosphomonoester linkage of 3′AMP.4. Synthetic polyribonucleotides are hydrolysed at a rate decreasing in the order: poly(U) > Poly(A) > Poly(C) > Poly(G).5. The endonuclease shows a high preference for single‐stranded nucleic acids. It does not depolymerise double‐stranded poly(U)· poly(A).6. The nuclease attacks covalently closed circular bacteriophage PM2 DNA, plasmid ColE1 and plasmid pBR322 DNA, converting it initially into the open‐circular (relaxed) form and subsequently into the linear form. This reaction is strongly dependent on ionic strength.7. The enzyme nicks the supercoiled DNA molecule of phage PM2 in only one place. At high enzyme concentrations several nicks in such a molecule are observed.
Energy‐Linked Reactions Catalyzed by the Purified ATPase Complex (F0F1) from Rhodospirillum rubrum Chromatophores
Authors: SCHNEIDER Erwin , SCHWULÉRA Udo , DOSE Klaus , FRIEDL Peter 
Published: March 2005

1. The isolation of the F0F1‐ATPase complex from Rhodospirillum rubrum chromatophores by the use of taurodeoxycholate is described.2. The enzyme preparation contains about 12 polypeptides; five are subunits of the F1 moiety.3. The ATPase activity of the purified enzyme is dependent on the addition of phospholipids.4. Km‐vales for Mg2+‐ATP and Ca2+‐ATP are similar to the values obtained for the membrane‐bound enzyme.5. The F0F1‐ATPase complex is more than 70% inhibited by oligomycin and N,N′‐dicyclohexyl‐carbodiimide.6. The F0F1‐ATPase complex was integrated into liposomes. The reconstituted proteoliposomes catalyzed energy transduction as shown by ATP‐dependent quenching of acridine dye fluorescence and ATP‐32Pi exchange.
Reconstitution of a Functional Acetylcholine Receptor
Authors: SOBEL André , HEIDMANN Thierry , CARTAUD Jean , CHANGEUX Jean‐Pierre 
Published: March 2005

The ‘reconstitution cycle’ is composed of the following sequence of operations. Highly purified receptor‐rich membranes prepared from Torpedo marmorata electric organ are exposed to pH 11 to remove the 43000‐Mr protein and dispersed into solution by sodium cholate under conditions where more than 85% of the receptor protein is in its 9‐S form. Elimination of the detergent by filtration on a Sephadex column (or dialysis) yields a ‘reconstituted receptor’ fraction, under conditions which conserve part of the endogenous lipids, or ‘reconstituted vesicles’ in the presence of an excess of exogenous lipids.The polypeptide composition of these fractions was analysed by sodium dodecylsulfate gel electrophoresis. Conditions are defined for quantitative measurements of the various polypeptide chains. The 40000‐Mr chain, which is labelled by the affinity reagent 4‐(N‐maleimido)phenyl [3H]trimethyl‐ammonium and therefore carries the acetylcholine receptor site, is the dominant polypeptide in the alkaline‐treated membranes and the reconstituted acetylcholine receptor.Electron microscopy discloses that many of the alkaline‐treated membranes no longer form closed vesicles and do not show the transverse asymmetry of the native membranes observed after tannic acid fixation. In the reconstituted receptor fractions, the receptor molecules reaggregate into discs and may be exposed on both faces of the discs. In the reconstituted vesicles, receptor rosettes are integrated to the lipid vesicles.With native membranes, the radioactive local anesthetic [3H]trimethisoquin binds to three classes of sites: non‐specific, low‐affinity and high‐affinity. Carbamylcholine causes an increase in the number of high‐affinity sites up to approximately 0.7 times the number of α‐125‐bungarotoxin sites. This ratio, the three classes of binding sites, and their regulation by carbamylcholine are conserved through the reconstitution cycle.
Reconstitution of a Functional Acetylcholine Receptor
Authors: HEIDMANN Thierry , SOBEL André , POPOT Jean‐Luc , CHANGEUX Jean‐Pierre 
Published: March 2005

The ‘functional’ state of the acetylcholine receptor protein has been followed during reconstitution with the fluorescent agonist [1‐(5‐dimethylaminonaplithalene)‐sulfonamido]‐n‐hexanoic acid‐β‐N‐trimethylammonium bromide ethyl ester (Dns‐C6‐Cho) and rapid‐mixing techniques. Under appropriate conditions, a majority of the acetylcholine receptor sites can be recovered in a low‐affinity state(s) for Dns‐C6‐Cho, similar to that found with the native membrane‐bound receptor. This state can be slowly interconverted to a high‐affinity state after rapid mixing with the agonist, and the non‐competitive channel blockers, like the local anesthetics, still regulate this transition in an allosteric manner.Several experimental conditions commonly used for the solubilization of the receptor and for its purification in the presence of sodium cholate result in the failure of reconstitution: the soluble receptor protein is stabilized in a low‐affinity state which can no longer be interconverted to a high‐affinity state in the presence of agonists or local anesthetics. On the other hand, it is demonstrated that if the concentration of lipids remains elevated in the presence of sodium cholate, a soluble (9 S) low‐affinity form of the receptor protein can be obtained which shows most of the characteristic properties of the membrane‐bound receptor and in particular the slow interconversion to the high‐affinity state and the effect of local anesthetics on this transition; furthermore, in these conditions the soluble protein can be manipulated ad libitum and submitted, for instance, to column filtrations and sucrose gradient centrifugations in the presence of detergent, without losing its characteristic conformational and allosteric transitions. After elimination of the detergent this form yields a reconstituted receptor which presents binding properties identical to those of the native membrane‐bound receptor and leads to the formation of vesicles which exhibit carbamylcholine‐sensitive ion fluxes.A necessary and sufficient condition for functional reconstitution is therefore the conservation, in the presence of lipids, of the allosteric properties of the receptor protein in its detergent‐soluble form.
Effect of Membrane Cholesterol on Action of Phospholipase A2 in Mycoplasma mycoides var. Capri
Authors: RIGAUD Jean‐Louis , LEBLANC Gérard 
Published: March 2005

Cells of Mycoplasma mycoides var. Capri grown in a medium containing 10 μg/ml cholesterol (native organisms) or in cholesterol‐free medium (adapted organisms) were treated with phospholipase A2. Hydrolysis of polar lipids (phosphatidylglycerol and diphosphatidylglycerol) only occurred in the adapted cells. Cholesterol replenishment of the membranes of these adapted cells in vitro which involves an increase from 7 μg to 66 μg cholesterol/mg membrane protein, completely abolished hydrolysis of polar lipid pools by phospholipase A2. This suggests that cholesterol incorporated either during growth or under conditions in vitro has an identical disposition and function in the membrane. This observation further indicates that cholesterol incorporation in M. mycoides var. Capri can be explained in terms of a simple physical adsorption process.Polar‐lipid breakdown products resulting from phospholipase A2 action on intact cells, isolated membranes and lipids extracted from adapted organisms were analyzed. In experiments with intact cells, [14C]oleic lysoderivatives but not [3H]palmitic lysoderivatives are accumulated within the membranes. In membrane preparations, again only [14C]oleic lysoderivatives are accumulated but transiently. Finally, both [14C]oleic and [3H]palmitic lysoderivatives were produced in phosphatidylglycerol‐diphosphatidylglycerol liposomal preparations. From these results it can be concluded that: (a) 80% of the phosphatidylglycerol and diphosphatidylglycerol have an unusual positional distribution of their fatty acid (unsaturated oleic acid in position 1) and (b) membranes of M. mycoides var. Capri contain an active lysophospholipase which more efficiently hydrolyzes palmitic‐acid‐containing lysoderivatives.