bluuebunny/crossref_metadata_2025_split huggingface dataset:
DNA‐Dependent RNA Polymerases of Thermoacidophilic Archaebacteria
Authors: PRANGISHVILLI David , ZILLIG Wolfram , GIERL Alfons , BIESERT Lothar , HOLZ Ingelore 
Published: July 2010

The component compositions of the DNA‐dependent RNA polymerases of the extremely thermophilic. anaerobic sulfur‐respiring archaebacteria Thermoproteus tenax and Desu/furococcus mucosus strongly resemble each other but also that of the RNA polymerase of Suljolobus acidocaldarius suggesting that both organisms belong to the same novel order Thermoproteales, which together with the order represented by Sulfolobus, forms the thermoacidophilic branch of archaebacteria.The component pattern of the RNA polymerase of T/iermoplasmu acidophilum, which does not belong to this branch, also appears homologuous. The archaebacterial type of the DNA‐dependent RNA polymerase is thus characterized by 9–10 components yielding a characteristic pattern which resembles that of yeast RNA polymerase A(1). In contrast to the α subunit of eubacterial RNA polymerases, the third largest component of archaebacterial RNA polymerases, although similar in size, is present only once per enzyme monomer.The polymerases of T. tenax and D. mucosus, like those previously isolated from other archaebacteria, are completely resistant against 100 μ/ml rifampicin and streptolydigin.The RNA polymerases of both organisms are highly thermostable. The enzyme from D. inucosus transcribes selectively and almost completely the H strand of phage T7 DNA.
Efficient Translation of the Coat Protein Cistron of Tobacco Mosaic Virus in a Cell‐Free System from Escherichia coli
Authors: GLOVER James F. , WILSON T. Michael A. 
Published: July 2010

Translation of tobacco mosaic virus (TMV) RNA in a cell‐free system derived from Escherichia coli (MRE 600) reveals several discrete polypeptides in the Mr range of 10000–50000. The major product is a polypeptide of Mr 17500 which comigrates with authentic TMV coat protein on sodium dodecyl sulphate/polyacrylamide gel electrophoresis. Structural investigations by peptide‐mapping techniques and differential radiolabelling confirm that the major product is TMV coat protein with an N‐terminal methionine. The major polypeptide product can be assembled in vitro into virus‐like ribonucleoprotein particles.The structural and evolutionary implications of this observation, and the values of TMV in elucidating eukaryotic mRNA interactions with the prokaryotic protein‐synthesizing machinery, are discussed.
‘Artificial’ Myosin Isozymes; Preparation and Characteristics
Authors: CARDINAUD Robert 
Published: July 2010

Using precisely monitored proteolytic digestion conditions rabbit fast skeletal muscle myosin could be selectively modified in different ways. A myosin isozyme with a 20‐kDa alkali light chain 1 (Al) could be obtained by digesting with papain in the presence of Ca2+. Under these conditions alkali light chain 2 (A2) was cleaved at Lys‐17 and lost a 2.3‐kDa N‐terminal fragment including the strongly basic N terminus and about half of the characteristic (Ala‐Pro) sequence. The Nbs2‐[5,5′ dithiobis(2‐nitrobenzoic acid)‐]light chain and A2 were left unmodified and less than 5% of the myosin heavy chain presented a break in the subfragment‐2 region. EDTA and Ca2+ ATPase activities were unchanged.A myosin isozyme with an 18‐kDa Nbs2‐light chain was obtained by limited digestion with trypsin in the presence of Ca2+. The 18.9→18‐kDa conversion was nearly 100% whereas less than 10% of the heavy chain was fragmented and only about 5% of A1 was converted to A1′. The Nbs2‐light chain was cleaved at Arg‐7 preserving Ser‐15 and consequently a phosphorylated modified myosin could be obtained. A quasi‐elastic light‐scattering study showed that this modified myosin in high‐ionic‐strength solutions exhibited physicochemical characteristics quite similar to those of unmodified myosin.
Biosynthesis of Vitamin B12 in Anaerobic Bacteria
Authors: LAMM Lydia , HECKMANN Gernot , RENZ Paul 
Published: July 2010

[1 ‐13C]Glycine, [2‐13C]glycine, and [15N]glycine were added to fermentations with the strictly anaerobic vitamin B12 producer Euhacterium limosum. The vitamin B12 isolated was almost exclusively labeled in its 5,6‐dimethylbenzimidazole moiety. It was methylated and degraded to 1,5,6‐trimethylbenzimidazole. The position of the 13C label was determined from the 13C NMR spectrum, using the natural‐abundance 13C NMR spectrum of 1,5,6‐triinethylbenzimidazole as reference. Thus it was found that C‐1 of glycine is incorporated into position 7a, C‐2 of glycine into position 3a of the 5,6‐dimethylbenzimidazole moiety of vitamin B12.In order to show whether the nitrogen atom of glycine is also incorporated, the vitamin B12 from the experiment with [15N]glycine was degraded to 5,6‐dimethylbenzimidazole. The H NMR spectrum of this compound exhibited in the region of the C‐2 proton a doublet from the 15N‐labeled part of the molecules (35 %) and a singlet from the unlabeled molecules. This doublet indicates that only one of the two nitrogen atoms of 5,b‐dimethyl‐ benzimidazole is labeled. From the incorporation pattern of 13C‐labeled glycine it must be deduced that this is N‐3 of the 5,6‐dimetliylbenzimidazole moiety of vitamin B12.These experiments reveal that one molecule of glycine is incorporated into 5,6‐dimethylbenzmidazole moiety of vitamin B12 in a regiospecific manner.
Direct Control of Glycogen Metabolism in the Perfused Rat Liver by the Sympathetic Innervation
Authors: HARTMANN Heinz , BECKH Karlheinz , JUNGERMANN Kurt 
Published: July 2010

The mode of action of hepatic nerves on the metabolism of carbohydrates was studied in the rat liver perfusedin sifu.

Electrical stimulation of the nerve bundles around the hepatic artery and the portal vein resulted in an increase of glucose and lactate output, an enhancement of phosphorylase a activity and a decrease of portal flow.
Sodium nitroprusside prevented the hemodynamic changes after nerve stimulation without affecting the metabolic alterations.
Phentolamine or an extracellular calcium level below 300 μmol × 1−1 abolished both hernodynamic and metabolic changes after nerve stimulation, while propranolol or atropine were without effect.
Norepinephrine infusion mimicked nerve stimulation only at the highly unphysiological concentration of 0.1 μM; it was not effective at a concentration of 0.01 μM, which might be reached in the sinusoidal blood due to an overflow from intrahepatic synapses.The present results suggest that, in rat liver, glycogen breakdown is regulated by α‐sympathetic nerves directly rather than indirectly via hemodynamic changes or via norepinephrine overflow.
Non‐ionic Adsorptive Immobilization of Proteins to Palmityl‐Substituted Sepharose 4B
Authors: NEMAT‐GORGANI Mohsen , KARIMIAN Khashayar 
Published: July 2010

Palmityl‐substituted Sepharose 4B prepared by the glycidyl either method of S. Hjertén et al. [J. Chromatogr. (1974) 101, 281–288] has been used as a non‐ionic matrix for protein adsorption. A number of proteins, some of which are catalytically active, may be immobilized on this adsorbent in the form of suspension or column.
Of the proteins examined, bovine serum albumin, hemoglobin, myoglobin, lysozyme, glutamate dehydrogenase, and β‐galactosidase were immobilized on the adsorbent irrespective of NaCl concentration. Trypsin, α‐chymotrypsin, papain, pepsin, and amyloglucosidase were totally adsorbed either in the absence of any additional salt or at high salt concentrations. Cytochrome c, used as a model protein, was totally immobilized only ar high ionic strength and low pH.
Immobilization normaly took place with an apparent in initial acitivity rates. In the case of trypsin using Nα‐benzoyl‐cl‐arginine p‐nitroanilide as substrate, adsorption resulted in anincrease in V (app).
Beef‐liver glutamate dehydrogenase, used as a model allosteric enzyme, was found to retain its allosteric properties towards ADP and GTP after immobilization.
Results are discussed in terms of specific interactions involving a smaller number of binding sites in protein molecules as compared to the multiple attachment to highly substituted adsorbents prepared with shorted ligands. Retention of the essential properties of the proteins examined in this study are ascribed to these characteristics of the adsorbent and to its non‐ionic nature. Relevance of these observations to in vivo processes and the potential use of the adsorbent for enzyme immobilization are also discussed.
The Interaction of the A and A* Proteins of Bacteriophage φX174 with Single‐Stranded and Double‐Stranded φX DNA in vitro
Authors: VAN DER ENDE Arie , LANGEVELD Simon A , VAN ARKEL Gerard A. , WEISBEEK Peter J 
Published: July 2010

The binding of the bacteriophage φX174‐coded A and A* proteins to single‐stranded (ssDNA) and double‐stranded (dsDNA) φX DNA was studied by electron microscopy. The interaction of the A* protein with ssDNA and dsDNA was also studied by sedimentation velocity centrifugation.It was shown that the binding of the A and A* proteins to ssDNA occurs in a non‐cooperative manner and requires no or very little sequence specificity under the conditions used here. Both protein‐ssDNA complexes have the same compact structure caused by intrastrand cross‐linking through the interaction of protein molecules with separate parts of the ssDNA molecule.The A protein does not bind to φX dsDNA in the absence of divalent cations. The A* protein does bind to dsDNA, although it has a strong preference for binding to ssDNA. The structure of the A* protein‐dsDNA complexes is different from that of the A* protein‐ssDNA complexes, as the former have a rosetre‐like structure caused by protein‐protein interactions. High ionic strengths favour the formation of large condenwi aggregates.
The Amino‐Acid Sequence of Histone H2A from Cuttlefish Sepia officinalis
Authors: WOUTERS‐TYROU Daniéle , MARTIN‐PONTHIEU Annie , BRIAND Gilbert , SAUTIERE Pierre , BISERTE Gérard 
Published: July 2010

The amino acid sequence of cuttlefish testis histone H2A (124 residues) was establijhed from structural data obtained by automated sequencing of large peptides generated by the cleavage of the protein with V8 staphylococcal protease or by limited chymotryptic hydrolysis.Compared to the calf thymus homologous histone, cuttlefish H2A shows 14 substirutions (most of them conservative) and 5 deletions.Extensive evolutionary changes were mainly observed in the basic amino‐terminal and carboxy‐terminal regions of the molecule, which are the primary DNA‐binding sites. Few punctual changes are observed in the central region (residues 18‐ 118), which interacts strongly with histone H2B to form the dimer H2A‐H2B.
The Specificity of Viral Sialidases
Authors: CORFIELD Anthony P. , WEMBER Margret , SCHAUER Roland , ROTT Rudolf 
Published: July 2010

The action of sialidases from Newcastle disease virus (NDV), influenzaA2 virus (IA2V) and fowl plague virus (FPV) on sialyloligosaccharide substrates containing α2 ‐ 3, α2 ‐ 6 or α2 ‐ 8 linkages was studied.
In all cases 2‐ 3‐linked sialic acids were preferentially released. Compared with II6Neu5AcLac, all 2–6‐ linked substrates, including sialyl‐N‐acetyllactosamine and its asparaginyl derivative, a urinary hexasaccharide and Neu5Ac(2–6)GalNAc were cleaved at improved rates by NDV and less by FPV sialidases. In the case of IA2V sialidase the asparaginyl oligosaccharide was very poorly cleaved, illustrating a variation in viral strain specificity.
A decrease in relative rates was observed in the order NDV >IA2V > FPV for substrates with 2–3 linkages relative to II6Neu5AcLac. The greatest relative rate was 470‐fold higher. The 2–3‐linked sialyl‐N‐acetyl‐lactosaminylasparagine and IV3Neu5AcLcOse4 were poor substrates for the IA2V sialidase, but the rates were greater than with the 2–6‐linked substrates.
The ganglioside substrate II3Neu5AcLacCer showed lower activity than its oligosaccharide analogue, but neither II3Neu5AcGgOse4Cer nor its oligosaccharide were substrates.
The Km values for 2–6‐linked substrates were generally of the order 10 mM while those for the 2–3‐ linked substrates were approximately 1 mM. The V values were consistently higher for the 2–3‐linked substrates. IV3Neu5AcLcOse4 showed high Km and very high V values, while the 2–8‐linked disialyllactose showed this trend only with NDV enzyme, the IA2V and FPV sialidases exhibiting high Km and low V values.
The results are discussed in the light of the current knowledge of viral sialidase specificity and relative to the binding of virus particles to cell surfaces.
A Cleavage Site of Ribonuclease F
Authors: GUREVITZ Michael , WATSON Ned , APIRION David 
Published: July 2010

A new endoribonuclease activity, RNase F, was partially purified from Escherichia coli cells. This activity can specifically cleave a precursor RNA molecule (of species 1) isolated from T4‐infected cells [N. Watson & D. Apirion (1981) Biochem. Biophys. Res. Commun. 103, 543–551], The cleavage results in products which are very similar to RNA molecules found in the cell, generating a 3′‐phosphate and a 5′‐hydroxyl groups. The cleavage takes place between a cytosine and an adenine moiety, within a possible loop and stem structure; the cut is in the border between the double‐stranded and single‐stranded regions of this structure.The specificity of this enzyme could be the introduction of a cleavage near the 3′ ends o f tRNA molecules and other RNAs like species 1 which could resemble tRNA in their three‐dimensional structure.
Quantification of Pathways of Glucose Utilization and Balance of Energy Metabolism of Rabbit Reticulocytes
Authors: SIEMS Werner , MÜLLER Marianne , DUMDEY Renate , HOLZHÜTTER Hermann‐Georg , RATHMANN Jörg , RAPOPORT Samuel Mitja 
Published: July 2010

In this work it is demonstrated that glucose constitutes the main substrate of energy metabolism of rabbit reticulocytes under aerobic conditions in the presence of 5 mM glucose. Amino acids and fatty acids are minor sources of energy. The shares of processes utilizing glucose in reticulocytes were estimated from tracer experiments. A new mathematical technique used permits the derivation of closed terms for the specific radioactivity of single positions of C atoms of the metabolites of the citrate cycle. By means of regression analysis, the undeter‐mined flux rates in the citrate cycle were calculated.On the basis of the data an overall balance sheet of glucose utilization and of ATP generation is given. About 45% of the glucose of reticulocytes is catabolized via the citrate cycle, about the same percentage yields lactate. Only 2% of the glucose was oxidized in the oxidative pentose pathway whereas the remainder is used for the formation of serine and glycine required for hemoglobin synthesis.These results are related to knowledge about the main processes utilizing ATP in reticulocytes, i.e. the synthesis of hemoglobin and the energy‐dependent proteolysis. Our approach to the investigation of metabolic relations in the reticulocytes can be applied to other tissues in which equilibria between large metabolite pools play a role
Rates of Flux through the Pentose Cycle in Perfused Rat Liver
Authors: KUEHN Alexander , SCHOLZ Roland 
Published: July 2010

A method for the determination of substrate flux through the pentose cycle was developed employing [l‐14C]glucose in experiments with perfused rat livers. The method consists first of a kinetic analysis which differentiates between the production of 14CO2 from [l‐14C]glucose via the pentose cycle and via the citrate cycle and, second of a calculation of the specific radioactivity of the hexose monophosphate pool from measured rates of glycolysis and the specific radioactivity of lactate released into the perfusate. The method was validated by experiments comparing the results of tracer infusions with [l‐14C]glucose, [6‐14C]glucose and [3‐14C]pyruvate.In livers from fed rats perfused with 10 mM glucose, the rate of substrate flux through the pentose cycle was around 0.2 μmol ×min−1× g−1; it was about 20% of the substrate flux via glycolysis. The kinetic data were inconsistent with the existence of an L‐type pentose cycle in liver.
Cucumber Mosaic Virus RNA 3
Authors: GOULD Allan R. , SYMONS Robert H. 
Published: July 2010

The complete sequence of the 2193 residues of RNA 3 (Mr 746000) of cucumber mosaic virus (Q strain) was determined by a combination of chemical and enzymic sequencing techniques utilizing cloned DNA fragments. The nucleotide sequence of RNA 3 also gave the complete sequence of 1027 residues of RNA 4 (Mr 349000), which codes for the viral coat protein and is derived from the 3′ end; there are 53 untranslated nucleotides at the 5′ end of RNA 4. The nucleotide sequence provided the amino acid sequences of the two proteins coded for by RNA 3: the 5′‐terminal 3A protein with 333 amino acids (Mr 36700) and the 3′‐terminal viral coat protein with 236 amino acids (Mr 26200). These two coding regions are in the same reading frame, are separated by an intercistronic region of 123 nucleotides and are flanked by two untranslated regions of 94 nucleotides at the 5′ terminus and of 263 nucleotides at the 3′ terminus.Secondary structure models are postulated for parts of the RNA 3 sequence. These are considered to be important in the control of the translation and replication of RNA 3 and in the processing of RNA 3 to give RNA 4.
A Transition‐State Analysis of the Enzyme Catalysis in the Affinity Labelling of Horse‐Liver Alcohol Dehydrogenase by Bromoacids
Authors: BRANDSNES Øystein , DAHL Knut H. , McKINLEY‐McKEE John S. 
Published: July 2010

The effect of temperature on the inactivation of liver alcohol dehydrogenase and on the alkylation of a model thiol free in solution by bromoacetate, 2‐bromopropionate, 3‐bromopropionate and 2‐bromo‐3‐(5‐imidazolyl)‐propionate has been studied and the thermodynamic activation parameters calculated.All the bromoacids had a favourable entropy of activation in reaction with the enzyme compared to the model reaction with the thiolate anion of cysteine. This results from the formation of a reversible bromoacid‐enzyme complex prior to the irreversible inactivation. The enthalpy of activation is however unfavourable, due to the lower intrinsic reactivity of the metal‐thiol in the enzyme, compared to the thiolate anion of cysteine.The difference in free energy of activation between the enzyme reaction and the model reaction was used to measure catalysis. The efficiency of the enzyme catalysis of alkylation increased in the order: bromoacetate < 2‐bromopropionate < 3‐bromopropionate < 2‐bromo‐3‐(5‐imidazolyl)propionate.Promotion by imidazole of enzyme inactivation by bromoacetate is a pure enthalpy effect. This is due to imidazole when bound to the active‐site metal improving the intrinsic reactivity of the metal‐thiol of Cys‐46.
Coordinate Regulation of Synthesis of Ribosomal Proteins during Encystation of Acanthamoeba castellanii
Authors: SCHULZE Ingrid , JANTZEN Helga 
Published: July 2010

The synthesis of ribosomal proteins has been examined during growth and encystment of Acanthamoeba castellanii. Cells have been radiolabelled with [35S]methionine and ribosomal proteins have been extracted either from ribosomes or from total cell extracts. The results show that there is less synthesis of ribosomal proteins relative to total cell proteins soon after transfer of growing cells into non‐nutrient medium, suggesting that the synthesis of all ribosomal proteins is under coordinate control. Furthermore, synthesis of ribosomal RNA and ribosomal proteins seems to be regulated coordinately, since within 1 h there is 70–80% decline in synthesis of both.Translation of total RNA in vitro indicates that the relative decrease in the synthesis of ribosomal proteins during development is not only due to a relative decrease in the content of ribosomal protein mRNA. However, a considerable (about 10‐fold) and coordinate decrease in the relative content of ribosomal protein mRNA occurs during encystment. These data suggest a difference in the control of the synthesis of total proteins and ribosomal proteins as well as in the regulation of the content of total mRNA and ribosomal protein mRNA.The comparison of patterns of ribosomal proteins indicates that during development no qualitative changes occur. The content of one protein of the small ribosomal subunit is low in growing cells and increases during encystation. Since the differences in the concentration of this protein are neither due to changing levels of the corresponding mRNA nor to changing rates of synthesis, changing affinities of this protein to ribosomes in vivo are suggested.
Adenosine Diphosphate Ribosylation of Chicken‐Erythrocyte Histones H1, H5 and High‐Mobility‐Group Proteins by Purified Calf‐Thymus Poly(adenosinediphosphate‐ribose) Polymerase
Authors: POIRIER Guy G. , NIEDERGANG Claude , CHAMPAGNE Madeleine , MAZEN Alice , MANDEL Paul 
Published: July 2010

Poly(ADP‐ribosylation) of histones H1, H5 and non‐histone chromosomal high‐mobility‐group proteins HMG 1, 2, 14 and 17 from chicken erythrocytes by purified calf thymus poly(ADP‐ribose) polymerase was studied using acid/urea/Triton gel electrophoresis and autoradiography.With histone H1, besides ADP‐ribosylated H1 supporting short chains of polymer, the appearance of H1 ‘dimer’ was observed and this reaction was dependent on NAD concentration and incubation time. In addition, highly modified and/or aggregated species of histone H1 were observed.Histone H5 was slightly ADP‐ribosylated at low NAD concentrations. At higher NAD concentrations or after longer incubations the formation of H5 ‘dimer’ and of more modified forms of H5 could be observed.HMG 1 and HMG 2 were found to be ADP‐ribosylated, the reaction being dependent on NAD concentration and time. Here again some discrete intermediates appeared. HMG 14 and HMG 17 were only slightly ADP‐ribosylated under our experimental conditions.These results indicate that the purified DNA‐independent poly(ADP‐ribose) polymerase can catalyse the formation of H1 ‘dimer’ as in nuclei and nucleosomes and that H5 and HMG proteins can also be ADP‐ribosylated and produce well‐defined higher complexes. These modifications of nuclear proteins may provide a means of localized conformational changes of the chromatin structure in vivo.
Methionyl‐tRNA Synthetase from Escherichia coli
Authors: BARKER David G. , EBEL Jean‐Pierre , JAKES Ross , BRUTON Chris J. 
Published: July 2010

A 3300‐base segment of Escherichia coli chromosomal DNA, cloned into pBR322, will complement a methionine auxotroph in which the lesion is a defective methionyl‐tRNA synthetase with a much reduced affinity for methionine. Crude extracts of these transformants contain elevated levels of a protein which has a subunit molecular weight of 66000, methionyl‐tRNA synthetase aminoacylation activity in vitro and which cross‐reacts with anti‐(methionyl‐tRNA synthetase) antibodies. This polypeptide is very slightly larger than the well‐characterised and crystallised tryptic fragment of methionyl‐tRNA synthetase.A DNA sequence of 1750 residues at one end of the cloned insert codes for a non‐terminated open reading frame in which we can locate a large number of methionyl‐tRNA synthetase tryptic and chymotryptic peptides. We have also sequenced 300 nucleotides upstream of this coding segment where we find a large invert repeat in the putative methionyl‐tRNA synthetase promoter region.