bluuebunny/crossref_metadata_2025 huggingface dataset:
Effect of insulin on glucose transporter distribution in white fat cells from hypophysectomized rats
Authors: Keller Susanne , Zapf Juergen
Published: July 2002

Glucose transport in white fat cells from hypophysectomized rats is increased and unresponsive to insulin. The goal of this study was to explain this observation. The number of glucose transporters, as determined by D‐glucose‐inhibitible cytochalasin B binding, in the plasma membranes from fat cells of hypophysectomized rats is: (1) elevated, (2) not increased by insulin, and (3) the same as in plasma membranes from insulin‐stimulated fat cells of control rats. In microsomal membranes from fat cells of hypophysectomized rats the number of glucose transporters is: (1) smaller than in basal and insulin‐stimulated fat cells from control rats, and (2) not changed by insulin.
Conserved residues of tumour necrosis factor and lymphotoxin constitute the framework of the trimeric structure
Authors: Tavernier Jan , van Ostade Xaveer , Hauquier Guido , Prange Thierry , Lasters Ignace , de Maeyer Marc , Lewit-Bentley Anita , Fourme Roger
Published: July 2002

Four distinct areas of primary sequence conservation between known tumour necrosis factor and lymphotoxin polypeptides from various species can be recognized. When these amino acid sequences are highlighted in the three‐dimensional structure, all are found in the same region, constituting the framework of the trimeric structure.
Electrophysiological expression of endothelin and angiotensin receptors in Xenopus oocytes injected with rat heart mRNA
Authors: Lory Philippe , Richard Sylvain , Rassendren François A. , Tiaho François , Nargeot Joël
Published: July 2002

Functional endothelin and angiotensin receptors have been expressed in Xenopus oocyte following the microinjection of rat heart mRNA. Under voltage clamp conditions, application of these peptides clearly induced oscillatory Ca2+‐activated chloride currents in a dose‐dependent manner. In addition, no direct modulation of expressed or native cardiac Ca channels was observed.
Crystallization of photosynthetic reaction centres from Chloroflexus aurantiacus
Authors: Yurkova E.V. , Tsygannik I.N. , Zargarov A.G. , Zolotarev A.S. , Abdulaev N.G. , Demin V.V.
Published: July 2002

Membrane protein crystallization; Reaction center; X‐ray; (Chloroflexus aurantiacus)
Site‐directed mutagenesis of the FAD‐binding histidine of 6‐hydroxy‐D‐nicotine oxidase
Authors: Mauch Ludwig , Bichler Veronika , Brandsch Roderich
Published: July 2002

6‐Hydroxy‐D‐nicotine oxidase; Site‐directed mutagenesis; Covalant flavinylation; Flavoenzyme; Covalent modification; Protein modification
Amino acid sequence of photosystem I subunit IV from the cyanobacterium Synechocystis PCC 6803
Authors: Rousseau Françoise , Lagoutte Bernard
Published: July 2002

We describe here the complete amino acid sequence of photosystem I subunit IV from Synechocystis 6803. The molecular mass of 8.0 kDa is lower than in higher plants and Chlamydomonas, due to the lack of a characteristic, proline‐rich, N‐terminal sequence. The remaining sequence exhibits a good conservation, with a hydrophilic and strongly basic N‐tenninal head followed by two hydrophobic domains. There is no possibility of classical membrane‐spanning alpha helices. This component is likely to be one of the most stroma accessible subunits of photosystem I.
Energy transfer via cell‐to‐cell junctions
Authors: Potapova T.V. , Aslanidi K.B. , Boitzova L.Ju.
Published: July 2002

Animal cell cooperation has been studied in a mixed cell culture. Membrane potentials of human embryonic cells and hamster BHK‐21 cells were recorded by intracellular microelectrodes. The Na+/K+‐ATPase inhibitor ouabain (1 × 10−6 M; 2 h) caused strong depolarisation of the human cells in pure culture. The same treatment reduced only slightly the membrane potential in the hamster cells in pure or mixed cultures, as well as in the human cells in mixed culture. The above data can be explained by effective ion fluxes through heterotypic gap junctions in mixed culture. Thus, in the presence of ouabain the Na+/K+‐ATPase of hamster cells creates transmembrane differences of the electrochemical potential of ions not only in the hamster cells, but in the human cells as well.
A nuclear‐encoded potato (Solanum tuberosum) mitochondrial tRNALeu and its cytosolic counterpart have identical nucleotide sequences
Authors: Maréchal-Drouard Laurence , Neuburger Michel , Guillemaut Pierre , Douce Roland , Weil Jacques-Henry , Dietrich André
Published: July 2002

Sequencing of potato mitochondrial (mt) tRNALeu(NAA) and of its cytosolic (cyt) counterpart revealed that these tRNAs are identical, except for a post‐transcriptional modification: a Gm is present at position 18 in mt tRNALeu, instead of a G in cyt tRNALeu. Hybridization studies have shown that potato mt tRNALeu(NAA) has a nuclear origin and must therefore be imported from the cytosol.
Formylation of tetrahydrofolate by formyl phosphate
Authors: Jahansouz Hossain , Scherübel Devin M. , Himes Richard H.
Published: July 2002

Formyl phosphate is the putative intermediate in the formylation of tetrahydrofolate (THF) catalyzed by N10‐formylTHF synthetase. In this study the non‐enzymic reaction between formyl phosphate and THF was examined at 5°C. 1H‐NMR, HPLC and kinetic analysis of the proton‐catalyzed conversion of the product to N5,10‐methenylTHF were used to identify the product. In contrast to the enzyme reaction, which produces N10‐form‐ylTHF, N5‐formylTHF was the only formylated THF derivative formed. The reaction was conducted at pH values of 3, 5, and 7, with the highest yield being obtained at pH 5 (64–85%, based on THF). The enzyme, therefore, changes the regioselectivity of this reaction by increasing the reactivity of the 10‐nitrogen and either decreasing the reactivity of the 5‐nitrogen or limiting its accessibility to formyl phosphate. 2‐Mercaptoethanol, present in the reaction mixture to protect THF from O2, was also formylated by formyl phosphate, at the oxygen position.
Biologically active transcripts of alfalfa mosaic virus RNA3
Authors: Dore Jean-Michel , Erny Claude , Pinck Lothaire
Published: July 2002

Transcripts of the bicistronic RNA3 of alfalfa mosaic virus were synthesized using the in vitro T7 run‐off transcription system. Synthetic RNA3 containing one additional G nucleotide at the 5' end were found to be infectious when coinoculated with RNA1 and RNA2 and coat protein.
Hydrodynamic data show that C1̄ inhibitor of complement forms compact complexes with C1̄r and C1̄s
Authors: Perkins Stephen J.
Published: July 2002

The C1̄ inhibitor of the complement cascade forms stoichiometric complexes with C1̄r and C1̄s and controls the activation of first component C1 of complement. Literature sedimentation coefficients s°20.w for the complexes formed between C1̄ inhibitor, C1̄r and C1̄s were analysed using frictional ratios and the hydrodynamic sphere approach. A head‐and‐tail two‐domain model for C1 inhibitor was combined with cylindrical hydrodynamic models for the six‐domain structures of C1̄r and C1̄s. The hydrodynamic data show that the heavily glycosylated N‐terminal domain of C1̄ inhibitor is positioned close to the two complement ‘short consensus repeat’ domains found in the centre of C1̄r and C1̄s.
Vectorial insulin secretion by pancreatic β‐cells
Authors: Cortizo Ana , Espinal Joseph , Hammonds Peter
Published: July 2002

Morphological studies of pancreatic β‐cells have suggested the presence of discrete sensory and secretory domains. In the present study we now provide functional evidence by demonstrating polarity of insulin release by HIT‐T15 cells. A significant diffusion barrier across a twin chamber culture system was verified in the presence of confluent HIT‐T15 cells. When stimulated with sulphonylurea, ionophore or high potassium, insulin was preferentially released into the lower chamber irrespective of whether secretagogues were added to the upper or lower chambers. Vectorial insulin secretion may be a significant determinant of islet hormone paracrine interactions in the maintenance of glucose homeostasis.
Haem binding to horse spleen ferritin
Authors: Kadir Fahmi H.A. , Moore Geoffrey R.
Published: July 2002

Horse spleen ferritin, a spherical protein shell of 24 subunits, contains no haem when extracted. This contrasts with ferritins isolated from bacterial sources which have the capacity to bind up to 24 haem groups [(1990) FEBS Lett. 271, 141–143] via two methionine residues [(1990) Nature 341, 771]. Here it is shown that horse spleen ferritin can bind between 15 and 17 haems per 24 subunits with an apparent association constant of 2.2–3.2 × 104 M−1. The strength of haem binding appears to be unaffected either by the presence of the core or by the oxidation state of the haem. The demonstration of the ability of animal ferritin to bind haem strengthens the similarity between it and bacterioferritin and could have major consequences for its mechanism of action in physiological iron uptake and release processes.
Expression in E.coli of the catalytic domain of rat poly(ADP‐ribose)polymerase
Authors: Thibodeau Jacques , Simonin Frédéric , Favazza Marisa , Gradwohl Gérard , Poirier Guy , de Murcia Gilbert
Published: July 2002

A 2 kilobase pair cDNA coding for the entire C‐terminal catalytic domain of rat poly(ADP‐ribose)polymerase has been expressed in E. coli. The overproduced 55 kDa polypeptide is active in synthesizing poly(ADP‐ribose) and the 4 kDa N‐terminal region of this domain is recognized by the monoclonal antibody C I,2 directed against the calf enzyme. Also, the minor αchymotrypsin cleavage site found in the human catalytic domain is not present in the rat enzyme as revealed by the absence of the 40 kDa specific degradation product in the E. coli cells expressing the rat domain. The expression of this partial rat cDNA should thus permit the rapid purification and subsequent crystallization of the catalytic domain of the enzyme.
A nucleotide sequence in the translation start signal region is involved in heat shock‐induced translation arrest in Escherichia coli
Authors: Kuriki Yoshitaka
Published: July 2002

In Escherichia coli synthesis of several proteins is transiently depressed upon heat shock treatment. A comparison of nucleotide sequences of the genes encoding these proteins revealed the occurrence of a consensus sequence, GAGGAA(N)3–6 ATG, in their translation start signal region. To examine whether this sequence is involved in heat shock‐induced depression of protein synthesis, DNA segments corresponding to this region of four of these genes, fusA, rpoB, glnS, and pheT, were synthesized, and each of them was fused in frame with the lacZ gene on the open reading frame vector pORFl. The effect of heat shock on the synthesis of β‐galactosidase encoded by these fused genes was then studied in E. coli. It was thus found that β‐galactosidase synthesis starting from the inserted translation start signal was arrested transiently upon temperature shift‐up from 30 to 42°C. I conclude that the heat shock‐induced depression of gene expression is an event taking place at the initiation of translation.
Replication‐linked histone acetylation in rat liver tissue is sensitive to alkylating agents
Authors: Talasz Heribert , Weiss Günter , Puschendorf Bernd
Published: July 2002

The effect of alkylating agents on histone acetyltransferase (EC 2.3.1.48) activity and thymidine incorporation was investigated in benign and malignant proliferating rat liver tissue and compared with the effect in normal non‐proliferating rat liver tissue. In both, benign and malignant proliferating tissue, but not in quiescent tissue, the histone acetylation is depressed by alkylating agents and this depression correlates with the inhibition of the thymidine incorporation. This effect suggests that the depression of the replication associated histone acetylation may be an important factor for the antiproliferative activity of alkylating agents.
Growth factor‐induced release of a glycosyl‐phosphatidylinositol (GPI)‐linked protein from the HEp‐2 human carcinoma cell line
Authors: Roberts Julia M. , Kenton Paul , Johnson Peter M.
Published: July 2002

The release of a GPI‐linked oncofetal protein (PLAP) from HEp‐2 cells has been studied as an indicator of endogenous PI‐specific PLC activation. A non‐hydrolysable GTP analogue, EGF and insulin all produced a dose‐dependent release of PLAP from these cells. This indicates that an increase in PLC activity following either growth factor binding or G protein activation may cause cellular release of PLAP.
Cooperativity between platelet‐activating factor and collagen in aggregation of bovine platelets III
Authors: Kojima Soichi , Sekiya Fujio , Inada Yuji , Sato Fumie , Tsukada Toshiyasu , Saito Yuji
Published: July 2002

In cooperative aggregation of bovine platelets induced by simultaneous addition of PAF and collagen at subthreshold concentrations the following observations were made. (i) Formation of phosphatidic acid and arachidonic acid metabolites, which characterize PAF and collagen alone‐induced aggregation, respectively, was observed very obviously. (ii) A thromboxane antagonist did not completely block the cooperative aggregation. The extent of residual aggregation activity was dependent on concentration of collagen used in the simultaneous administration with PAF. These results suggest that both signal transduction pathways activated by PAF and collagen alone at high concentrations are attained by simultaneous addition of both agonists at subthreshold concentrations through unknown mechanisms.
Estimation of adenosine triphosphate utilization of rat mast cells during and after anaphylactic histamine secretion
Authors: Johansen Torben
Published: July 2002

Determination of the cellular content of adenosine triphosphate (ATP) and the rate of ATP‐synthesis were used to estimate the cellular utilization ofATP in relation to anaphylactic histamine secretion. There was an increased rate of oxidative ATP‐synthesis and a decreased cellular ATP content during the time period of histamine secretion and immediately after its completion. During secretion the additional ATP‐utilization above the basal level of ATP‐synthesis was 0.51 cells. 2.5 min after cell activation, the rate of additional ATP‐utilization was 0.30
, and the persistent ATP‐decrease observed after 30 and 120 min may be due to a decreased rate of oxidative ATP‐synthesis.
Inactivation of thioredoxin by sulfite ions
Authors: Würfel Martina , Häberlein Ingo , Follmann Hartmut
Published: July 2002

Oxidized thioredoxin undergoes sulfitolysis of its single disulfide bond at low concentrations of sulfite ions and protein and in the absence of denaturing agents. The reaction, which has an optimum at pH 8, was studied using [35S]sulfite and E. coli thioredoxin as model. The product, thioredoxin‐S‐sulfonate, has a half‐life of several hours in solution. It is unable to activate chloroplast NADP malate dehydrogenase. Thioredoxin sulfitolysis may therefore be a physiologically important factor in mediating the phytotoxic effects ofsulfur dioxide in plants.
The primary structure of DNA binding protein II from the extreme thermophilic bacterium Thermus thermophilus
Authors: Zierer Rainer , Choli Dora
Published: July 2002

The primary structure of DNA binding protein II (DNA bp II) from the extreme thermophilic bacterium Thermus thermophilus has been established by combination of manual and automated techniques. The protein has 95 residues and a molecular mass of 11843. Comparison of the primary structure with the known sequence data of DNA bp II from Clostridium pasteurineum, Baccillus stearothermophilus, Escherichia coli, Rhizobium meliloti, Anabena, Thermoplasma acidophilum, Pseudomonas aeruginosa and Bacillus caldolyticus reveals a clear homology among these small basic proteins. In particular, two short sequences in the middle and C‐terminal part of the proteins (residues N‐Gly‐Phe‐Gly‐X‐Phe and Pro‐X‐Thr at positions 46–51 and 63–65, respectively) are completely conserved.
The influence of oligonucleotide‐effector on the selectivity of sequence specific modification of 16 S rRNA
Authors: Zenkova M.A. , Fedorova O.S. , Levina A.S. , Mamaev S.V. , Karpova G.G.
Published: July 2002

The influence of duplex stabilizing oligonucleotide‐effector (oligonucleotide, carrying N‐(2‐hydroxyethyl)phenazinium residues on both ends), on selectivity of site‐directed modification of E. coli 16 S rRNA (1542 nucleotides in length) under the conditions of its secondary structure stability was studied. The constant of cooperative binding of the reagent and the oligonucleotide‐effector with 16 S rRNA was determined. The accuracy of modification was shown to double in the presence of 50 μM effector at 5 μM concentration of the reagent.
Characterization of a chemically synthesized RNA having the sequence of the yeast initiator tRNAMet
Authors: Bratty John , Wu Taifeng , Nicoghosian Krikor , Ogilvie Kelvin K. , Perreault Jean-Pierre , Keith Gérard , Cedergren Robert
Published: July 2002

A 75‐unit long oligoribonucleotide corresponding to the sequence of the Saccharomyces cerevisiae initiator tRNA was synthesized chemically. The crude RNA was purified, and the sequence was verified by RNA sequencing techniques. A particularly useful purification step involved hydrophobic chromatography on BND‐cellulose. The purified RNA could be aminoacylated to 28% of a bona fide initiator tRNAMetsample and threonylated to 76 % of the level observed with native tRNAfMet from E. coli.
Prediction of arenavirus fusion peptides on the basis of computer analysis of envelope protein sequences
Authors: Glushakova Svetlana E. , Lukashevich Igor S. , Baratova Ludmila A.
Published: July 2002

Theoretical search and selection criteria for putative fusion peptides of enveloped viruses are proposed. Arena virus fusion peptides are predicted on the basis of computer‐assisted analysis of amino acid sequences of arenavirus envelope proteins and elements of their secondary and tertiary structure. Accordingly, two regions of GP2 surface protein from 5 viruses of Arenaviridae family have been detected with properties typical of fusion peptides of other enveloped viruses. One region, named peptide IV, located at the N‐terminus of the GP2 protein, is followed by the other region or peptide V, more likely candidate for the arenavirus fusion peptide.
Determination of tRNA nucleotide residues directly interacting with proteins in the post‐ and pretranslocated ribosomal complexes
Authors: Abdurashidova G.G. , Tsvetkova E.A. , Budowsky E.I.
Published: July 2002

Nucleotide residues of E.coli tRNA interacting directly with proteins in pre‐ and posttranslocated ribosomal complexes have been identified by analysis of photo‐induced tRNA‐protein cross‐links. A9, G 18, A26 and U59 residues of NAcPhePhe‐tRNA, located in the Ab‐site (pretranslocated complex) have been cross‐linked with proteins S10, L27, S7 and L2 respectively. In deacylated tRNA, located in the Pb‐site, residues C17, G44, C56 and U60 have been cross‐linked with proteins L2, L5, L27 and S9 respectively. The G44‐L5 cross‐link disappeared after translocation (NAc‐PhePhe‐tRNA located in the Pt‐site).
Transfection of DHFR− and DHFR+ mammalian cells using methotrexate‐resistant mutants of mouse dihydrofolate reductase
Authors: Thillet Joëlle , Pictet Raymond
Published: July 2002

Site‐directed mutagenesis was used to generate mutants of mouse dihydrofolate reductase more resistant to methotrexate than the wild type enzyme. The mutant genes were used to transfert either DHFR− or DHFR+ cell lines. These mutants, as well as the wild type gene, were able to confer methotrexate resistance to DHFR− CHO cells. The number of selected colonies decreased with increased concentrations of methotrexate. The number of colonies observed at 10 μM methotrexate is correlated with the Ki
(MTX) of the enzyme: the higher the Ki
, the higher the number of colonies for the corresponding mutant. In contrast, the transfections of DHFR+ cells gave a few numbers of colonies not different for the wild type and the mutants.
Cobalamin inactivation induces formyltetrahydrofolate synthetase
Authors: Deacon Rosemary , Perry Janet , Lumb M. , Chanarin I.
Published: July 2002

Loss of cobalamin function produces profound changes in the metabolism of formate. There is impaired synthesis of formyltetrahydropteroylglutamate synthetase (CHO‐H4,PteGlu), accumulation of endogenous formate and impaired utilization of [14C]formate. There are contradictory reports on the effect of cobalamin inactivation on CHO‐H4PteGlu synthetase. This study confirms a significant increase in synthetase activity following cobalamin inactivation.
Subcloning and nucleotide sequence of the 3,4‐dihydroxyphenylacetate (homoprotocatechuate) 2,3‐dioxygenase gene from Escherichia coli C
Authors: Roper David I. , Cooper Ronald A.
Published: July 2002

A cloned gene encoding the Escherichia coli C homoprotocatechuate (HPC) dioxygenase, an aromatic ring cleavage enzyme, was used to produce large amounts of the protein. Preparations of E. coli C HPC dioxygenase, whether expressed from the cloned gene or produced by the bacterium, lost activity very rapidly. The pure protein showed one type of subunit of M
1 33000. The first 21 N‐terminal amino acids were sequenced and the data used to confirm that the open reading frame of 831 bp, identified from the nucleotide sequence, encoded HPC dioxygenase. Comparison of the derived amino acid sequence with those of other extradiol and intradiol dioxygenases showed no obvious similarity to any of them.
Induction of differentiation of human leukemia cells by inhibitors of myosin light chain kinase
Authors: Makishima Makoto , Honma Yoshio , Hozumi Motoo , Sampi Kazumi , Hattori Masao , Motoyoshi Kazuo
Published: July 2002

Inhibitors of myosin light chain kinase, 1‐(5‐chloronaphthalene‐1‐sulfonyl)‐1H‐hexanydro‐1,4‐diazepine hydrochloride (ML‐9) and 1‐(5‐iodonaph‐thalene‐1‐sulfonyl)‐1 H‐hexahydro‐1,4‐diazepine hydrochloride (ML‐7), induced Nitroblue tetrazolium reducing activity, lysozyme activity and morphological maturation of human monoblastic U937, THP‐1 and promyelocytic HL‐60 cells, but not of erythroblastic K562 cells. However, three analogs of ML‐9, which are an inhibitor and an activator of protein kinase C, and a calmodulin antagonist, respectively, did not induce differentiation of the cells.
Effect of α‐ketoglutarate and its structural analogues on hysteretic properties of α‐ketoglutarate dehydrogenase
Authors: Bunik V.I. , Romash O.G. , Gomazkova V.S.
Published: July 2002

The burst of product accumulation during the KGD reaction was investigated. It has been shown not to be the obligatory feature of catalysis, but appears when increasing the enzyme saturation by KG. Structural analogues of KG and the SH‐group modification suppress the initial burst without preventing catalysis. The results obtained are in favour of the existence of the regulatory site for binding KG and its structural analogues essential for hysteretic properties of KGD.
Suicide process of renal cell carcinoma cells encountering mumps virus
Authors: Miki Shunji , Yamada Hidekazu , Orita Tomoaki , Yamamoto Masahiro , Miki Yoshitsugu
Published: July 2002

Renal cell carcinoma cells produced the substance(s) which killed them (suicide factor(s)) after co‐culture with mumps virus. The suicide factor(s) were heat‐sensitive and were degraded with trypsin. Furthermore, actinomycin D inhibited the production of the substance(s) by cancer cells. Considering these facts, the substance(s) were thought to be protein(s) derived from de novo synthesis in cancer cells. It was demonstrated that renal cell carcinoma cells proliferated with the autocrine loop of interleukin‐6 (IL‐6). Mumps virus almost completely inhibited the IL‐6 production in several hours. Because of these two facts, the suicide process might be initiated in renal cell carcinoma cells after encountering mumps virus, i.e. inhibition or the autocrine growth loop of IL‐6 followed by the induction of an autocrine killing loop of unknown substance(s).
A conformational switch involving the 915 region of Escherichia coli 16 S ribosomal RNA
Authors: Leclerc Daniel , Brakier-Gingras Léa
Published: July 2002

A novel alternative conformation, which involves an interaction between the 5′ terminal and 915 regions (E. coli numbering), is proposed after a screening of compiled sequences of small subunit ribosomal RNAs. This conformation contains a pseudoknot helix between residues 12–16 and 911–915, and its formation requires the partial melting of the 5′ terminal helix and the disruption of the 17–19/916–918 pseudoknot helix of the classical 16 S rRNA secondary structure. The alternate pseudoknot helix is proximal to the binding site of streptomycin and various mutations in rRNA which confer resistance to streptomycin have been located in each strund of the proposed helix. It is suggested that the presence of streptomycin favours the shift towards the alternate conformation, thereby stabilizing drug binding. Mutations which destabilize the novel pseudoknot helix would restrict the response to streptomycin.
The transport of glycolic acid by Chlamydomonas reinhardtii
Authors: Wilson Barbara J. , Tolbert N.E.
Published: July 2002

Evidence for a transport system for glycolate in Chlamydomonas was obtained, [14C]Glycolate was taken up rapidly reaching an equilibrium in less than 2 % at 4°C. Glycolate uptake was stimulated by valinomycin and high KCl or high KCl alone and inhibited by N‐ethylmaleimide. This uptake was not dependent on temperature or pH in contrast to uptake of benzoate by diffusion which decreased by orders of magnitude with increasing external pH. Based on these data, a transporter for glycolate is proposed.
The effect of F0 inhibitors on the Vibrio alginolyticus membrane ATPase
Authors: Capozza G. , Dmitriev O.Yu. , Krasnoselskaya I.A. , Papa S. , Skulachev V.P.
Published: July 2002

The inhibition of membrane ATPase from the marine alkalotolerant bacterium Vibrio alginolyticus by DCCD, triphenyltin and venturicidin was studied. DCCD proved to be an irreversible inhibitor, while venturicidin and triphenyltin produced a reversible inhibitory effect. The DCCD‐binding proteolipid was identified in the membrane preparations. The effect of the inhibitors on ATPase activity and ATP‐dependent Na+‐transport in V. alginolyticus subcellular vesicles is discussed.
Electrolyte induced vesiculation of MA782 mice mammary adenocarcinoma ascites cells
Authors: Zheng Tan , Shi-guo Fang , Shu-sen Liu
Published: July 2002

MA782 ascites cells were found to vesiculate when incubated in electrolyte solutions and release giant plasma membrane vesicles with diameters ranging from 5 μm to 15μm. The vesicles, generated from the plasma membrane and free of cellular organelles, as visualized by microscopy and marker‐enzyme analysis, could be proposed as a useful model membrane system.
What is the true ATPase activity of contracting myofibrils?
Authors: Houadjeto Maurice , Barman Tom , Travers Franck
Published: July 2002

The ATPase activity of contracting myofibrils was obtained in a solvent of ionic contents that mimic in vivo conditions: pH 7.4 and 0.1 M potassium acetate. Contracting myofibrils are fleeting structures and their ATPase activity is of short duration: even at 4°C it was over after a reaction time of 1 s and rapid reaction equipment was needed for its study. The ATPase (1.1 s−1) was very similar to that of crosslinked actoS1 (1 s−1). It was 100 × that of relaxed myofibrils.
Two types of linkage between codon usage and gene‐expression levels
Authors: Nakamura Takeshi , Suyama Akira , Wada Akiyoshi
Published: July 2002

The relation between codon usage and gene‐expression levels is an intensively investigated and discussed topic in the field of molecular evolution. We statistically analyzed 25 Escherichia coli gene sequences by a new classification of synonymous codons and found that (i) there are two distinct types of linkage between codon usage and gene‐expression levels in E. coli, and (ii) one of the two kinds of codon preferences (the codon preference concerned with interaction of GC/AT choice at three codon positions) is observed significantly in weakly expressed genes.
Chondroitinases release acetylcholinesterase from chick skeletal muscle
Authors: Pérez-Tur Jordi , Barat Ana , Ramos Milagros , Ramirez Galo
Published: July 2002

Bacterial chondroitinases (both ABC and AC types) release asymmetric and globular forms of AChE from chick skeletal muscle samples. Heparinases, however, including heparitinase I, fail to do so under different incubation conditions. These results do not support the direct implication of the heparin/heparan sulfate family of GAGs in the interaction of the different AChE molecular forms with the muscle ECM. GAGs of the chondroitin/dermatan sulfate group could however be involved, either directly or indirectly, in the attachment of the AChE collagen‐like tail to the muscle basal lamina.
Pentasialogangliosides of human brain
Authors: Miller-Podraza Halina , Månsson Jan-Eric , Svennerholm Lars
Published: July 2002

A pentasialoganglioside fraction of the ganglio series of glycosphingolipids has been isolated from human cerebellar cortex with a yield of 8.8 nmol NeuAc per g fresh tissue. The structural analysis showed that the material was a mixture of GP1b and GP1c gangliosides.
Augmentation of haptoglobin production in Hep3B cell line by a nuclear factor NF‐IL6
Authors: Natsuka Shunji , Isshiki Hiroshi , Akira Shizuo , Kishimoto Tadamitsu
Published: July 2002

The nuclear factor NF‐IL6 had been suggested to be responsible for the IL‐6‐mediated induction of several acute‐phase proteins. To obtain evidence for the involvement of NF‐IL6 in the induction of acute‐phase proteins, we introduced the NF‐IL6 gene and its truncated mutant (delNFIL6) gene into a hepatoma cell line Hep3B. Then, we examined the effect of the overproduced NF‐IL6 and delNFIL6 on the expression of haptoglobin, fibrinogen and albumin. As a result, basal production as well as induction of haptoglobin by IL‐6 were augmented by the expression of NF‐IL6, whereas delNFIL6 blocked the production of haptoglobin, fibrinogen and albumin.
Targeting of ricin A chain into pea chloroplasts
Authors: Roberts Lynne M. , Meadows Julie W. , Robinson Colin
Published: July 2002

Achimaeric gene was constructed encoding the pre‐sequence of the 33 kDa oxygen‐evolving complex protein from wheat (a thylakoid lumen protein) linked to ricin A chain. The fusion protein is efficiently imported by isolated pea chloroplasts and localised partly in the stroma, with the remainder bound to the stromal surface of the thylakoids. The imported protein is fully processed by both the stromal and thylakoidal processing peptidases, indicating that partial or complete translocation across the thylakoid membrane has taken place.
Impaired affinity for phenylalanine in Escherichia coli phenylalanyl‐tRNA synthetase mutant caused by Gly‐to‐Asp exchange in motif 2 of class II tRNA synthetases
Authors: Kast Peter , Wehrli Claudia , Hennecke Hauke
Published: July 2002

Phenylalanyl‐tRNA synthetase (PheRS: αβ2 subunit structure) is a member of class II of tRNA synthetases. We report here the genetic analysis of an Escherichia coli mutant strain which is auxotrophic for phenylalanine because it has a PheRS with a decreased affinity for phenylalanine. The mutant pheS gene encoding the PheRS α subunit was cloned and sequenced, and the deviation from the wild‐type gene was found to result in a Gly191‐to‐Asp191 exchange. This alteration is located within motif 2, one of 3 conserved sequence motifs characteristic for class II aminoacyl‐tRNA synthetases. Motif 2 may thus participate in the formation of the phenylalanine binding site in PheRS.
Correlation of apoproteins with the genes of the major chlorophyll a/b binding protein of photosystem II in Arabidopsis thaliana Confirmation for the presence of a third member of the LHC IIb gene family
Authors: Morishige Daryl T. , Thornber J.Philip
Published: July 2002

The major light‐harvesting complex in higher plants is LHC IIb. The LHC IIb of Arabidopsis thaliana contains 2 pigment‐binding apoproteins of 28 and 25 kDa. To determine the relationship between them and the LHC IIb gene family members, each protein was purified to homogeneity, subjected to direct protein sequencing, and the sequences compared with those deduced from LHC IIb genes in this organism. The 28 kDa protein is the product of Type I LHC IIb genes. The 25 kDa LHC IIb component is distinctly different from the 28 kDa LHC IIb protein, and is more closely related to the type III LHC IIb gene product of barley. Type III gene products lack the first 9–11 residues found in proteins of the Type I and II genes, a region that contains a phosphorylatable threonine residue. The lack of the N‐terminal residues explains why this LHC IIb apoprotein has never been seen to be phosphorylated, and partly or wholly why it is smaller. The implication of the missing N‐terminus on uptake of LHC II precursor proteins into the plastid and of the relative organization of the LHC IIb subunits in the PS II antenna is discussed.
Effects of 17‐hydroxywortmannin on serine/threonine‐protein kinases in human blood platelets
Authors: Kocher Markus , Clemetson Kenneth J.
Published: July 2002

Protein kinases are involved in signal transduction in human blood platelets. A number of renaturable protein kinases have increased in vitro activities compared to controls due to covalent modifications when intact platelets are activated by thrombin. The effect of the platelet inhibitor 17‐hydroxywortmannin (HWT) on these protein kinases was investigated in intact platelets and in vitro. HWT inhibits the increase in activity of these protein kinases but it does not interact directly with their catalytic subunits. We conclude that HWT blocks a step in signal transduction which is necessary to activate these protein kinases via covalent modifications.
Identification of the phosphorylation sites of H2B histone by a catalytic fragment of p72syk from porcine spleen
Authors: Sakai Keiko , Tanaka Yukie , Asahi Momoyo , Shimomura Reiko , Taniguchi Takanobu , Hashimoto Eikichi , Yamamura Hirohei
Published: July 2002

Phosphorylated sites of calf thymus H2B histone were investigated with a catalytic fragment of 72 kDa protein‐tyrosine kinase (p72
syk
). Three of five tyrosine residues in H2B histone can be phosphorylated by this kinase. In this analysis, H2B histone was thoroughly phosphorylated in vitro with [γ‐32P]ATP and the kinase, and then digested with a lysylendopeptidase. The resulting radioactive phosphopeptides were separated by a reverse‐phase column on high performance liquid chromatography. Subsequent sequential Edman degradation of the purified phosphopeptides revealed that 40Y, 83Y and 121Y were phosphorylated. 121Y is the major phosphorylated residue in H2B histone. No phosphorylation was detected in 37Y and 42Y. Although the consensus sequence was not defined from these analyses, our data suggest that higher‐order structure(s) in addition to primary one may participate in recognition of H2B histone by this protein kinase.
Interaction of cholesterol‐conjugated alkylating oligonucleotide derivatives with cellular biopolymers
Authors: Ryte A.S. , Karamyshev V.N. , Nechaeva M.V. , Guskova Z.V. , Ivanova E.M. , Zarytova V.F. , Vlassov V.V.
Published: July 2002

Interactions of oligonucleotide derivatives with mammalian cells and cellular biopolymers have been investigated. The derivatives were oligonucleotides bearing an alkylating 2‐chloroethylamino group at the 3′‐end and a cholesterol residue at the 5′‐terminal phosphate. These compounds are readily taken up by cells and react with cellular DNA, RNA and some proteins which may play a role in delivery of the compounds into cells.
Rapid agonist‐induced beta‐adrenergic receptor kinase translocation in C6 glioma cells
Authors: Garcia-Higuera Irene , Mayor Federico
Published: July 2002

Exposure of C6 glioma cells to 1 μM isoproterenol leads to fast desensitization of the β‐adrenergic receptor/adenylyl cyclase system and transient receptor sequestration. It also triggers a very rapid and transient translocation to the plasma membrane or β‐adrenergic receptor kinase (βARK), a specific cytoplasmic kinase that phosphorylates only the agonist‐occupied form of several G protein‐coupled receptors. βARK‐mediated receptor phosphorylation appears to be a suitable mechanism for the rapid regulation or adrenergic receptor function in the nervous tissue.
Proton NMR of the interaction of benzo(a)pyrene with rat liver microsomal cytochrome P‐450
Authors: Castro-Maderal Luz , Sullivan Paul D.
Published: July 2002

1H nuclear magnetic resonance longitudinal relaxation time (T
1) measurements were used to study the interaction of 1,3,6‐H‐benzo(a)pyrone (1,3,6‐H‐BaP) with microsomal cytochrome P‐450 from livers of phenobarbital‐ and β‐naphthoflavone‐treated rats. Upon addition of various amounts of cytochrome P‐450 to solutions of BaP, the T
1 values for the three BaP protons decreased. When the hemeprotein was converted to its carbon monoxyferrous derivative in the presence of BaP, the T
1 values of the BaP protons increased. The paramagnetic contributions to the observed T
1 values were calculated and resulted in distance estimates of > 6 Å for the three BaP protons in both cases.
2‐Aminopurine inhibits RNA and protein synthesis and reduces catecholamine desensitization in C6‐2B rat glioma cells
Authors: DeBernardi Maria A. , Brooker Gary
Published: July 2002

We previously proposed that intracellular cyclic AMP accumulation induces a putative, rapidly turning over protein inhibitory to further hormone activation of adenylate cyclase. In the present study, 2‐aminopurine, which has been reported to selectively block c‐fos gene expression, was used to test the hypothesis that c‐fos protein might be involved in the desensitization process. Indeed, a reduction in heterologous desensitization to catecholamines was observed in 2‐aminopurine‐treated C6‐2B rat glioma cells. However, we found 2‐aminopurine to inhibit, in a concentration‐dependent manner, total cellular RNA and protein synthesis in C6‐2B. HeLa, Swiss 3T3 and BALB/e cells. mRNA synthesis was also markedly reduced in 2‐aminopurine‐treated cells. These unexpected findings, while supporting our hypothesis of a protein synthesis‐sensitive step in the development of refractoriness, raise concern about the specificity of action of 2‐aminopurine to inhibit c‐fos induction and thus any cellular process, including desensitization, which might be regulated by c‐fos gene expression.
Primary structure of three minor isoforms of amphioxus sarcoplasmic calcium‐binding proteins
Authors: Takagi Takashi , Valette-Talbi Laurence , Cox Jos A.
Published: July 2002

Previously we reported the amino acid sequences of 4 well‐defined sarcoplasmic, high‐affinity Ca2+‐binding proteins in the protochordate amphioxus, Branchiostoma lanceolatum [1]. Here we report on the complete amino acid sequence determination of 3 additional minor isoforms. The seven isoforms differ from each other in 9 positions of a contiguous 17‐residue‐long segment (positions 20–36) and can be classified in a α (ASCP I, III and IV) and a β lineage (ASCP II, V, VI and VII).
The polypeptide chain fold in tyrosine phenol‐lyase, a pyridoxal‐5′‐phosphate‐dependent enzyme
Authors: Antson Alfred A. , Strokopytov Boris V. , Murshudov Garib N. , Isupov Michail N. , Harutyunyan Emil H. , Demidkina Tatyana V. , Vassylyev Dmitry G. , Dauter Zbigniew , Terry Howard , Wilson Keith S.
Published: July 2002

The tyrosine phenol lyase (EC 4.1.99.2) from Citrobacter intermedius has been crystallised in the apo form by vapour diffusion. The space group is P21212. The unit cell has dimensions a = 76.0 Å, b = 138.3 Å, c = 93.5 Å and it contains two subunits of the tetrameric molecule in the asymmetric unit, Diffraction data for the native enzyme and two heavy atom derivatives have been collected with synchrotron radiation and an image plate scanners. The structure has been solved at 2.7 Å resolution by isomorphous replacement with subsequent modification of the phases by averaging the density around the non‐crystallographic symmetry axis. The electron density maps clearly show the relative orientation of the subunits and most of the trace of the polypeptide chain. Each subunit consists of two domains. The topology of the large domain appears to be similar to that of the aminotransferases.
Heterogeneous expression of four MAP kinase isoforms in human tissues
Authors: Gonzalez Fernando A. , Raden David L. , Rigby Mark R. , Davis Roger J.
Published: July 2002

Mitogen‐activated protein kinases (MAP kinases) are a group of closely related enzymes implicated in signal transduction pathways. We report the molecular cloning of four human proteins (p40
mapk
, p41
mapk
, p44
mapk
and p63
mapk
, with high homology to members of the MAP kinase family. Sequence analysis demonstrated that p44
mapk
and p63
mapk
were the products of distinct genes. However, the p40
mapk
and p41
mapk
were found to be related, and are likely to result from alternative processing of transcripts from a single gene. The heterogeneous expression of these human MAP kinase isoforms in different tissues may reflect the diversity of signal transduction pathways in differentiated cells.
Abolishment of bradykinin‐induced calcium oscillations in ras‐transformed fibroblasts by the expression of 80 kDa diacylglycerol kinase
Authors: Fu Tao , Sugimoto Yoshikazu , Okano Yukio , Kanoh Hideo , Nozawa Yoshinori
Published: July 2002

Our previous study showed bradykinin‐induced periodic Ca2+ changes (Ca2+ oscillations) in v‐Ki‐ras‐transformed NIH/3T3 (DT) cells in which protein kinase C (PKC) activity is partially down‐regulated by a sustained high level of 1,2‐diacylglycerol (DAG) [FEBS Lett. (1991) 281. 263–266].In the present study, DAG kinase with 80 kDa mass (80K DGK) has been successfully transfected in DT cells, which exhibited enhanced cellular DAG kinase activities, decreased cellular DAG contents, and increased PKC activities compared to the control vector‐transfected cells. Furthermore, these DGK‐transfectants showed strong inhibition in bradykinin‐induced Ca2+ oscillations. The results suggest that the sustained DAG increase down‐regulates the PKC activity, thereby leading to the induction of Ca2+ oscillations in DT cells.
Differences in the thermostabilities of barley (1→3,1→4)‐β‐glucanases are only partly determined by N‐glycosylation
Authors: Doan Danny N.P. , Fincher Geoffrey B.
Published: July 2002

Barley (1→3,1→4)‐β‐glucan 4‐glucanohydrolase (EC 3.2.1.73) isoenzyme EII carries 4% by weight carbohydrate and is more stable at elevated temperatures than isoenzyme EI, which has no associated carbohydrate. The relationship between carbohydrate content and thermostability has been investigated by treatment of the two isoenzymes with N‐glycopeptidase F (EC 3.5.1.52). Removal of carbohydrate from isoenzyme EII results in a decrease in the enzyme's thermostability, but treatment of isoenzyme EI with the N‐glycopeptidase F has no effect. In addition, removal of a single N‐glycosylation site in isoenzyme EII (Asn190‐Ala‐Ser) by site‐directed mutagenesis of the corresponding cDNA led to a reduction in thermostability, while the introduction of this site into isoenzyme EI enhanced stability. We conclude that N‐glycosylation of Asn190 enhances the stability of isoenzyme EII at elevated temperatures, but that other factors related to their primary structures also contribute to the differences in thermostabilities of the barley (1→3,1→4)‐β‐glucanases.
Direct RNA polymerase chain reaction for TMV detection in crude cell extracts
Authors: Drygin Yuri F. , Korotaeva Stalina G. , Dorokhov Yuri L.
Published: July 2002

A part of the 30,000 bp transport protein gene of tobacco mosaic virus (TMV) RNA was amplified via direct RNA PCR and via traditional reverse transcription followed by cDNA PCR. Both amplified cDNA products were restricted with NcoI or HaeIII endonucleases and identical restriction fragments were produced. Two efficient methods of viral RNA concentration from an infected tobacco leaf extract were used; both 3–3.5 M sodium acetate alone and 3 M LiCl with 4 M urea quantitatively precipitated TMV RNA from the extracts. TMV RNA thus obtained could be readily amplified by direct RNA PCR. These results demonstrate that direct RNA PCR can be applied for the detection or/and analysis of high molecular weight RNA and for diagnosis of viral infections.
Mitogen‐induced expression of the primary response gene cMG1 in a rat intestinal epithelial cell‐line (RIE‐1)
Authors: Gomperts Miranda , Corps Anthony N. , Pascall John C. , Brown Kenneth D.
Published: July 2002

cMG1 is a primary response gene first identified in a rat intestinal epithelial (RIE‐1)cell‐line[(1990) Oncogene 5, 1081–1083]. A number of mitogens, including epidermal growth factor (EGF), angiotensin II (AII), serum and insulin rapidly induced 2‐ to 6‐fold increases of cMG1 mRNA in RIE‐1 cells, while transforming growth factor‐β caused a small reduction. Cyclic AMP‐elevating agents blocked the increase of cMG1 mRNA induced by EGF. The All‐stimulated increase of cMG1 mRNA was blocked by the depletion of protein kinase C, whereas the EGF‐stimulated increase was not affectcd, indicating that protein kinase C‐dependent and ‐independent signalling pathways stimulate cMG1 expression.
Regeneration of vitamin E in rat polymorphonuclear leucocytes
Authors: Ho Chuong T. , Chan Alvin C.
Published: July 2002

The objectives of this study were to determine whether the recycling of tocopherol occurs in elicited rat polymorphonuclear leukocytes and if so, whether the recycling process is anzymic or chemical. When incubated with hemoglobin. tocopherol was oxidized in cell homogenates in a time‐and concentration‐dependent manner. The oxidized tocopherol could be regenerated by addition of ascorbate, glutathione or nordihydroguaiaretic acid. Time course studies showed a rapid regeneration of tocopherol which peaked at 1 min after the addition of reductants. Determination of the regeneration reaction in the presence of CHCl3 and MeOH indicated that under these enzyme‐denaturing conditions, a considerable amount of tocopherol was still regenerated, suggesting that the regeneration reaction is predominantly a chemical reaction. This study provided direct evidence from mass analysis that oxidized vitamin E can be regenerated by cellular water‐soluble reductants such as ascorbate and glutathione.
Rabbit fast skeletal muscle phospholipase C Molecular weight determination by renaturation after polyacrylamide gel electrophoresis in the presence of sodium dodecylsulfate
Authors: Windhofer Volker , Varsànyi Magdolna , Heilmeyer Ludwig M.G.
Published: July 2002

Phosphoinositide specific phospholipase C from rabbit fast skeletal muscle has been enriched ca. 1,000‐fold with a specific activity of 40 μmol × min−1 × mg−1. Following SDS‐PAGE, renaturation of the enzyme protein in the presence of deoxycholate allowed the determination of an apparent molecular weight of 110 kDa. Gel‐filtration of the native enzyme resulted in a very similar apparent molecular weight of 115 kDa, however, associated proteins of higher molecular weight were also found. Free Ca2+ concentrations needed for half‐maximal activation of PtdIns(4,5)P2, PtdIns4P and PtdIns hydrolysis are 6.3 μM, 85 μM and 1.8 mM, and the Km
values for these substrates 102, 340 and 937 μM, respectively.
Crucial role of pyrophosphate in the aminoacylation of E. coli tRNAPhe by yeast phenylalanyl‐tRNA synthetase
Authors: Khvorova A.M. , Motorin Yu.A. , Wolfson A.D.
Published: July 2002

Rapid inactivation of the yeast phenylalanyl‐tRNA synthetase in the course of aminoacylation of the heterologous E. coli tRNAPhe is observed. This inactivation occurs due to the formation of the tight complex of the enzyme with the pyrophosphate formed during the aminoacylation reaction. This complex is shown to be the normal intermediate of the reaction. Possible inactivation mechanism and correlation between structural differences of yeast and E. coli tRNAsPhe with the changes in the enzymatic mechanism of aminoacylation are discussed.