bluuebunny/crossref_metadata_2025 huggingface dataset:
Third letters in codons counterbalance the (G + C)‐content of their first and second letters
Authors: Wada Akiyoshi , Suyama Akira
Published: July 2002

The correlation among the frequencies of appearance of G and C bases at the first, second and third positions of codons is examined. It is found that the redundancy at the wobbling third base is utilized to counter‐balance the local (G + C)‐content variation due to the first and second bases of codons, so that a homogeneous (G + C)‐content is provided to each individual gene. We speculate that this levelling tendency of (G + C)‐content comes from the functional constraint to achieve a uniform double‐helix stability in a gene.
Expression of plasmid encoded Escherichia coli 5 S ribosomal ribonucleic acid in Pseudomonas putida
Authors: Hartmann Roland K. , Henze Peter P. , Ulbrich Norbert , Erdmann Volker A.
Published: July 2002

The recombinant plasmid pNRK 36, which represents the plasmids RSF 1010, a small multicopy plasmid of the incompatability group IncQ that confers resistance to streptomycin and sulfoamide to its host cells, and pKK‐ 223‐3, which contains the try‐lac (tac) promoter followed by a polylinker and a DNA segment containing the 5 S rRNA (rrn B) with the ribosomal RNA transcription terminators, was employed to transform Pseudomonas putida 2440 cells. The plasmid encoded 5 S rRNA from Escherichia coli was transcribed and processed properly in P. putida cells thus demonstrating for the first time the expression of a plasmid encoded ribosomal RNA in a heterologous system. The fact that the E. coli 5 S rRNA was not incorporated into assembled ribosomes suggests that the in vivo incorporation of 5 S rRNA into the 50 S ribosomal sub‐unit is closely linked to 23 S rRNA and/or ribosomal protein synthesis.
A monoclonal antibody specific for the duplex DNA poly[d(TC)]· poly[d(GA)]
Authors: Lee Jeremy S. , Latimer Laura J.P. , Woodsworth Mary L.
Published: July 2002

Although most duplex DNAs are not immunogenic some synthetic DNAs such as polyl[d(Tm5C)] · poly‐[d(GA)] are weakly immunogenic allowing the production of monoclonal antibodies. The specificity of one of these antibodies, Jel 172, was investigated in detail by a competitive solid‐phase radioimmune assay. Jel 172 bound well to poly[d(TC)]·poly[d(GA)] but not to other duplex DNAs such as poly[d(TTC)] · poly‐[d(GAA)] and poly[d(TCC)]· poly[d(GGA)]. The binding to poly[d(Br5UC)]· poly[d(GA)] was enhanced while that to poly[d(TC)]· poly[d(IA)] was decreased compared to poly[d(TC)]· poly[d(GA)]. Thus, not only is the antibody very specific for a sequence of duplex DNA but it also appears to recognize functional groups in both grooves of the helix.
Discovery of a γ‐carboxyglutamic acid‐containing protein in human spermatozoa
Authors: Soute Berry A.M. , Müller-Esterl Werner , de Boer-van den Berg Marian A.G. , Ulrich Magda , Vermeer Cees
Published: July 2002

Here we describe the identification of a γ‐carboxyglutamic acid‐containing protein in human spermatozoa. After thermal decarboxylation the protein is a good substrate for vitamin K‐dependent carboxylase from various origins. A quick purification procedure for the decarboxylated protein is presented and in a preliminary characterization we have established its M
r, (28000–30000) and its amino acid composition.
Ca2+ requirement for photosynthetic oxygen evolution of spinach and mangrove photosystem II membrane preparations
Authors: Preston Christopher , Critchley Christa
Published: July 2002

CaCl2 significantly increased the photosynthetic oxygen‐evolution activity of photosystem II membrane preparations isolated from spinach and the mangrove Avicennia marina, but not that of spinach thylakoids. This increase is composed of two parts; one due to Cl−, the other to Ca2+. Under Cl−‐sufficient conditions photosystem II particle preparations required only 2 mM Ca2+ for maximum oxygen evolution, irrespective of the presence of the 23‐ and 18‐kDa polypeptides. It is suggested that there is no immediate interaction between Ca2+ and the 23‐ or 18‐kDa polypeptides and that instead Ca2+ acts directly on the oxygen‐evolving centre, perhaps via another, as yet unidentified protein.
The use of carotenoids and oxonol VI as probes for membrane potential in proteoliposomes
Authors: van Walraven H.S. , Krab K. , Hagendoorn M.J.M. , Kraayenhof R.
Published: July 2002

Carotenoids present in lipids extracted from the cyanobacterium Synechococcus 6716 indicate trans‐membrane potential in proteoliposomes reconstituted from these lipids and the ATPase complex isolated from the same organism. A carotenoid absorbance band shift to a longer wavelenght is obtained with valinomycin‐induced potassium ion diffusion potentials, irrespective of the polarity of the potassium gradient. In contrast to this, the (externally added) probe oxonol VI only shows an absorbance band shift when the external potassium ion concentration is higher than the internal one. In liposomes without ATPase complex, no carotenoid absorbance band shifts were observed.
Study of hepatic binding protein activity in jejuno‐ileal by‐passed rat hepatocytes
Authors: Serbource-Goguel Nathalie , Dodeur Michèle , Borel Béatrice , Feger Jeanne , Agneray Jean , Durand Geneviève
Published: July 2002

The kinetic constants of internalization of asialoorosomucoid were determined for normal and jejuno‐ileal by‐passed rat hepatocytes. In by‐passed rats the maximum velocity of asialoorosomucoid internalization is decreased 3‐fold, without any modification of apparant constant of internalization. Moreover, the rate constant of internalization was the same in the two groups of rats. These data suggest that the process of asialoorosomucoid internalization is not altered in by‐passed hepatocytes and that the decrease of maximal velocity is only due to a decrease of total uptake receptor number.
Potentiation of the antagonistic effect of ACTH11–24 on steroidogenesis by synthesis of covalent dimeric conjugates
Authors: Fauchère Jean-Luc , Rossier Michel , Capponi Alessandro , Vallotton Michel B.
Published: July 2002

Covalent dimerization of the adrenocorticotropin fragment ACTH11–24 increases its antagonistic potency on the ACTH‐induced steroidogenesis in isolated bovine fasciculata/reticularis cells by 3 orders of magnitude when the C‐termini are linked via a 10 Å long spacer. This strong potentiation, probably mediated by cross‐linking of the receptors, was shown to be dependent on the point of attachment of the monomeric fragment to the spacer, thus providing information about the position of the binding site in the hormonal segment and about the distance of the receptors on the cell surface.
The dynamic nature of DNA‐strand breaks present in differentiating muscle cells and quiescent lymphocytes
Authors: Farzaneh Farzin , Shall Sydney , Johnstone Alan P.
Published: July 2002

DNA‐strand break
ADP‐ribosylation
Differentiation
Mitogen activation
Use of phosphorothioate analogs of poly(dA‐dT) · poly(dAdT) to study steroidal‐diamine induced conformational change in poly(dA‐dT) · poly(dA‐dT)
Authors: Suggs J.William , Taylor Donna A.
Published: July 2002

31P NMR
DNA phosphorothioate
Steroidal diamine
DNA conformation
Incorporation of 19F‐substituted aromatic amino acids into membrane proteins from chromatophores of Rhodospirillum rubrum G9+
Authors: Ghosh R. , Bachofen R. , Hauser H.
Published: July 2002

Chromatophore membranes were isolated from cells of the carotenoidless mutant Rhodospirillum rubrum G9+ grown in the presence of several fluorinated aromatic amino acids, solubilized using SDS and the extent of incorporation analyzed using high‐resolution 19F‐NMR spectroscopy. 3‐ and 4‐19F‐phenylalanine, 6‐19F‐tryptophan and 3‐19F‐tyrosine were biosynthetically incorporated into membrane proteins whereas 5‐19F‐tryptophan and 2‐19F‐phenylalanine were inhibitors of cell growth. The polypeptide chains of the major chromatophore membrane protein the light‐harvesting complex, were isolated and shown by high‐resolution 19F‐NMR to contain 3‐19F‐phenylalanine, which is known to be situated principally within the membrane hydrocarbon layer. Broad‐band 19F‐NMR spectra of 3‐19F‐phenylalanine‐labelled chromatophores showed the phenyl ring to be immobilized within the membrane.
Alcohol dehydrogenase II and fructose‐1,6‐bisphosphatase appear to be co‐regulated in wild‐type yeast
Authors: Wills Christopher , Martin Tracy , Melham Thomas , Walker Diana
Published: July 2002

Activity gel electrophoresis
Gluconeogenesis
Regulation
Cardiolipin vesicles can accommodate cholesterol up to 0.80 mole fraction, i.e. one molecule per cardiolipin fatty acid chain
Authors: Gallay Jacques , de Foresta Béatrice , Vincent Michel
Published: July 2002

Cardiolipin‐cholesterol interactions were studied by time‐resolved fluorescence spectroscopy with 1,6‐diphenyl‐1, 3,5‐hexatriene as probe. The residual anisotropy parameter, r∞ (reflecting the fatty acid chain packing), was measured in the liquid crystalline phase as a function of cholesterol addition. Two main results are reported: (i) a slight increase of the order parameter, S, computed from the r
∞ value as S = (r
∞/r
0)

, in the physiological concentration range of cholesterol; (ii) a sharp enhancement of S from a cholesterol mole fraction (X̄
chl) of 0.20 and up to X̄
chl of 0.80. This is in contrast to unsaturated kcithin systems for which a continuous increase of the order parameter was monitored, culminating at X̄
chl = 0.50, the wellknown maximum level of incorporation of cholesterol into lecithin model membranes.
Functionally important site in the vicinity of the amino‐ terminus of the Escherichia coli RNA polymerase β subunit
Authors: Nikiforov V.G. , Kalyaeva E.S. , Moiseyeva E.P. , Yakubov L.Z.
Published: July 2002

We have analyzed the interaction of monoclonal antibodies against Escherichia coli RNA polymerase with products of its limited proteolysis. Two major proteolytic fragments of molecular masses 107 and 43 kDa originate as a result of a single cleavage in the vicinity of the 980th amino acid residue. Anti‐β subunit monoclonal antibody PYN‐2 inhibiting RNA polymerase activity at the stage of RNA elongation reacts with an epitope located between the amino‐terminus and the 50th amino acid residue of the β subunit. DNA sequencing has shown that the RNA polymerase mutation rpoB22 converts the Gln(1111) codon of the β subunit gene into the amber codon. An epitope for the monoclonal antibody PYN‐6 was located between the major site of proteolytic cleavage and Gln(1111) of the β subunit.
Biogenesis of liver δ‐aminolevulmate synthase
Authors: Friedland David M. , Ades Ibrahim Z.
Published: July 2002

In primary cultures of chick embryo hepatocytes pulse labeled with [35S]methionine, immunochemical analyses indicated that adenosine 3':5'‐cyclic monophosphate (cAMP) did not affect either the rate of production or the maturation of δ‐aminolevulinate synthase (ALA synthase). In addition, allylisopropylacetamide caused a slight drop in intracellular cAMP while testosterone caused the levels of cAMP to rise to 260% of the basal levels measured in hepatocytes in culture. Thus the results of this study did not indicate a direct short‐term role for cAMP in the regulation of production of ALA synthase.
Agarose gel electrophoretic evidence for domains of nuclear DNA linked with bonds cleavable with sulfhydryl molecules
Authors: Taş Sinan , De Larco Joseph , Altiok Ender
Published: July 2002

Complexes of intact nuclear DNA with proteins undisssociable by 2.0 M NaCl and nonionic detergents were analyzed by agarose gel electrophoresis following physical or enzymatic fragmentation. Sulfhydryl molecules converted these DNAs (but not the bacteriophage λ DNA) into smaller‐M
r forms. Following limited restriction endonulease digestion of complexes with PstI most of the nuclear DNA formed a high‐molecular‐mass band in the 60–110 kbp range. These 60–110 kbp fragments, releasable from the rest of nuclei by sulfhydryl molecules, have similar sizes to nuclear DNA loops detected by other techniques and may derive from supranucleosomal organizational units in the chromatin complex.
The role of carbohydrates in the radioimmunoassay of human low‐molecular‐mass kininogen
Authors: Turpeinen Ursula , Kärkkäinen Tytti
Published: July 2002

The immunoreactivity of human low‐molecular‐mass kininogen from Cohn plasma fraction IV was investigated after deglycosylations and carbohydrate modifications by radioimmunoassay using the conformation‐specific antiserum. Removal of all sialic acids, 44% of amino sugars and 63% of neutral sugars did not alter the immunoreactivity of the protein but the periodate‐treated concanavalin A fractions showed strikingly diminished immunoreactivity. A conformational change could account for the observed effect of periodate on the decreased reactivity of the protein in radioimmunoassay. Externally added carbohydrates had no effect on immunoreactivity. The results suggest that the carbohydrate part of kininogen is not involved in the immunoreactivity although it accounts for the observed lectin‐binding heterogeneity.
A small phospholipase inhibitory factor released by cultured cell lines
Authors: Wood John N. , Coote Paul R. , Salmon John , Rhodes John
Published: July 2002

Cells of the mouse macrophage‐like cell line RAW264 release a dialysable inhibitor of phospholipase activity into their culture medium. This inhibitor can be detected in saline solution, Hanks solution and a variety of tissue culture media in the presence or absence of serum. The inhibitor is stable at 4°C, unaffected by trypsin, nucleases, or boiling, and partially extractable with chloroform/methanol. The release of both arachidonic acid and prostaglandins from mouse macrophages or human monocytes is inhibited by this material. A variety of other cell types release the inhibitor, which is effective against stimulation of arachidonic acid release from cultured macrophages by zymosan, serum, immune complexes and the calcium ionophore A23187.
Locus‐specific detection of HLA‐DQ and ‐DR antigens by antibodies against synthetic N‐terminal octapeptides of the β chain
Authors: Deufel Thomas , Grove Anne , Kofod Hans , Lernmark Åke
Published: July 2002

Antibodies against synthetic peptides representing the class‐II antigen HLA‐DR and ‐DQ β chain N‐terminal sequences were prepared in rabbits. The two octapeptides only share two amino acids and enzyme‐linked immuno‐assays showed the antisera only to bind to its own antigen. Both peptide antisera detected a 29 kDa component in immunoblots of Raji and AL‐34 cell plasma membrane proteins separated by SDS gel electrophoresis. The binding of either N‐terminal peptide antiserum was selectively inhibited only by the peptide used as antigen. Indirect immunofluorescence.analysis by flow cytofluorometry showed specific surface immunofluorescence in 1:100–1:1000 dilutions in lymphoblastoid and blood mononucleated cells. In the latter the binding was primarily confined to monocytes and a subpopulation of lymphocytes. It is concluded that locus‐specific immunological reagents to distinguish between β chains of HLA‐DR and ‐DQ have been prepared by the preparation by the production of antibodies against the N‐terminal sequences of each polypeptide
Rapid proton release accompanying photosynthetic electron transport in intact cells of Rhodopseudomonas capsulata
Authors: Taylor M.A. , Jackson J.B.
Published: July 2002

The release of protons from intact cells of Rhodopseudomonas capsulata after either 4μs flashes or during brief periods of continuous illumination has been measured with the indicator, cresol red. The half‐time for H+‐release after a flash was 35 ms and the extent, 1H+ per 134 bacteriochlorophyll. Myxothiazol completely inhibited the flash‐induced H+‐release and antimycin A reduced it by 37%. The proton‐releasing reaction is discussed with reference to the protonmotive Q‐cycle. During continuous illumination the rapid phase of H+ release is followed by a lag and then by another period of acidification, suggesting that other protolytic reactions may be in operation.
Effects of bc1‐site electron transfer inhibitors on the absorption spectra of mitochondrial cytochromes b
Authors: Kamensky Yuriy , Konstantinov Alexander A. , Kunz Wolfram S. , Surkov Sergey
Published: July 2002

Changes are described that are brought about by antimycin, NoHOQnO, funiculosin, myxothiazol and mucidin in the α‐, β‐ and γ‐absorption bands of reduced and oxidized cytochromes b in the isolated complex bc
1, form beef heart mitochondria. The inhibitors can be divided into 2 groups. Antimycin, funiculosin and NoHOQnO are likely to shift the spectrum of b‐562 and compete for specific binding with complex bc
1 with each other but not with myxothiazol and mucidin. The spectral effects of the latter two inhibitors are more difficult to interpret and may involve contributions not only from b‐562 but from b‐566 as well. The existence of 2 independent inhibitor binding‐sites in the complex bc
1 corroborates the Q‐cycle hypothesis.
Evaluation and significance of kinetic parameters governing function of protein synthesis initiation factors eIF‐2 and eIF‐2B
Authors: Manchester K.L.
Published: July 2002

Initiation factor eIF‐2
Initiation factor eIF‐2B
Ternary complex
Crystallization and structure at 3.2 Å resolution of a terbium parvalbumin
Authors: Wéry J.P. , Dideberg O. , Charlier P. , Gerday C.
Published: July 2002

Crystal
X‐ray crystallography
Three‐dimensional structure
Tb3+

Parvalbumin
Ca2+‐binding site
Fluorescent and radiolabelling of pepsin‐digested human glomerular basement membrane with a newly developed hydroxy‐coumarin derivative (CASE)
Authors: Rand-Weaver Mariann , Abuknesha Ramadan A. , Price Robert G.
Published: July 2002

Iodination
Protein labelling
Human glomerular basement membrane
7‐Hydroxy‐coumarin‐3‐acetic acid N‐hydroxysucciniimydyl ester
Fluorescence
Inhibition of poly(ADP‐ribose) polymerase causes increased DNA strand breaks without decreasing strand rejoining in alkylated HeLa cells
Authors: Moran Michael F. , Ebisuzaki Kaney
Published: July 2002

Treatment of alkylated HeLa cells with 3‐aminobenzamide, an inhibitor of poly(ADP‐ribose) polymerase, increased the number of DNA strand breaks but did not affect the rate of strand rejoining. This suggests that an increase in DNA incision, not a decrease in ligation, results from the inhibition ofpoly(ADP‐ribose) polymerase in cells recovering from DNA damaged by alkylating agents. Poly(ADP‐ribose) DNA strand break DNA repair
Loss of heat‐shock acquisition of thermotolerance in yeast is not correlated with loss of heat‐shock proteins
Authors: Cavicchioli Rick , Watson Kenneth
Published: July 2002

Yeast cells when subjected to a primary heat shock, defined as a temperature shift from 23 to 37° for 30 min, acquired tolerance to heat stress (52°C/5 min). Primary heat shocked cells incubated at 23°C for up to 3 h, progressively lost thermotolerance but retained high levels of the major heat‐shock proteins as observed on polyacrylamide gels. On the other hand, a temperature shift back up to 37°C for 30 min fully restored thermotolerance. The major high‐molecular‐mass heat‐shock proteins (hsp) identified were of approximate molecular mass 100 kDa (hsp 100), 80 kDa (hsp 80) and 70 kDa (hsp 70). The results indicate that loss of heat‐shock acquisition of thermotolerance is not correlated with loss of heat‐shock proteins.
The roles of HPr and FPr in the utilization of fructose by Escherichia coli
Authors: Kornberg Hans
Published: July 2002

A mutant impaired in FPr activity was isolated. The altered gene (fpr), which was located near min. 44 on the E. coli genome, was transferred by phage‐mediated transduction to appropriate recipients that lack HPr (ptsH), or Enzyme IIman (ptsM), or neither. The rates of growth on fructose of such transductants indicate that phosphate from PEP is transferred predominantly via FPr to fructose that enters the cells by Enzyme IIfru, but that HPr can play a role in transferring phosphate to fructose taken up via Enzyme IIman
The effect of different types of physical exercise on glucose and citrulline synthesis in isolated rat liver parenchymal cells
Authors: Bobyleva-Guarriero Valentina , Lardy Henry A.
Published: July 2002

Rates of gluconeogenesis from lactate or pyruvate in hepatocytes from untrained rats were not increased by an acute (1 h) bout of exercise (running at 20 ). Hepatocytes from rats that had been exercise‐trained for 1 month had lower rates of gluconeogenesis from lactate than cells from unexercised controls; the rates with pyruvate were identical. Hepatocytes from livers of trained animals immediately after 1 h of exercise synthesized glucose more rapidly and accumulated more citrulline than cells from resting rats.
Dynamic aspects of affinity labelling as revealed by alkylation and phosphorylation of pancreatic ribonuclease with reactive deoxyribodinucleotide derivatives
Authors: Knorre D.G. , Buneva V.N. , Baram G.I. , Godovikova T.S. , Zarytova V.F.
Published: July 2002

Affinity labelling of pancreatic RNase with 4‐(N‐2‐chloroethyl‐N‐methylamino)benzylamide and (N → P) N‐methylimidazolide of d(pTpA) results in the formation of monomodified enzyme derivatives retaining partially enzymatic activity. These data together with some cases described in the literature are considered as suggesting the dynamic nature of the enzyme‐reagent complex represented by a set of states differing in the probability of intra‐complex reaction. In particular, modification may proceed in a low probability state with an especially favorable mutual orientation of reagent and some protein residue remote from the active site of the enzyme resulting in the removal of the covalently attached reagent moiety from the active center.
Energetics of interaction of oligopeptide (lac 53–57) with DNA base sequences and origin of sequence‐specific recognition
Authors: Kalia A. , Royyuru A.K. , Mrigank  , Kothekar V.
Published: July 2002

Computer model building with a dynamic energy minimization procedure is used here to study the interaction of a pentapeptide sequence from the lac repressor headpiece (lac 53–57) with different base sequences of DNA. The peptide fragment for this purpose was considered in the classical β‐antiparallel as well as the β‐associated conformation. The model of its interaction with DNA was optimised for various binding positions and base sequences. Partitioning of energy is analysed for different dielectric constant values and the main contributing factors to sequence‐specific binding are discussed.
Lysine‐sensitive plant aspartate kinase is not regulated by calcium or calmodulin
Authors: Bonner P.L.R. , Hetherington A.M. , Lea P.J.
Published: July 2002

Lysine‐sensitive plant aspartate kinase purified from Daucus carota to a specific activity 20‐fold higher than previously reported was not activated by calcium or calmodulin. The calmodulin antagonists calmodazolium and compound did not inhibit activity. However, trifluoperazine exhibited an inhibitory effect, but only at high concentrations. The results do not agree with a previous proposal that calcium and calmodulin play a key role in regulating higher plant lysine‐sensitive aspartate kinase.
Heat shock induces variably the major heat shock proteins of CV1 clones
Authors: Matthopoulos Demetrios P.
Published: July 2002

CV1 cells have been subcloned several times. Five of these clones were studied for the induction of the major heat shock proteins. These CV1 clones exhibit morphological differences as well as differences in SDS‐PAGE protein profiles. These clones responded to heat shock variably as judged from the induction of the major heat shock proteins, 70, 72 and 92 kDa. Variable expression of the heat shock proteins suggests that the selective pressure for isolation of cell clones may affect gene expression differently.
Modification of some properties of spherical lipid membranes induced by the association with fructose‐1,6‐bisphosphate aldolase
Authors: Langner M. , Gutowicz J. , Gomułkiewicz J.
Published: July 2002

The influence of fructose‐1,6‐bisphosphate aldolase, as a membrane peripheral protein, on some electrical and transport properties of spherical lipid membranes was investigated. It was found that the association of the enzyme with the membrane did not effect markedly the electrical conductance or capacity of the membrane but decreased the water filtration coefficient and the cationic transferance number. The enzyme association also modifies temperature characteristics of the membrane parameters.
Identification of a c‐myc oncogene lacking the exon 1 in the normal cells of a patient carrying a thyroid carcinoma
Authors: Del Senno L. , Uberti E.Degli , Rossi M. , Buzzoni D. , Barbieri R. , Rossi P. , Patracchini P. , Bernardi F. , Marchetti G. , Conconi F. , Gambari R.
Published: July 2002

In this paper we describe an alteration of the c‐myc oncogene present in the white blood cells and normal as well as neoplastic thyroid cells of a subject carrying a thyroid carcinoma. Restriction enzyme mapping and hybridization to human c‐myc probes specific for different regions of this gene demonstrate that this subject carries, in addition to the normal one, a c‐myc oncogene lacking the first exon and part of the first intron. The levels of the c‐myc mRNA in thyroid cells of this subject do not show differences with respect to thyroid cells from other subjects. Taken together, these findings indicate that the deletion of the first exon of the c‐myc oncogene, in itself, does not produce overtranscription of this oncogene nor hematopoietic malignancies.
The N‐terminal 21 amino acids of a 70 kDa protein of the yeast mitochondrial outer membrane direct E. coli β‐galactosidase into the mitochondrial matrix space in yeast cells
Authors: Hase Toshiharu , Nakai Masato , Matsubara Hiroshi
Published: July 2002

The intracellular location of fusion proteins was investigated in yeast cells. They consisted of the N‐terminal 21,61 or 292 amino acids of the 70 kDa protein of the yeast mitochondrial outer membrane and an enzymatically active E. coli β‐galactosidase. The hybrids containing 61 or 292 residues of the 70 kDa protein, as well as the original 70 kDa protein, were localized on the outer membrane in a tightly membrane‐bound form. In contrast, the other hybrid was exclusively localized in the mitochondrial matrix space as a soluble protein.
Electron microscopy of the hydrogenase from the hydrogen‐oxidizing bacterium Alcaligenes eutrophus Z1
Authors: Tsuprun V.L. , Utkin I.B. , Popov V.O. , Egorov A.M. , Berezir I.V. , Kiselev N.A.
Published: July 2002

The quaternary structure of the NAD‐dependent hydrogenase from the hydrogen‐oxidizing bacterium Alcaligenes eutrophus Z1 has been studied by electron microscopy. A model of the hydrogenase molecule with two small subunits arranged between two large ones in linear fashion is proposed. The electron microscopy studies are in accordance with the previously proposed linear electron transport chain of the hydrogenase and distribution of the substrate binding sites within the enzyme molecule.
The role of Cu(I)‐thiolate clusters during the proteolysis of Cu‐thionein
Authors: Weser Ulrich , Mutter Wolfgang , Hartmann Hans-Jürgen
Published: July 2002

Rat liver Cu,Zn‐[35S]thionein and yeast Cu‐thionein were subjected to proteolysis in vitro using equilibrium dialysis. The partially copper‐loaded vertebrate thionein (2–7 ) was affected by different proteases including thermolysin, proteinase K, protease from Streptomyces griseus and lysosomal enzymes. Unlike the 2Cu‐thionein the respective 7Cu‐thiolate‐centred metallothionein was hardly proteolytically digested. In contrast to fully copper‐loaded native yeast Cu‐thionein both the H2O2‐oxidized and the metal‐free protein were effectively cleaved in the presence of proteinase K. It is important to realize that the native Cu(I)‐thiolate chromophore survives the proteolytic attack. When the copper‐sulphur bonding is broken and the same amount of copper is unspecifically bound to the thionein portion, proteolysis proceeds identically with respect to the rate observed in the presence of the apoprotein. The unsuccessful proteolysis of native Cu‐thionein is not attributable to a simple copper‐dependent inhibition of the proteinases. It is suggested that prior to proteolysis the copper‐sulphur clusters must be destroyed.
Amino acid sequences from the N‐terminal domain of Bacillus thuringiensis, subspecies alesti, δ‐endotoxin
Authors: Chestukhina G.G. , Tyurin S.A. , Osterman A.L. , Khodova O.P. , Stepanov V.M.
Published: July 2002

The tryptic peptides derived from the N‐terminal domain of δ‐endotoxin produced by Bacillus thuringiensis ssp. alesti have been sequenced and aligned by homology with the primary stuctures of δ‐endotoxins formed by B.t. kurstaki subspecies (K−1 and K−73). The N‐terminal regions of the domains (residues 1–346) show only marginal variations: 9 amino acid substitutions were found among 158 aligned residues of B.t. ssp. alesti δ‐endotoxin. In contrast, the following stretch of the sequence starting from residue 347 reveals ~ 50% of amino acid substitutions when compared with the structures of K−1 or K−73. As a rule, the fragments of B.t. ssp. alesti δ‐endotoxin that differ from K‐1 in structure precisely reproduce the corresponding sequences of K−73 δ‐endotoxin and, vice versa, those different from the K−73 sequences coincide with the respective structures of K−1 δ‐endotoxin. A few fragments present in B.t. ssp. alesti were also found but not in both B.t. ssp. kurstaki δ‐endotoxins. It appears that the structure of this region of B. thuringiensis δ‐endotoxins (347–625 residues) contains a pattern of relatively conservative and highly variable fragments. These variations might be responsible for the variations in specificity of the entomocidal action of δ‐endotoxins.
Deacylation kinetics of γ‐chymotrypsin in solution and in the crystal
Authors: Merli A. , Rossi G.L.
Published: July 2002

The rate of catalyzed hydrolysis of the acyl enzyme analogs indolacryloyl‐γ‐chymotrypsin and furylacryloyl‐γ‐chymotrypsin in the crystal has been measured by single crystal microspectrophotometry and compared with the rate of catalyzed hydrolysis of acyl‐γ‐chymotrypsin in solution and acyl‐α‐chymotrypsin both in solution and in the crystal. The maximal deacylation rate is the same for both species and independent of the physical state. However, the pH dependence of the deacylation rate of crystalline acyl‐γ‐chymotrypsin shows a 0.9 unit shift in the pK of the catalytic system which is unique and probably consequent to specific lattice interactions.
The occurrence and distribution of α‐hydroxy‐acid dehydrogenase in some members of the order Kinetoplastida
Authors: Taylor Mark B. , Gutteridge Winston E.
Published: July 2002

LDH‐X is the isoenzyme of lactate dehydrogenase found in mammalian spermatozoa, occurring in cytosolic and mitochondrial locations. Gossypol strongly inhibits it, and the spermicidal action of this compound is attributed to the disruption of a reducing shuttle. The flagellated protozoan, Trypanosoma cruzi, contains an enzyme activity similar to LDH‐X, called α‐hydroxy‐acid dehydrogenase, which is here shown to possess cytosolic and glycosomal components. The glycosome is a microbody‐like organelle containing the early glycolytic enzymes. We postulate that the inhibition of replication of T. cruzi by gossypol derives from interference with glycosomal reducing shuttles. T. lewisi resembles T. cruzi in this respect.
Requirement for bivalent cations in the actions of insulin and sodium nitroprusside on metabolism in rat adipocytes
Authors: Emami Shahin , Perry Michael C.
Published: July 2002

The requirement for Ca2+ and Mg2+ in the actions of insulin and sodium nitroprusside on rat adipocyte metabolism was investigated: (i) sodium nitroprusside, but not insulin, increased cGMP levels in cells incubated in the absence of Ca2+ and/or Mg2+; (ii) sodium nitroprusside and insulin are unable to increase the incorporation of [14C]glucose into triglycerides and [14C]leucine into proteins in the absence of Ca2+ and Mg2+; (iii) sodium nitroprusside and insulin showed antilipolytic actions in Ca2+‐ and Mg2+‐free medium. We conclude that in the absence of Ca2+ and Mg2+, sodium nitroprusside and insulin have very similar regulatory properties on triglyceride, protein synthesis and adrenaline‐stimulated lipolysis, but not on cGMP levels in rat adipocytes. This could provide evidence that omission of bivalent cations was inhibitory at more than one site, or that sodium nitroprusside mimics insulin's actions by another mechanism that does not involve cGMP.
Synthesis of pyrophosphate coupled to the reverse energy‐linked transhydrogenase reaction in Rhodospirillum rubrum chromatophores
Authors: Nore Beston F. , Husain Iqbal , Nyrén Pål , Baltscheffsky Margareta
Published: July 2002

Chromatophores from the photosynthetic bacterium Rhodospirillum rubrum contain a membrane‐bound transhydrogenase catalyzing the transfer of a hydride ion between NADH and NADP+. The reverse reaction, i.e. reduction of NAD+ by NADPH, can furnish sufficient energy (ΔgmH+) to drive the phosphorylation of inorganic orthophosphate (Pi) to pyrophosphate (PPi). The rate of ppi synthesis is 50 nmol PPi formed/min per μmol Bchl which is 5% of the rate of light‐induced PPi synthesis. PPi synthesis is inhibited by both the H+‐PPase inhibitor fluoride and the specific transhydrogenase inhibitor palmitoyl‐CoA. The effects of both DCCD and uncouplers on the system provide additional evidence that the ΔgmH+ generated by the reverse transhydrogenase reaction drives PPi synthesis. The rate of PPi synthesis can be partially inhibited by the addition of NADP+ a substrate of the forward energy‐consuming reaction. The ΔgmH+ generated can also be used to drive ATP synthesis by the H+‐ATPase, but at a lower rate than the ppi synthesis.
Protein phosphorylation as a control for excitation energy transfer in Rhodospirillum rubrum
Authors: Holmes Nigel G. , Allen John F.
Published: July 2002

Changes in cooperativity of the light‐harvesting system of Rhodospirillum rubrum were monitored in whole cells by measurements of in vivo bacteriochlorophyll fluorescence yield transients. Under non‐cooperative conditions a polypeptide of 10.5 kDa (provisionally identified as B880‐β) was specifically phosphorylated; under cooperative conditions a polypeptide of 13 kDa (provisionally identified as B880‐α) was specifically phosphorylated. These results are discussed in relation to the regulation of excitation energy distribution in purple photosynthetic bacteria and other photosynthetic organisms.
Insulin release has no absolute requirement for extracellular monovalent ions
Authors: Hermans M.P. , Henquin J.C.
Published: July 2002

Current chemiosmotic models of exocytosis ascribe an essential role to the influx of extracellular monovalent anions or cations into the secretory granules apposed to the plasma membrane. These hypotheses were tested by measuring insulin release in sucrose media devoid of monovalent ions. A small response to glucose (25% of controls) was still observed, which could be potentiated by isobutylmethylxanthine and suppressed by cobalt or low temperature. Substitution of Ba2+ for Ca2+ triggered a practically normal release of insulin that was inhibited by blockers of Ca2+ channels (cobalt or D 600) and abolished by low temperature. These results show that insulin release remains possible in the absence of extracellular monovalent ions and, therefore, that the chemiosmotic models of exocytosis do not entirely apply to insulin release.
The regulation of phosphofructokinase in epimastigote Trypanosoma cruzi
Authors: Taylor Mark , Gutteridge Winston E.
Published: July 2002

Phosphofructokinase
D‐Fructose 2,6‐bisphosphate (Trypanosoma cruzi, Parasitic protozoan)
Accumulation and subcellular localization of glutelin‐2 transcripts during maturation of maize endosperm
Authors: Pérez-Grau Lluis , Cortadas Jordi , Puigdomènech Pere , Palau Jaume
Published: July 2002

The expression of glutelin‐2 genes has been studied during maize endosperm maturation and compared to the expression of two different zein genes. Glutelin‐2 mRNA in endosperm accumulates in parallel to zein mRNAs, nevertheless, the opaque‐2 mutation does not affect glutelin‐2 transcription. Subcellular fractionation of endosperm tissue shows that glutelin‐2 mRNA is accumulated in the protein body fraction indicating that these organelles are the place of translation of glutelin‐2 mRNA. The results indicate that the levels of glutelin‐2 transcripts in maize endosperm increase following a temporal pattern similar to that displayed by zein transcripts, the typical maize storage protein.
Reinvestigation of the roles of the carboxyl groups of glutathione with yeast glyoxalase I
Authors: D'Silva Claudius
Published: July 2002

A number of carboxyl‐substituted s‐blocked glutathiones have been shown to be competitive inhibitors of yeast glyoxalase I at 25°C, pH 6.6. Amidation of the glycyl carboxyl group of S‐(p‐bromobenzyl)glutathione has no appreciable effect on binding whilst methylation reduces binding by 8.9‐fold, indicating a steric constraint and the possible presence of a hydrogen bond in this region of the enzyme. Amidation of both carboxyl groups of S‐(p‐bromobenzyl)glutathione reduces binding significantly by 237‐fold; this result agrees with electrostatic interaction of the Glu COO− group with a group located within the enzyme surface as opposed to the Gly COO− group, previously proposed.
Isolation of a human ceruloplasmin cDNA clone that includes the N‐terminal leader sequence
Authors: Mercer Julian F.B. , Grimes Andrew
Published: July 2002

A number of cDNA clones encoding human ceruloplasmin were identified using two mixed oligonucleotide probes. One of these clones was shown by DNA sequence analysis to span from the complete N‐terminal leader sequence to 114 amino acids short of the C‐terminus. The leader sequence consists of 19 primarily hydrophobic amino acids. Northern blot analysis of RNA from human liver showed two species of ceruloplasmin mRNA; a minor species of 3600 nucleotides and a major one of 4400 nucleotides.
Immunological studies on the light‐harvesting polypeptides of photosystems I and II
Authors: Williams R.S. , Ellis R.J.
Published: July 2002

Monoclonal and polyclonal antibodies have been raised against the three apoproteins of the peripheral light‐harvesting complex of photosystem I (LHC I) from Pisum sativum L. These antibodies have been used to study the immunological relatedness of the light‐harvesting polypeptides of photosystems I and II. The results suggest that there is no immunological/structural relationship between the two light‐harvesting systems. The apoproteins of the LHC I fall into two distinct groups corresponding to the two chlorophyll complexes comprising the PS I antenna.
Metabolic turnover of proliferation‐related nuclear proteins in serum‐stimulated Swiss mouse 3T3 cells
Authors: O'Farrell Minnie K.
Published: July 2002

Stimulation of quiescent Swiss mouse 3T3 cells either by serum or by pure growth factors induces DNA synthesis after a lag period of about 15 h. Following restimulation by serum or by growth factors there is an overall increase of 2–4‐fold in the rate of biosynthesis of nuclear proteins. Two nuclear polypeptides show specific temporal correlations with the transition from quiescence to proliferation. The synthesis of p30 (30 kDa, pI 5.2) is at a maximum within 5 h of restimulation, while the synthesis of p36 (36 kDa, pI 4.25) is first seen at 10–12 h after restimulation. The synthesis of p36 correlates well with the initiation of DNA biosynthesis. The metabolic turnover of both of these proteins has been estimated by pulse‐chase and by cycloheximide inhibition experiments. They both have a half‐life of 10–15 h and appear to be cell‐cycle related.
The gramicidin A channel
Authors: Etchebest Catherine , Pullman Alberte
Published: July 2002

The effect of the conformational freedom of the ethanolamine tail of gramicidin A on the energy profile for the transfer of Na+, computed in the presence of water, shows an appreciable lowering of the minimum at 10.5 Å, and a splitting of the entrance barrier. The deep energy region at the channel mouth remains, however, the deepest one and contains a site of strong interaction of the ion with the Trp 11 carbonyl, at about 12 Å from the center.
An infrequent generation of catenated network of pBR322 in Escherichia coli
Authors: Komiyama Naoki , Shishido Kazuo
Published: July 2002

It was demonstrated that Escherichia coli infrequently generates the catenated network of pBR322. This complex pBR322 form was detected when DNA molecules could hardly enter the agarose gel during electrophoresis and was found to comprise monomers and dimers of the plasmid.
Quaternary structure of the liver microsomal cytochrome P‐450
Authors: Tsuprun V.L. , Myasoedova K.N. , Berndt P. , Sograf O.N. , Orlova E.V. , Chernyak V.Ya. , Archakov A.I. , Skulachev V.P.
Published: July 2002

Cytochrome P‐450LM2 was isolated from rabbit liver microsomes in a form which was shown to be homogeneous in AcA‐22 Ultrogel and ultracentrifugation studies. The molecular mass determined by sedimentation equilibrium roughly corresponded to hexamer composed of 56 kDa monomers. Hexamer structure of the cytochrome was directly demonstrated by electron microscopic study. In the cytochrome P‐450LM2 hexamer, monomers seem to be arranged in two layers (three monomers in the layer) in such a way that each monomer occupies aposition at the vertices of a triangular antiprism with a 32 point group symmetry.
Identification of three protein kinases which phosphorylate threonyl‐tRNA synthetase from rat liver
Authors: Pendergast Ann Marie , Traugh Jolinda A.
Published: July 2002

Threonyl‐tRNA synthetase is phosphorylated in Chinese hamster ovary cells labeled with 2Pi[(1984) J. Biol. Chem. 259,11160–11161]. Phosphorylation of the purified synthetase from rat liver has been examined with five different protein kinases. Three of the enzymes phosphorylate the synthetase, protease activated kinase I, the cAMP‐dependent protein kinase, and the Ca2+, phospholipid‐dependent protein kinase. Phosphorylation occurs exclusively on seryl residues. Two‐dimensional phosphopeptide maps of tryptic digests of the phosphorylated synthetase are distinct with each protein kinase. These data suggest that multiple phosphorylation of the synthetase may occur in vivo.
Hornetin: the lethal protein of the hornet (Vespa flavitarsus) venom
Authors: Ho Chewn-Lang , Ko Jane-Ling
Published: July 2002

By gel permeation on a Fractogel TSK HW 50 column followed by ion‐exchange chromatography on carboxymethylcellulose CM 52, a lethal protein, designated hornetin, was purified from the venom of Vespa flavitarsus. Hornetin is a highly basic protein (pI 10.2) with a molecular mass of about 32 kDa. Its amino acid composition is characterized by a high content of lysine, aspartic and glutamic acid, and is devoid of tryptophan and cysteine. The lack of cysteine in the molecule is distinct from other known vespid venom proteins of comparable size. The i.v. LD50 of the toxin is 0.42 μg per g mouse. Assayed on the red blood cells of the mouse and guinea‐pig as well as isolated nerve muscle preparations of the chick and mouse, hornetin showed direct hemolytic activity and presynaptic neurotoxicity at microgram level and displayed musculotropic effect at higher concentrations.
An organ‐specific hypomethylation of cotyledon genomic rDNA in Pisum sativum L
Authors: Waterhouse Rosemary N. , Boulter Donald , Gatehouse John A.
Published: July 2002

The CCGG and GCGC sequences in pea genomic rDNA were found to be hypomethylated relative to the leaf in the developing cotyledon. No rRNA gene copies were detected with only unmethylated CCGG sequences and the majority appeared to contain a mixture of doubly methylated (mCmCGG) and partially methylated (CmGG) sites. Some CCGG sequences were shown to have a greater probability of being undermethylated. Seed development did not appear to act as a stimulus for de novo methylation.
Restoration of hydroperoxide‐dependent lipid peroxidation by 3‐methylcholanthrene induction ofcytochrome P‐448 in hepatoma microsomes
Authors: Borrello Silvia , Galeotti Tommaso , Palombini Guglieimo , Minotti Giorgio
Published: July 2002

Microsomal membranes from the slow‐growing Morris hepatoma 9618A catalyze, in the presence of t‐butyl hydroperoxide, lower rates of lipid peroxidation than rat liver microsomes. The cytochrome P‐450 content of hepatoma microsomes is about 40% that of the liver. SKF 525‐A, an inhibitor of mixed‐function oxidase, produces in hepatoma microsomes a P‐450 type I binding spectrum similar to that of hepatic microsomes. The concentration of the inhibitor required for half‐maximal spectral change is about 2 μM in both microsome types. SKF 525‐A or ethylmorphine inhibit lipid peroxidation of normal and tumor microsomes to the same extent (about 60%). Treatment of the tumor‐bearing rats with 3‐methylcholanthrene increases the hepatoma cytochrome P‐450 to values comparable to those of control membranes, although the hemoprotein has a peak in the CO‐reduced difference absorption spectrum at 448 nm. The cytochrome P‐448 induction is accompanied by an almost complete restoration of the hydroperoxide‐dependent lipid peroxidation.
The branched respiratory chain of heterotrophically dark‐grown Chloroflexus aurantiacus
Authors: Zannoni Davide
Published: July 2002

The respiratory electron‐transport chain of heterotrophically dark‐grown Chloroflexus aurantiacus has been investigated. Membranes isolated from these cells have been shown to contain at least three c‐type cytochromes (E
m, 7.0 255,180, and 10 mV), three b‐type cytochromes (E
m, 7.0 of 210, 60 and −65 mV) and two cytochromes of the a type with E
m, 7.0 of 330 and 190 mV. Spectroscopic evidence from CO‐difference spectra, CN−‐duference spectra and spectra at fixed oxidation‐reduction potentials suggests that the two a‐type components may be analogous to cytochromes a and a
3 of mitochondria. The analyses of the effects induced by CN−, myxothiazol and antimycin A on both steady‐state respiratory activities and semi‐rapid oxidation‐reduction kinetic patterns of c‐ and a‐type cytochromes indicate the presence of a branched respiratory chain. Growth of Chloroflexus in medium lacking added copper diminished the concentration of the a‐type cytochromes but not those of cytochromes of the b and c type.
Scorpion venom (Leiurus quinquestriatus) elicits accumulations of inositol phosphates and cyclic AMP in guinea pig cortical synaptoneurosomes
Authors: Gusovsky Fabian , Daly John W.
Published: July 2002

Scorpion (Leiurus quinquestriatus) venom (ScV) stimulated accumulations of cyclic AMP and turnover of phosphatidylinositol in guinea pig cortical synaptoneurosomes. The concentrations of ScV that were necessary to increase cyclic [3H]AMP accumulation were lower than those required to stimulate formation of [3H]inositol phosphates from phosphatidylinositol. In the presence of 10 μM 2‐chloroadenosine, ScV induced a dose‐dependent synergistic accumulation of cyclic AMP with an EC50 value that was comparable to the EC50 required for stimulation of phosphatidylinositol turnover. Tetrodotoxin partially inhibited cyclic AMP accumulations elicited by ScV indicating that at least part of such responses are due to activation of voltage‐dependent sodium channel. Tetrodotoxin virtually completely blocked formation of inositol phosphate stimulated by ScV. High concentrations of Mg2+ (30 mM) did not block responses to ScV indicating that release of neurotransmitters was not involved. Membrane potential changes could not be detected at concentrations of ScV that triggered the biochemical responses. Stimulation of phosphatidylinositol turnover by ScV appears to depend on an increase in influx of Na+ in synaptoneurosomes, presumably due to slowing of the inactivation of voltage‐dependent sodium channels by α‐scorpion toxin, a component of ScV. At least in part, the stimulation of cyclic AMP accumulation by ScV correlates with increases in phosphatidylinositol turnover.