PMID(sorted ascending)
reconstitution of biological molecular generators of electric current. inorganic pyrophosphatase.proteoliposomes have been reconstituted from soy-bean phospholipids (asolectin) and inorganic pyrophosphatase isolated from rhodospirillum rubrum chromatophores. in the presence of mg2+ ions, pyrophosphatase proteoliposomes were incorporated into a phospholipid-impregnated teflon filter separating two solutions of an identical electrolyte content. addition of inorganic pyrophosphate to the same compartment as proteoliposomes was found to induce generation of an electric potential difference betw ...19806109629
energy-linked reactions in photosynthetic bacteria: pi in equilibrium with hoh oxygen exchange catalyzed by the membrane-bound inorganic pyrophosphatase of rhodospirillum rubrum. 19816114711
[role of inorganic pyrophosphate in the cell bioenergetics (author's transl)]. 19816121321
reconstitution of highly purified proton-translocating pyrophosphatase from rhodospirillum rubrum. 19826128256
a sensitive and rapid method for determination of pyrophosphatase activity.a new, rapid and sensitive method of measuring the rate and amount of ppi hydrolysis is described. the method is based on registration of small ph changes, caused by decrease of hydrogen-ion concentration during the pyrophosphatase reaction. with this detected value of hydrogen-ion concentration change and the buffering capacity of each reaction mixture determined by titration, the rate and amount of ppi hydrolysis can be calculated. the results obtained by ph method are in good agreement with v ...19826131564
inorganic pyrophosphate-driven atp-synthesis in liposomes containing membrane-bound inorganic pyrophosphatase and f0-f1 complex from rhodospirillum rubrum.ppi driven atp synthesis has been reconstituted in a liposomal system containing the membrane-bound energy-linked ppiase and coupling factor complex, both highly purified from rhodospirillum rubrum. this energy converting model system was made by mixing both enzyme preparations with an aqueous suspension of sonicated soybean phospholipids and subjecting to a freeze-thaw procedure. in the presence of adp, mg2+, pi and ppi the system catalyzed phosphorylation by up to 25 nmol atp formed x mg prote ...19836132837
evidence for a glutamine synthetase-chromatophore association in the phototroph rhodospirillum rubrum: purification, properties, and regulation of the enzyme.the characteristics of soluble and membrane-bound glutamine synthetase (gs) from rhodospirillum rubrum were compared with those of the enzyme located in situ (measured in detergent-treated cells). the results suggest that in vivo gs may be associated with, or bound to, the chromatophore membranes. gs was found to reversibly associate and dissociate from purified chromatophores as a function of the ionic strength of the buffer or the mg2+ concentration. solubilized gs was purified to homogeneity ...19836132914
[effect of 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone, a ubiquinone analog, and sh-reagents on the electrogenic function of pyrophosphatase from rhodospirillum rubrum chromatophores].inorganic pyrophosphate-induced membrane potential generation in rhodospirillum rubrum chromatophores is inhibited by 2,5-dibromo-2-methyl-6-isopropyl-p-benzoquinone (dbmib). the inhibition is increased by menadione and dithionite and is not relieved by ascorbate; this effect is probably due to a displacement of the intrachromatophore quinones by dbmib. the sh-reagents, n-ethylmaleimide and p-chloromercurybenzoate at high concentrations inhibit membrane potential generation driven by inorganic p ...19836135455
h2 metabolism in photosynthetic bacteria and relationship to n2 fixation.the photosynthetic bacteria can evolve h2 in the light through a nitrogenase-mediated reaction. the nitrogenase enzyme in the photosynthetic bacteria is similar to other nitrogenases. it is made of two soluble components: a) the fe protein (dinitrogenase reductase or component ii) which receives electrons from ferredoxin, and b) the mo-fe protein (dinitrogenase or component i) on which the substrates (including protons) are reduced. in photosynthetic bacteria, the physiological regulation of nit ...19836139053
derepression of nitrogenase by addition of malate to cultures of rhodospirillum rubrum grown with glutamate as the carbon and nitrogen source.rhodospirillum rubrum grown in continuous culture with glutamate as the sole fixed c and n source produced no nitrogenase, and the cultures were characterized by high extracellular ammonium concentrations. addition of organic acids derepressed nitrogenase. glutamate dehydrogenase, glutamine synthetase, glutamate synthase, malate dehydrogenase, nitrogenase, and ammonium were assayed before and after malate addition.19846145702
membrane-bound inorganic pyrophosphatase. 19846151664
purification and immunological studies of glutathione reductase from rat liver. evidence for an antigenic determinant at the nucleotide-binding domain of the enzyme.glutathione reductase has been purified to homogeneity by a method which is an improvement of an earlier procedure (carlberg, i. and mannervik, b. (1975) j. biol. chem. 250, 5475-5480). the new steps in the purification scheme include affinity chromatography on 2',5' adp-sepharose 4b. antibodies to glutathione reductase from rat liver were raised in rabbits and used for analysis of the enzyme by quantitative 'rocket' immunoelectrophoresis. glutathione reductase from human erythrocytes, porcine e ...19816170346
sequential removal and reconstitution of subunits beta and gamma from a membrane-bound fo.fl-atp synthase. 19816171271
the nucleotide sequence of phenylalanine trna and 5s rna from rhodospirillum rubrum.the complete nucleotide sequence of trnaphe and 5s rna from the photosynthetic bacterium rhodospirillum rubrum has been elucidated. a combination of in vitro and in vivo labelling techniques was used. the trnaphe sequence is 76 nucleotides long, 7 of which are modified. the primary structure is typically prokaryotic and is most similar to the trnaphe of escherichia coli and anacystis nidulans (14 differences of 76 positions). the 5s ribosomal rna sequence is 120 nucleotides long and again typica ...19816174192
biosynthesis of bacterial glycogen: purification and structural and immunological properties of rhodopseudomonas sphaeroides adpglucose synthetase.adpglucose synthetase from the photosynthetic bacterium rhodopseudomonas sphaeroides was purified to greater than 95% purity. the molecular weight of the r. sphaeroides enzyme, as determined by sucrose density gradient ultracentrifugation, was approximately 204,000. the subunit molecular weight of the enzyme based on sodium dodecyl sulfate-gel electrophoresis was 46,000. although the amino acid composition of the enzyme was similar to that found for the enzymes from escherichia coli, salmonella ...19826178721
isolation and purification of an active gamma-subunit of the f0.f1-atp synthase from chromatophore membranes of rhodospirillum rubrum. the role of gamma in atp synthesis and hydrolysis as compared to proton translocation.we have earlier shown that extraction of rhodospirillum rubrum chromatophores with licl removed completely the beta-subunit of their coupling factor atpase complex leaving the other four subunits attached to the membrane (philosoph, s., binder, a., and gromet-elhanan, z. (1977) j. biol. chem. 252, 8747-8752). further treatment of these beta-less chromatophores with libr, under the described optimal conditions, resulted in specific removal of one additional subunit, the gamma-subunit, and both su ...19826181058
chemical modification of the beta-subunit isolated from a membrane-bound fo-f1-atp synthase: modification by 4-chloro-7-nitrobenzofurazan does not inhibit restoration of atp synthesis or hydrolysis. 19826184056
immunochemical relationship of the major outer membrane protein of rhodopseudomonas sphaeroides 2.4.1 to proteins of other photosynthetic bacteria.immunoblots of sodium dodecyl sulfate-polyacrylamide gels derived from outer membrane preparations of various strains of rhodopseudomonas sphaeroides revealed polypeptides which cross-reacted with antibody directed against the major outer membrane protein of r. sphaeroides 2.4.1. immunochemical quantitation of the major outer membrane protein of strain 2.4.1 showed approximately 5.5 x 10(4) molecules per cell whether cells were grown chemoheterotrophically or photoheterotrophically. rhodospirill ...19836188744
neutron small angle scattering of matched proteoliposomes with incorporated f0f1 atpase complex from rhodospirillum rubrum fr1. an approach to the structure of membrane proteins in their natural environment.purified f0f1 atpase from rhodospirillum rubrum fr1 has been incorporated into lipid vesicles from the partially deuterated phospholipid dimyristoylglycerophosphocholine (dmpc-d54). these proteoliposomes were able to carry out energy transducing reactions. the incorporation of the membrane protein was controlled by freeze fracture electron microscopy. a method for structural research of the membrane protein in its natural environment has been developed by means of neutron small angle scattering. ...19836195064
n-terminal sequences of subunits l and m of the photosynthetic reaction centre from rhodospirillum rubrum g-9+. separation of the subunits by gel filtration on hydroxypropylated sephadex g 100 in organic solvents.a new method has been developed by which subunits l and m of the photosynthetic reaction centre from rhodospirillum rubrum g-9+ can be obtained in pure form, starting form freeze-dried chromatophore membranes. the method employs extraction into a mixture of chloroform/methanol and gel permeation chromatography on a column of hydroxypropylated sephadex g 100. cross-contamination of the purified subunits was less than 5% (mol/mol), as estimated by manual edman degradation. automated edman degradat ...19836199280
polyadenylated mrna from the photosynthetic procaryote rhodospirillum cellular rna extracted from rhodospirillum rubrum cultured in butyrate-containing medium under strict photosynthetic conditions to the stationary phase of growth has been fractionated on an oligodeoxythymidylic acid-cellulose column into polyadenylated [poly(a)+] rna and poly(a)- rna fractions. the poly(a)+ fraction was 9 to 10% of the total bulk rna isolated. analysis of the poly(a)+ rna on a denaturing urea-polyacrylamide gel revealed four sharp bands of rna distributed in heterodisperse ...19846199342
antibodies to the f1-atpase of rhodospirillum rubrum and its purified native beta-subunit: inhibition of atp-linked activities in r. rubrum and in lettuce.1. antibodies prepared against the rhodospirillum rubrum f1-atpase (rrf1) and its purified, native-beta-subunit, exhibited cross-reactivity with the following soluble preparations of r. rubrum atpase: rrf0 . f1, rrf1 and the beta-subunit. anti-rrf1, but not anti-beta antibodies, also formed precipitin lines with soluble beta-less rrf1, indicating that antigenic determinants of both the beta-subunit and the other four rrf1-subunits are expressed in the whole rrf1 molecule. both antibodies aggluti ...19816210525
the interaction of 4-chloro-7-nitrobenzofurazan with rhodospirillum rubrum chromatophores, their soluble f1-atpase, and the isolated purified beta-subunit.atp synthesis and hydrolysis by rhodospirillum rubrum chromatophores as well as the soluble rrf1-atpase activity are inhibited by 4-chloro-7-nitrobenzofurazan (nbd-c1) in a dithiothreitol-reversible manner. using the method earlier developed in these chromatophores to remove specifically the beta-subunit from their membrane-bound rrf1 leaving all other subunits attached to the resulting inactive beta-less chromatophores (philosoph, s., binder, a., and gromet-elhanan, z. (1977) j. biol. chem. 252 ...19836219996
the interaction of carboxyl group reagents with the rhodospirillum rubrum f1-atpase and its isolated beta-subunit.the carboxyl group reagents dicyclohexylcarbodiimide (dccd) and n-ethoxycarboxyl-2-ethoxy-1,2-dihydroquinoline (eedq) inactivate the soluble rhodospirillum rubrum f1-atpase (rrf1). the inactivation is both time- and concentration-dependent and also ph-dependent, being more marked at acid ph. under the same conditions, n-ethyl-5-phenylisoxazolium 3'-sulfonate causes almost no inactivation of the rrf1-atpase. complete inhibition of the enzyme activity requires the binding of 1 mol of dccd/mol of r ...19836219997
[reconstruction of highly purified proton-translocating pyrophosphatase from rhodospirillum rubrum].using the freezing-thawing procedure, a highly purified preparation of ppase from r. rubrum chromatophore membranes was incorporated into soybean phospholipid liposomes. the activity of reconstituted ppase was increased in the presence of the uncoupler, fccp, and the antibiotics, valinomycin (+kcl) and nigericin (+kcl). oligomycin did not exert any inhibiting action, while imidodiphosphate and naf significantly decreased the activity of the ppase incorporated into the liposomes. preincubation of ...19836226320
response of soluble and membrane-bound f1 atpase of rhodospirillum rubrum to the photoaffinity label 8-azido atp.soluble atpase (f1 atpase) and membrane-bound atpase (chromatophores) of rhodospirillum rubrum have been photoaffinity labeled with 8-azidoadenosine 5'-triphosphate (8-n3atp). the specific binding of 8-n3atp to nucleotide binding sites of the chromatophore-bound enzyme is established by competition experiments and it is suggested that hydrolysis and phosphorylation involve the same nucleotide binding sites. the experimental results also indicate that the function of the coupling factor f1 is inf ...19836237651
preparative purification of the subunits of chloroplast and rhodospirillum rubrum coupling factors by flat-bed electrofocusing in granulated gels.a rapid method for the preparative purification of the subunits of oligomeric proteins like chloroplast and rhodospirillum rubrum coupling factors is presented. it involves the dissociation of the protein in urea and the separation of its subunits by isoelectric focusing in flat-beds of sepharose cl-4b or sephadex g-75 superfine, in the presence of urea and in an overnight run. using this procedure in the ph range 5-7, we have purified to homogeneity the alpha, beta and delta subunits of chlorop ...19846240506
purification and identification of the factor capable of converting ca2+-atpase into mg2+-atpase present in rhodospirillum rubrum chromatophores. 19806244266
cytochrome c and the evolution of energy metabolism. 19806244618
structure of cytochrome c': a dimeric, high-spin haem protein. 19806250058
two sequence-specific deoxyribonucleases from rhodospirillum rubrum. 19806250895
electron transfer reactions of high-potential iron-sulfur proteins and c-type cytochromes.studies of electron transfer by biological oxidation-reduction proteins frequently focus on the interaction of a particular protein with nonphysiological oxidants and/or reductants. this approach, although valuable, is limited by the size and chemistry of the nonphysiological reactants. to further the understanding of biological electron transfer, we have investigated the interaction of two examples of high-potential iron-sulfur proteins (hipip's) with mitochondrial cytochrome c (horse heart) an ...19806252957
nuclear magnetic resonance studies of rhodospirillum rubrum cytochrome c'.cytochrome c' from rhodospirillum rubrum has been studied by proton magnetic resonance (nmr) at 270 mhz. the ph and temperature-dependence properties as well as proton water relaxation enhancement and bulk susceptibility measurements were examined. we conclude that the fifth ligand to the iron is histidine. the ph-dependent shift of the heme methyl resonances of the ferric protein shows pka's at 5.8 and 8.7. the low-ph equilibrium causes only minor changes in the properties of the protein. howev ...19816258631
comparison of the redox reactions of various types of cytochrome c with iron hexacyanides.the dynamic behavior of various types of cytochromes c in the redox reaction with iron hexacyanides was studied using a temperature-jump method in order to elucidate the molecular mechanism of the redox reaction of cytochromes with their oxidoreductants. transmittance after the temperature jump changed through a single exponential decay for all cytochromes investigated. under a constant concentration of anion, the redox reaction of various types of cytochrome c with iron hexacyanides was analyze ...19816258647
biosynthesis of bacterial glycogen: purification and structural properties of rhodospirillum tenue adenosine diphosphate glucose synthetase.adenosine diphosphate glucose synthetase from the photosynthetic bacterium rhodospirillum tenue has been purified greater than 95%. the molecular weight of the enzyme is approximately 215,000, with a subunit molecular weight of about 51,000. the enzyme appears to be composed of four similar if not identical subunits. although the amino acid composition of the enzyme is similar to that of escherichia coli and salmonella typhimurium, no apparent homology has been observed between their n-terminal ...19816263861
on the evolutionary relationship of the 4-alpha-helical heme proteins. the comparison of cytochrome b562 and cytochrome c'.the atomic models of the cytochrome b562 and cytochrome c' monomers have been compared. when the respective heme groups are superimposed, the four alpha-helices of each nearly coincide. four aromatic side chains, including the heme ligands, and a methionine occur in spatially equivalent positions in contact with the heme groups. this structural evidence suggests that the two cytochrome families may have diverged from a common molecular ancestor.19816267021
biosynthesis of bacterial glycogen: activator specificity of the adenosine diphosphate glucose pyrophosphorylases from the genus rhodospirillum.the adenosine diphosphate (adp) glucose pyrophosphorylases from rhodospirillum fulvum, rhodospirillum molischianum, and rhodospirillum tenue were partially purified, and their kinetic properties were studied. the enzyme from the three organisms was found to be activated by pyruvate and thus was similar to the rhodospirillum rubrum enzyme that had been previously studied (c. e. furlong, and j. preiss, j. biol. chem. 244:2539-2548, 1979). the enzymes from r. fulvum, r. molischianum, and r. tenue w ...19816268603
epr studies of a nonphotosynthetic mutant of rhodospirillum rubrum. 19816271071
incorporation of atp synthetase into long-term stable liposomes of a polymerizable synthetic sulfolipid. 19816271594
metal coordination centres of class ii cytochromes c.the class ii cytochromes rhodospirillum molischianum cytochrome c', rhodopseudomonas palustris cytochrome c556 and agrobacterium tumefaciens (b2a) cytochrome c556 have been investigated with a variety of spectroscopic techniques. the cytochrome c' was found to be high-spin and the two cytochromes c556 were found to be mainly low-spin and sx-coordinate with the fifth and sixth ligands being histidine and methionine. the implications of the different types of iron coordination are discussed.19826279397
crystallographic structure of rhodospirillum molischianum ferricytochrome c' at 2.5 a resolution. 19816279876
correlations between structural and spectroscopic properties of the high-spin heme protein cytochrome c'.the cytochromes c' are a class of heme proteins whose native spectroscopic properties have been suggested to represent a quantum mechanical admixture of intermediate-(s = 3/2) and high-(s = 5/2) spin states. here features of the cytochrome c' heme environment, as revealed by x-ray crystallographic studies of the dimeric cytochrome c' from rhodospirillum molischianum, are related to the observed spectroscopic properties. the environment of the heme group in cytochrome c' supports the existence of ...19826293536
ribulose-1,5-bisphosphate carboxylase: enzyme-catalyzed appearance of solvent tritium at carbon 3 of ribulose 1,5-bisphosphate reisolated after partial reaction.when ribulose 1,5-bisphosphate is allowed to react with carbon dioxide in tritiated water in the carboxylation reaction catalyzed by ribulose-1,5-bisphosphate carboxylase from rhodospirillum rubrum, the ribulose 1,5-bisphosphate reisolated after partial reaction is found to be labeled. the specific radioactivity of the remaining substrate pool rises during the course of the reaction. experiments in deuterium oxide show that the isotopic label resides on carbon 3. earlier failures to detect this ...19826293539
[generation of the differences of the electric potentials by rhodospirillum rubrum reaction center complexes devoid of the heavy subunit].the electrogenic activity of rhodospirillum rubrum p870 reaction center complexes devoid of the heavy (h) subunit and retaining the light (l) and medium (m) subunits, was studied. the proteoliposomes containing such reaction center complexes were formed by a self-assembly procedure, using soya bean phospholipids. in the presence of ca2+ the reaction center proteoliposomes were incorporated into a phospholipid-impregnated teflon filter separating two solutions of identical composition. after addi ...19826293591
individual 1h-nmr assignments for the heme groups and the axially bound amino acids and determination of the coordination geometry at the heme iron in a mixture of two isocytochromes c-551 from rhodopseudomonas gelatinosa.this paper describes chemical and physicochemical studies of two small isocytochromes c-551 (approx. 9000 dalton) from rhodopseudomonas gelatinosa. in spite of numerous amino acid substitutions in the n-terminal half of the sequence the two isoproteins could not be separated by the procedures used, presumably because they have identical size, charge and isoelectric points. individual assignments of the 1h-nmr lines of heme c and the axial ligands to the heme iron were therefore obtained by nucle ...19836297597
nitrogen fixation and ammonia switch-off in the photosynthetic bacterium rhodopseudomonas viridis.rhodopseudomonas viridis atcc 19567 grows by means of nitrogen fixation in yeast extract-n2 or nitrogen-free medium when sparged with 5% co2 and 95% n2 in the light at 30 degrees c. acetylene reduction assays for nitrogenase activity revealed an initially high level of activity during early-logarithmic growth phase, a lower plateau during mid- to late-logarithmic phase, and a dramatic reduction of activity at the beginning of the stationary phase. when viewed by electron microscopy, nitrogen-fix ...19836305906
two types of essential carboxyl groups in rhodospirillum rubrum proton atpase.two different types of essential carboxyl groups were detected in the extrinsic component of the proton atpase of rhodospirillum rubrum. chemical modification of r. rubrum chromatophores or its solubilized atpase by woodward's reagent k resulted in inactivation of photophosphorylating and atpase activities. the apparent order of reaction was nearly 1 with respect to reagent concentration and similar k1 were obtained for the soluble and membrane-bound atpases suggesting that inactivation was asso ...19836307154
characterization of a rhodospirillum rubrum plasmid: loss of photosynthetic growth in plasmidless strains.a single plasmid of 55 kilobases was found in crude cell lysates of each of nine strains of rhodospirillum rubrum. restriction endonuclease analysis showed identical fragment patterns with a given nuclease for all plasmids except one, for which an additional ecori site was observed. elimination of the plasmid required that the cells be passaged several times in 25 mm calcium-containing medium, followed by at least two passages under photosynthetic growth conditions in low-calcium medium before t ...19836313617
the dibromothymoquinone effect on membrane potential generation in rhodospirillum rubrum chromatophores.2,5-dibromo-3-methyl-6-isopropyl benzoquinone (dbmib) inhibits the light-dependent membrane potential generation in rhodospirillum rubrum chromatophores. the inhibition is relieved by electron donors and is obviously due to oxidation of the photosynthetic electron transfer chain components. in addition, high dbmib concentrations elicit another effect probably caused by disruption of quinone functions in chromatophores. however, in quinone-depleted chromatophores and proteoliposomes containing th ...19836316108
identification of nitrogenase and carboxylase genes in the photosynthetic bacteria and cloning of a carboxylase gene from rhodopseudomonas sphaeroides.the presumptive genes for the ribulose 1,5-bisphosphate carboxylase large subunit and for nitrogenase-specific components from rhodopseudomonas sphaeroides and several other photosynthetic bacteria were identified and located by interspecific probing. restriction digests of r. sphaeroides genomic dna were hybridized under stringent conditions to cloned dna from rhodospirillum rubrum (plasmid prr2119 carrying the carboxylase gene) and klebsiella pneumoniae (psa30 carrying the nitrogenase genes). ...19836319239
reconstruction of photosynthetic, cyclic electron transport system from photoreaction unit, ubiquinone-10 protein, cytochrome c2 and polar lipids purified from rhodospirillum was previously reported that in chromatophores of rhodospirillum rubrum, reaction center, which consists of three kinds of protein (mm, about 78k), is a small fragment of a large protein complex (pru; photoreaction unit), which contains six other kinds of protein including light-harvesting bacteriochlorophyll protein, has mm of about 700k and is free of phospholipid [j. biochem. 86, 1211-1224 (1979); 94, 1815-1826 (1983(]. in the present study, the photosynthetic, cyclic electron transport sy ...19846325401
rna ligase in bacteria: formation of a 2',5' linkage by an e. coli extract.ligase activity was detected in extracts of escherichia coli, clostridium tartarivorum, rhodospirillum salexigens, chromatium gracile, and chlorobium limicola. ligase was measured by joining of trna halves produced from yeast ivs-containing trna precursors by a yeast endonuclease. the structure of trnatyr halves joined by an e. coli extract was examined. the ligated junction is resistant to nuclease p1 and rnaase t2 but sensitive to venom phosphodiesterase and alkaline hydrolysis, consistent wit ...19836347395
molecular cloning of the phosphoenolpyruvate carboxylase gene, ppc, of escherichia coli.the cole1 hybrid plasmid, plc20-10, carrying the ppc gene and the argecbh gene cluster of escherichia coli k-12, was characterized. the ppc gene coding for phosphoenolpyruvate carboxylase (ec, was subcloned into the plasmid pbr322 to give the plasmids ps2 and ps3. these plasmids carried a 4.4-kb sali segment containing the ppc gene, in both orientations. the specific activity of the enzyme was increased approx. 20-fold by these plasmids. experiments with maxicells harboring ps2 showed ...19846396163
cloning and expression in escherichia coli of the form ii ribulose 1,5-bisphosphate carboxylase/oxygenase gene from rhodopseudomonas sphaeroides.the gene encoding the form ii ribulose 1,5-bisphosphate carboxylase/oxygenase (rubpc/o) from rhodopseudomonas (r.) sphaeroides has been identified on a 3-kb ecori fragment and cloned into a broad-host-range, high-copy-number plasmid, using the gene from rhodospirillum (rs.) rubrum as a hybridization probe. subclones of the gene from r. sphaeroides in pbr322 and puc8 show substantial levels of expression and enzymatic activity in whole cells and crude cell extracts of escherichia coli. this enzym ...19846396166
derepression of the synthesis of d-ribulose 1,5-bisphosphate carboxylase/oxygenase from rhodospirillum rubrum.the synthesis of ribulose 1,5-bisphosphate carboxylase/oxygenase in rhodospirillum rubrum was greatly influenced by the conditions of culture. when grown photolithotrophically in an atmosphere containing low levels of co2 (1.5 to 2%), enzyme synthesis was derepressed, with the result that the enzyme comprised up to 50% of the soluble protein of the cells as determined by immunological quantitation. this response was not observed when r. rubrum was grown photolithotrophically in an atmosphere of ...19836401286
overproduction of nitrogenase by nitrogen-limited cultures of rhodopseudomonas palustris.rhodopseudomonas palustris cells grown on limiting nitrogen produced four- to eightfold higher nitrogenase specific activity relative to cells sparged with n2. the high activity of n-limited cells was the result of overproduction of the nitrogenase proteins. this was shown by four independent techniques: (i) titration of the mo-fe protein in cell-free extracts with fe protein from azotobacter vinelandii; (ii) direct detection of the subunits of mo-fe protein by sodium dodecyl sulfate-polyacrylam ...19836402491
identification of two distinct lactate dehydrogenases in rhodospirillum rubrum.the activities of pyridine nucleotide-independent d- and l-lactate dehydrogenases were detected in membranes from rhodospirillum rubrum grown under aerobic and phototrophic conditions. crossed immunoelectrophoretic analysis revealed two antigenically distinct enzymes that were further distinguished by specificity for d- and l-stereoisomers of lactate and by the sensitivity of the d-lactate dehydrogenase to inhibition by oxamate and oxalate.19836402502
[purification of adenyl cyclase from the phototrophic bacterium rhodospirillum rubrum]. 19836403060
adenine nucleotide levels in and nitrogen fixation by the cyanobacterium anabaena sp. strain 7120.adenine nucleotide levels were determined in whole filaments of anabaena sp. 7120 grown under different n2-fixing or non-n2-fixing conditions. these were compared with levels in isolated heterocysts, rhodospirillum rubrum, and azotobacter vinelandii. adenine nucleotides in whole filaments of anabaena sp. do not reflect the energetic expense of n2 fixation as they do in r. rubrum and a. vinelandii. however, adenine nucleotide levels in heterocysts were similar to the levels found in n2-fixing r. ...19836403506
isolation and sequencing of an active-site peptide from rhodospirillum rubrum ribulosebisphosphate carboxylase/oxygenase after affinity labeling with 2-[(bromoacetyl)amino]pentitol 1,5-bisphosphate.2-[(bromoacetyl)amino]pentitol 1,5-bisphosphate was reported to be a highly selective affinity label for ribulosebisphosphate carboxylase/oxygenase from rhodospirillum rubrum [fraij, b., & hartman, f. c. (1982) j. biol. chem. 257, 3501-3505]. the enzyme has now been inactivated with a 14c-labeled reagent in order to identify the target residue at the sequence level. subsequent to inactivation, the enzyme was carboxymethylated with iodoacetate and then digested with trypsin. the only radioactive ...19836404301
action of chlorophyllase purified from rye seedlings on light-harvesting bacteriochlorophyll of chromatophores and spheroplasts from rhodospirillum rubrum.1. the chlorophyllase [ec] purified from greened rye seedlings hydrolyzed the bacteriochlorophyll isolated from rhodospirillum rubrum, but not the pigment bound to the membrane of chromatophores or spheroplasts from the bacterium. 2. acetone, if added at such concentrations that the bound bacteriochlorophyll would not be solubilized, enabled the enzyme to hydrolyze the bound pigment. the acetone concentrations required for half the maximum hydrolysis rates were 16% with chromatophores a ...19836404894
hybridization of cloned rhodopseudomonas capsulata photosynthesis genes with dna from other photosynthetic bacteria.the homology of rhodopseudomonas capsulata dna segments carrying photosynthesis genes with sequences present in total dna from certain other photosynthetic and non-photosynthetic bacterial species was determined by hybridization. r. capsulata dna fragments that carry loci for production of peptide components of the photosynthetic reaction center and light-harvesting i antenna complex were found to hybridize to dna from some photosynthetic species. however, fragments that carry carotenoid or bact ...19836406432
[interaction of redox mediators with chromatophores of the photosynthetic bacterium rhodospirillum rubrum]. 19836408397
incorporation of adenine into the modifying group of inactive iron protein of nitrogenase from rhodospirillum rubrum.adenine was fed to cells of rhodospirillum rubrum grown on glutamate. the adenine was found to be incorporated into the modifying group of the inactive form of iron protein. adenine labelled in the 8-position ([8-3h]adenine) and the 2-position ([2-3h]adenine) was specifically incorporated into the electrophoretic 'upper-band' subunit of iron protein. incorporation of label from the 2-position into many proteins was observed if histidine was not present in the medium. label was removed by the act ...19836409076
ribulose bisphosphate carboxylase/oxygenase in toluene-permeabilized rhodospirillum rubrum.toluene-permeabilized rhodospirillum rubrum cells were used to study activation of and catalysis by the dual-function enzyme ribulose bisphosphate carboxylase/oxygenase. incubation with co2 provided as hco3-, followed by rapid removal of co2 at 2 degrees c and subsequent incubation at 30 degrees c before assay, enabled a determination of decay rates of the carboxylase and the oxygenase. half-times at 30 degrees c with 20 mm-mg2+ were 10.8 and 3.7 min respectively. additionally, the concentration ...19836409101
architectonic natures of proteins bound to rhodospirillum rubrum chromatophores as detected by trypsin treatment and sonication. 19836409893
structure of the extracellular ferredoxin from rhodospirillum rubrum: close similarity to clostridial ferredoxins.the amino acid sequence of an [8fe-8s] ferredoxin isolated from the culture medium of rhodospirillum rubrum, a photosynthetic purple non-sulfur bacterium, was determined by a combination of various conventional procedures. the sequence was a-y-k-i-e-e-t-c-i-s-c-g-a-c-a-a-e-c-p-v-n-a-i-e-q-g-d-t-i-f-v-v-n-a-d-t-c-i-d-c - g-n-c-a-n-v-c-p-v-g-a-p-v-a-e (55 amino acid residues). it lacked methionine, leucine, histidine, arginine, and tryptophan. the molecular weight was calculated to be 5,568 exclud ...19836411697
the role of mg2+ and mn2+ in the enzyme-catalysed activation of nitrogenase fe protein from rhodospirillum rubrum.the activation of the fe protein of nitrogenase (rr2) from glutamate-grown rhodospirillum rubrum by activating enzyme (ae) was investigated. ae is confirmed to have mr about 20 000 and is shown to operate catalytically. there is a role in activation for metal-ion-atp, which can be met by either mnatp or mgatp. there is also a site of action for free metal ions. this site prefers mn2+ (apparent kd approx. 20 microm) over mg2+ (apparent kd approx. 20 mm) by a factor of 1000-fold. non-activated rr2 ...19836412690
[intramolecular dynamics and electron transfer in photosynthetic reaction centers. a study using luminescence].the temperature dependences of fluorescence and phosphorescence spectra maxima of chromophor labels--endogenic (tryptophan) and exogenic (eosinisothiocyanate)--were measured for the preparations of photosynthetic membranes and reaction centers from rhodospirillum rubrum. it was found that the dipole mobility of protein-lipid matrix in the vicinity of the chromophores intensified markedly with a temperature rise from 150 to 300k resulting in the corresponding relaxation time tau r decrease from 1 ...19836413837
[dicyclohexylcarbodiimide as an inhibitor of light- and pyrophosphate-induced formation of membrane potential in chromatophores of purple bacteria].n,n'-dicyclohexylcarbodiimide (dccd) suppresses the uptake of penetrating tetraphenylborate anions by rhodospirillum rubrum chromatophores during cyclic and non-cyclic electron transfer and atp and pp i hydrolyses. the photochemical activity of the bacteriochlorophyll reaction centers of the chromatophores in insensitive to dccd. this supports the view that dccd inhibits the electron transfer between the primary and secondary quinones of the photosynthetic chain. incorporation of the chromatopho ...19836414533
inhibition of ribulose bisphosphate carboxylase by substrate ribulose 1,5-bisphosphate.substrate ribulose bisphosphate is a potent and a weak inhibitor of the rate of co2/mg2+ activation in the carboxylase purified from spinach leaves and rhodospirillum rubrum, respectively. at 2 degrees c, the concentration of ribulose bisphosphate required for 50% inhibition of the initial rate of co2/mg2+ activation was less than 0.4 microm for the spinach enzyme, but between 67 and 270 microm for the r. rubrum carboxylase. activator 14co2 trapping experiments demonstrated that ribulose bisphos ...19836417133
uptake of methionine sulfoximine by some n2 fixing bacteria, and its effect on ammonium transport.the n2 fixing bacteria klebsiella pneumoniae, azospirillum brasilense, rhodopseudomonas sphaeroides and rhodospirillum rubrum, but not azotobacter vinelandii accumulate the glutamine analogue methionine sulfoximine in the cell. in the accumulating cells methionine sulfoximine inhibits ammonium transport. accumulation and inhibition are prevented by glutamine.19836418571
2-(4-bromoacetamido)anilino-2-deoxypentitol 1,5-bisphosphate, a new affinity label for ribulose bisphosphate carboxylase/oxygenase from rhodospirillum rubrum. determination of reaction parameters and characterization of an active site peptide.a new affinity label for ribulose bisphosphate carboxylase/oxygenase from rhodospirillum rubrum, 2-(4-bromoacetamido)anilino-2-deoxypentitol 1,5-bisphosphate, has been prepared, reductive amination of ribulose-p2 with p-phenylenediamine in the presence of sodium cyanoborohydride yielded an epimeric mixture which was resolved by chromatography on quaternary aminoethyl-sephadex. subsequent bromoacetylation of the isolated amino bisphosphates gave reagents a and b (ribo and arabino epimers of 2-(4- ...19846421817
2-[8-14c]naphthyl 2-diazo-3,3,3-trifluoropropionate, a new carbene generating reagent for probing hydrophobic membrane domains.a new precursor of a lipophilic photolabel, 2-[8-14c]naphthyl 2-diazo-3,3,3-trifluoropropionate (nadit) has been synthesized. the suitability of the reagent for labeling the hydrophobic core of membranes is demonstrated by studying its reactivity in chromatophores of rhodospirillum rubrum g-9+. the label binds preferentially to the phospholipids and intrinsic membrane proteins. in isolated reaction centers treated with nadit the hydrophobic subunits m and l are more labeled than the h subunit. t ...19846422985
chemical nature of protein complex of photoreaction unit including reaction center in chromatophores of photosynthetic bacterium, rhodospirillum rubrum, as detected by successive dissociation method.reaction center of chromatophores of rhodospirillum rubrum consists of three kinds of protein, h-, m-, and l-subunit, and is bound with many other kinds of protein to form a larger protein complex (pru; photoreaction unit), which contains all the bacteriochlorophyll. in the present study, purified pru was dissociated in a stepwise manner in the presence of various mixtures of lithium dodecyl sulfate, sodium cholate and/or sodium deoxycholate, and separated into five, smaller protein complexes (p ...19836423620
the shape of ribulose bisphosphate carboxylase/oxygenase in solution as inferred from small angle neutron scattering.small angle neutron scattering of both activated and deactivated hexadecameric ribulose bisphosphate carboxylase/oxygenase from spinach revealed that its structure in solution very closely resembles that determined for the deactivated crystalline enzyme from tobacco (baker, t.s., eisenberg, d., and eiserling, f. (1977) science 196, 293-295). scattering from both forms of the enzyme in h2o most closely fits that expected from a hollow sphere with an outer radius of 56.4 a and inner radius of 14.3 ...19846423629
x-ray diffraction studies on chromatophore membrane from photosynthetic bacteria. ii. comparison of diffraction patterns of photosynthetic units from various purple bacteria.comparative x-ray diffraction studies, in conjunction with infrared absorption spectroscopy, were performed on chromatophores isolated from various purple photosynthetic bacteria in order to achieve a better understanding of the molecular structure of the photosynthetic unit. purple non-sulfur bacteria used were rhodospirillum rubrum, rhodospirillum molischianum, rhodopseudomonas sphaeroides, and rhodopseudomonas palustris. chromatophores of chromatium vinosum, as a typical example of purple sul ...19846425275
x-ray diffraction studies on photosystem i fragments from a blue-green alga, anabaena variabilis, and spinach.photosystem i fragments were prepared from thylakoid membranes of a blue-green alga (anabaena variabilis) and spinach by treatment with a detergent, triton x-100. equatorial x-ray diffraction patterns were recorded on films for oriented specimens of thylakoid membranes and photosystem i fragments. the thylakoid membranes and photosystem i fragments gave essentially the same equatorial diffraction patterns in both anabaena variabilis and spinach, indicating that the major x-ray scatterers in thes ...19846425276
effect of ammonia, darkness, and phenazine methosulfate on whole-cell nitrogenase activity and fe protein modification in rhodospirillum rubrum.a procedure for the immunoprecipitation of fe protein from cell extracts was developed and used to monitor the modification of fe protein in vivo. the subunit pattern of the isolated fe protein after sodium dodecyl sulfate-polyacrylamide gel electrophoresis was assayed by coomassie brilliant blue protein staining and autoradiographic 32p detection of the modifying group. whole-cell nitrogenase activity was also monitored during fe protein modification. the addition of ammonia, darkness, oxygen, ...19846427184
intermediates in the ribulose-1,5-bisphosphate carboxylase least two intermediates of the d-ribulose-1,5-bisphosphate carboxylase/oxygenase (ec reaction were liberated in detectable amounts when the functioning enzyme from rhodospirillum rubrum was quenched in acid. using substrate labeled with 32p in c-1, [32p]orthophosphate (pi) was found when the quenched solution was rapidly processed for extraction of pi as the acid molybdate complex. reaction with sodium borohydride under mildly alkaline conditions immediately after acid quenching of ...19846427222
preliminary x-ray diffraction study of ribulose-1,5-bisphosphate carboxylase from rhodospirillum rubrum.crystals from the dimeric enzyme ribulose-1,5-bisphosphate carboxylase of the photosynthetic bacterium rhodospirillum rubrum have been obtained from the gene product expressed in escherichia coli. the crystals are of the quarternary complex comprising enzyme: activator co2 (as a carbamate): mg2+: 2- carboxyarabinitol -1,5-bisphosphate (as a transition state analog). x-ray diffraction photographs show symmetry consistent with space group p4(1)2(1)2 or the corresponding enantiomorphic space group. ...19846427471
complete primary structure of ribulosebisphosphate carboxylase/oxygenase from rhodospirillum rubrum.of the 14 cyanogen bromide fragments derived from rhodospirillum rubrum ribulosebisphosphate carboxylase/oxygenase, four are too large to permit complete sequencing by direct means [f. c. hartman, c. d. stringer, j. omnaas, m. i. donnelly, and b. fraij (1982) arch. biochem. biophys. 219, 422-437]. these have now been digested with proteases, and the resultant peptides have been purified and sequenced, thereby providing the complete sequences of the original fragments. with the determination of t ...19846430239
carbon monoxide dehydrogenase from rhodospirillum rubrum.the carbon monoxide dehydrogenase from the photosynthetic bacterium rhodospirillum rubrum was purified over 600-fold by deae-cellulose chromatography, heat treatment, hydroxylapatite chromatography, and preparative scale gel electrophoresis. in vitro, this enzyme catalyzed a two-electron oxidation of co to form co2 as the product. the reaction was dependent on the addition of an electron acceptor. the enzyme was oxygen labile, heat stable, and resistant to tryptic and chymotryptic digestion. opt ...19846430875
preliminary structural studies of ribulose-1,5-bisphosphate carboxylase/oxygenase from rhodospirillum rubrum.ribulose-1,5-bisphosphate carboxylase/oxygenase (ec from rhodospirillum rubrum has been crystallized in a form that is suitable for structural studies by x-ray diffraction. the asymmetric unit of the crystal contains one dimeric enzyme molecule of molecular mass 101,000 da. the enzyme was activated prior to crystallization and is presumed to be in the co2-activated state in the crystal. the method of hydrophobicity correlation has been used to compare the amino acid sequence of this mo ...19846432800
the light-harvesting polypeptides of rhodospirillum rubrum. i. the amino-acid sequence of the second light-harvestng polypeptide b 880-beta (b 870-beta) of rhodospirillum rubrum s 1 and the carotenoidless mutant g-9+. carotenoidless mutant g-9+.the light-harvesting complex b 880 from rhodospirillum rubrum s 1 (wild type) and b 870 from the carotenoidless mutant g-9+ was shown to consist mainly of an organic solvent-(chloroform/methanol-) soluble and an organic solvent-insoluble polypeptide. the isolation and separation of these two low-molecular-mass polypeptides (mr 6101 and mr 6079) were achieved by a two-step extraction procedure of chromatophores using in the first step chloroform/methanol containing 0.1m ammonium acetate. followin ...19846434396
the light-harvesting polypeptides of rhodospirillum rubrum. ii. localisation of the amino-terminal regions of the light-harvesting polypeptides b 870-alpha and b 870-beta and the reaction-centre subunit l at the cytoplasmic side of the photosynthetic membrane of rhodospirillum rubrum g-9+.the unspecific proteinase k and the specific proteases alpha-chymotrypsin, trypsin and s. aureus v 8 protease were used in order to determine the orientation of the polypeptides b 870-alpha and b 870-beta from the major antenna complex b 870 of rs. rubrum g-9+ within the chromatophore membrane (inside-out vesicle). although b 870-alpha exhibits cleavable peptide bonds, treatment with specific proteases yielded splitting only in b 870-beta within the n-terminal region. in the case of proteinase k ...19846434397
plasmidless, photosynthetically incompetent mutants of rhodospirillum rubrum.ethyl methanesulfonate rendered a high percentage of rhodospirillum rubrum cells plasmidless and photosynthetically incompetent (kuhl et al., j. bacteriol. 156:737-742, 1983). by probing restriction endonuclease-digested chromosomal dna from these plasmidless strains with 32p-labeled r. rubrum plasmid dna, we showed that no homology exists between the plasmid and the chromosomal dna of the mutant. loss of the plasmid in all the nonphotosynthetic isolates was accompanied by the synthesis of spiri ...19846434514
nature and location of amide-bound (r)-3-acyloxyacyl groups in lipid a of lipopolysaccharides from various gram-negative has previously been demonstrated [eur. j. biochem. 124, 191-198 (1982) and 137, 15-22 (1983)] that the lipid a component of salmonella and proteus lipopolysaccharides contains amide-linked (r)-3-acyloxyacyl residues. in the present study lipid a of other gram-negative bacteria was analysed for the presence of amide-bound 3-acyloxyacyl residues. it was found that such residues are constituents of all lipid a tested (agrobacterium tumefaciens, chromobacterium violaceum, pseudomonas aeruginosa, ...19846437812
the mechanism of the attachment of esterifying alcohol in bacteriochlorophyll a biosynthesis.the mechanism through which the c-17(3) carboxy group of bacteriochlorophyllide a is esterified to produce bacteriochlorophyll aphytyl of rhodopseudomonas spheroides and bacteriochlorophyll ageranylgeranyl of rhodospirillum rubrum was studied by using 5-aminolaevulinate labelled with 18o at its c-1 carboxy oxygen atoms. the latter species was prepared by an exchange reaction in which 5-aminolaevulinate hydrochloride was heated in h218o in an autoclave. a method for the determination of the 18o c ...19846439193
the distribution of microsomal glutathione transferase among different organelles, different organs, and different the present study we have used both enzyme assay with 1-chloro-2,4-dinitrobenzene as substrate and immunochemical quantitation to examine the distribution of microsomal glutathione transferase in different organelles, in different organs, and in different organisms. this enzyme was found to constitute 3% and 5%, respectively, of the total protein recovered in the microsomal and outer mitochondrial membrane fractions from rat liver. microsomal glutathione transferase present in other subcellul ...19846439207
purification and properties of the activating enzyme for iron protein of nitrogenase from the photosynthetic bacterium rhodospirillum rubrum.the oxygen-labile, activating enzyme for iron protein from the photosynthetic bacterium, rhodospirillum rubrum, was purified 11,800-fold using a combination of chromatophore washing, de52-cellulose chromatography, hydroxylapatite chromatography, reactive red-120 cross-linked agarose chromatography, reactive red-120 cross-linked agarose chromatography, and sephadex g-75 gel filtration. activating enzyme appeared homogeneous on silver-stained sodium dodecyl sulfate-polyacrylamide gels, and the sta ...19846439722
[effect of dehydration on the primary picosecond stages of charge separation in rhodospirillum rubrum].effects of dehydration on the quantum yield of charge separation in the reaction centres, fluorescence and nanosecond recombination luminescence in r. rubrum chromatophores have been investigated. it has been shown that dehydration results in more than a 10 times decrease in the quantum efficiency of photosynthesis. besides, photoinduced fluorescence changes practically disappear in dehydrated samples and the parameters of nanosecond luminescence substantially change. these observations indicate ...19846441114
adenine nucleotide levels in rhodospirillum rubrum during switch-off of whole-cell nitrogenase activity.adenine nucleotide pools were measured in rhodospirillum rubrum cultures that contained nitrogenase. the average energy charge [([atp] + 1/2[adp])/([atp] + [adp] + [amp])] was found to be 0.66 and 0.62 in glutamate-grown and n-limited cultures respectively. treatment of glutamate-grown cells with darkness, ammonia, glutamine, carbonyl cyanide m-chlorophenylhydrazone, or phenazine methosulphate resulted in perturbations in the adenine nucleotide pools, and led to loss of whole-cell nitrogenase ac ...19846441571
x-ray diffraction studies on chromatophore membrane from photosynthetic bacteria. iii. basic structure of the photosynthetic unit and its relation to other bacteriochlorophyll forms.we have performed x-ray diffraction studies on photosynthetic units of rhodospirillum rubrum and solubilized *b800 + b890 complex from chromatophores of chromatium vinosum, to investigate the homology of their molecular structures. the native chromatophores of chromatium vinosum, which contain other bacteriochlorophyll forms, were examined by an x-ray diffraction technique, in order to assess the interactions between the complexes as well as the molecular structures of the bacteriochlorophyll fo ...19846442292
[pyrophosphate-dependent d-fructose-6-phosphate-phosphotransferase in rhodospirillaceae (author's transl)].a pyrophosphate-dependent fructose-6-phosphate phosphotransferase from the photosynthetic bacterium rhodospirillum rubrum was partially purified and characterized in respect to kinetic and regulatory properties. the enzyme had a molecular weight of about 95 000 dalton and required mg2+-ions for catalysis and maintenance of activity. the phosphotransferase was specific for fructose-6-phosphate (f-6-p) and inorganic pyrophosphate (pp) as substrates of the fructose-1,6-bisphosphate-forming reaction ...19806446207
energy-linked reactions catalyzed by the purified atpase complex (f0f1) from rhodospirillum rubrum chromatophores.1. the isolation of f0f1-atpase complex from rhodospirillum rubrum chromatophores by the use of taurodeoxycholate is described. 2. the enzyme preparation contains about 12 polypeptides; five are subunits of the f1 moiety. 3. the atpase activity of the purified enzyme is dependent on the addition of phospholipids. 4. km-vales for mg2+-atp and ca2+-atp are similar to the values obtained for the membrane-bound enzyme. 5. the f0f1-atpase complex is more than 70% inhibited by oligomycin and n,n'-dicy ...19806447594
conversion of the ca2+-atpase from rhodospirillum rubrum into a mg2+-dependent enzyme by 1,n6-etheno atp. 19806448051
atp synthesis catalyzed by the atpase complex from rhodospirillum rubrum reconstituted into phospholipid vesicles together with bacteriorhodopsin. 19806451197
unusual lipid a types in phototrophic bacteria and related species.photosynthetic bacteria of the rhodospirillaceae family (sulfur-free purple bacteria) possess lipopolysaccharides (lps) that deviate markedly from the salmonella lipopolysaccharides in the chemical makeup of the lipid a component and in their biologic properties. lps of rhodopseudomonas gelatinosa is highly toxic and pyrogenic, while that of rhodospirillum tenue shows cryptic toxicity. two lps types are completely non-toxic. the rhodopseudomonas sphaeroides lipid a has the same backbone as that ...19846474014
protein on the cell surface of the moderately halophilic phototrophic bacterium rhodospirillum salexigens.a cell surface protein (mr 68,000) of the moderately but obligately halophilic phototrophic bacterium rhodospirillum salexigens was identified by two independent methods: first, by labeling the cell surface with radioactive iodine and lactoperoxidase, and second, by washing cells in 30% sucrose to remove proteins attached to the cell surface by ionic bonds. the identified protein very likely represents the outermost layer of the cell envelope of r. salexigens as observed by electron microscopy. ...19846480555
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