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effects of perturbations of the nitrogenase electron transfer chain on reversible adp-ribosylation of nitrogenase fe protein in klebsiella pneumoniae strains bearing the rhodospirillum rubrum dra operon.the redox state of nitrogenase fe protein is shown to affect regulation of adp-ribosylation in klebsiella pneumoniae strains transformed by plasmids carrying dra genes from rhodospirillum rubrum. the dra operon encodes dinitrogenase reductase adp-ribosyltransferase and dinitrogenase reductase-activating glycohydrolase, enzymes responsible for the reversible inactivation, via adp-ribosylation, of nitrogenase fe protein in r. rubrum. in bacteria containing the dra operon in their chromosomes, inac ...200010850982
purification and characterization of a membrane-bound hydrogenase from the hyperthermophilic archaeon pyrococcus furiosus.highly washed membrane preparations from cells of the hyperthermophilic archaeon pyrococcus furiosus contain high hydrogenase activity (9.4 micromol of h(2) evolved/mg at 80 degrees c) using reduced methyl viologen as the electron donor. the enzyme was solubilized with n-dodecyl-beta-d-maltoside and purified by multistep chromatography in the presence of triton x-100. the purified preparation contained two major proteins (alpha and beta) in an approximate 1:1 ratio with a minimum molecular mass ...200010852873
rhythmic activity of uptake hydrogenase in the prokaryote rhodospirillum rubrum.growth of rhodospirillum rubrum was followed in cultures kept under anoxic conditions at constant temperature in either continuous light (ll, 32 degrees c) or continuous darkness (dd, 32 degrees c and 16 degrees c). in dd, only small modifications of the turbidity were detected; linear regression analysis nevertheless gives a very significant slope (t(34) = 13.07, p < 10(-14), with r2 of 0.834). mean generation times reflected these differences of growth with 11.9+/-0.5 h in ll and 43.2+/-1.1 h ...200010885876
altering the specificity of cooa, the carbon monoxide-sensing transcriptional activator: characterization of cooa variants that bind cyanide in the fe(ii) form with high affinity.cooa is a carbon monoxide- (co-) sensing homodimeric heme protein that activates the transcription of genes required for the anaerobic oxidation of co to co(2) in the phototrophic bacterium rhodospirillum rubrum. in this study, we demonstrate that mutational alteration of the histidine residue (his(77)) that serves as a heme ligand in the fe(ii) form of cooa allows high-affinity binding of cyanide (k(d) approximately 0.4 mm) to the heme. in contrast, neither these same variants in the fe(iii) fo ...200010889037
evidence for a ligand co that is required for catalytic activity of co dehydrogenase from rhodospirillum rubrum.radiolabeling studies support the existence of a nonsubstrate co ligand (co(l)) to the fe atom of the proposed [feni] cluster of carbon monoxide dehydrogenase (codh) from rhodospirillum rubrum. purified codh has variable amounts of co(l) dissociated depending on the extent of handling of the proteins. this dissociated co(l) can be restored by incubation of codh with co, resulting in a 30-40% increase in initial activity relative to as-isolated purified codh. a similar amount of co(l) binding is ...200010891076
crystallization of the di component of transhydrogenase, a proton-translocating membrane protein.nicotinamide nucleotide transhydrogenase couples the exchange of a hydride-ion equivalent between nad(h) and nadp(h) to the translocation of protons across an energy-transducing membrane. peripheral components of 380 and 200 residues bind nad(h) (di) and nadp(h) (diii), respectively, while a third component forms a membrane-spanning region (dii). the nad(h)-binding component di of rhodospirillum rubrum transhydrogenase has been crystallized in a form which diffracts to beyond 3.0 a resolution an ...200010957636
protein-protein recognition, hydride transfer and proton pumping in the transhydrogenase complex.membrane-bound ion pumps are involved in metabolic regulation, osmoregulation, cell signalling, nerve transmission and energy transduction. how the ion electrochemical gradient interacts with the scalar chemistry and how the catalytic machinery is gated to ensure high coupling efficiency are fundamental to the mechanism of action of such pumps. transhydrogenase is a conformationally coupled proton pump linking a proton gradient to the redox reaction between nad(h) and nadp(h). the enzyme has thr ...200010997900
characterization of variants altered at the n-terminal proline, a novel heme-axial ligand in cooa, the co-sensing transcriptional activator.cooa, the carbon monoxide-sensing transcription factor from rhodospirillum rubrum, binds co through a heme moiety resulting in conformational changes that promote dna binding. the crystal structure shows that the n-terminal pro(2) of one subunit (met(1) is removed post-translationally) provides one ligand to the heme of the other subunit in the cooa homodimer. to determine the importance of this novel ligand and the contiguous residues to cooa function, we have altered the n terminus through two ...200011007793
solution structure of the nadp(h)-binding component (diii) of proton-translocating transhydrogenase from rhodospirillum rubrum.transhydrogenase is a proton pump found in the membranes of bacteria and animal mitochondria. the solution structure of the expressed, 21.5 kda, nadp(h)-binding component (diii) of transhydrogenase from rhodospirillum rubrum has been solved by nmr methods. this is the first description of the structure of diii from a bacterial source. the protein adopts a rossmann fold: an open, twisted, parallel beta-sheet, flanked by helices. however, the binding of nadp(+) to diii is profoundly different to t ...200011004437
dissociation and recombination between ligands and heme in a co-sensing transcriptional activator cooa. a flash photolysis study.cooa from rhodospirillum rubrum is a transcriptional activator in which a heme prosthetic group acts as a co sensor and regulates the activity of the protein. in this study, the electronic relaxation of the heme, and the concurrent recombination between ligands and the heme at approximately 280 k were examined in an effort to understand the environment around the heme and the dynamics of the ligands. upon photoexcitation of the reduced cooa at 400 nm, electronic relaxation of the heme occurred w ...200010978334
the orf162b sequence of rhodobacter capsulatus encodes a protein required for optimal levels of photosynthetic pigment-protein complexes.the orf162b sequence, the second open reading frame 3' of the reaction center (rc) h protein gene puha in the rhodobacter capsulatus photosynthesis gene cluster, is shown to be transcribed from a promoter located 5' of puha. a nonpolar mutation of orf162b was generated by replacing most of the coding region with an antibiotic resistance cartridge. although the mutant strain initiated rapid photosynthetic growth, growth slowed progressively and cultures often entered a pseudostationary phase. the ...200010986247
structure of the co sensing transcription activator cooa.cooa is a homodimeric transcription factor that belongs to the catabolite activator protein (cap) family. binding of co to the heme groups of cooa leads to the transcription of genes involved in co oxidation in rhodospirillum rubrum. the 2.6 a structure of reduced (fe2+) cooa reveals that his 77 in both subunits provides one heme ligand while the n-terminal nitrogen of pro 2 from the opposite subunit provides the other ligand. a structural comparison of cooa in the absence of effector and dna (o ...200011017196
pigment-protein architecture in the light-harvesting antenna complexes of purple bacteria: does the crystal structure reflect the native pigment-protein arrangement?structural analysis of crystallized peripheral (lh2) and core antenna complexes (lh1) of purple bacteria has revealed circular aggregates of high rotational symmetry (c8, c9 and c16, respectively). quantum-chemical calculations indicate that in particular the waterwheel-like arrangements of pigments should show characteristic structure-sensitive spectroscopic behavior in the near infrared absorption region. laser-spectroscopic data obtained with non-crystallized, isolated lh2 of rhodospirillum m ...200011034303
a transcriptional regulator of a pristinamycin resistance gene in streptomyces coelicolor.pip is a pristinamycin-induced transcriptional regulator protein detected in many streptomyces species by its ability to specifically bind sequence motifs within the promoter of a streptomyces pristinaespiralis multidrug resistance gene (ptr). to investigate the possible role of pip in regulating multidrug resistance, it was purified from a genetically characterized species, streptomyces coelicolor, utilizing an affinity matrix of the ptr promoter conjugated to magnetic beads. reverse genetics i ...200111050092
genetic analysis of carboxydothermus hydrogenoformans carbon monoxide dehydrogenase genes coof and coos.carboxydothermus hydrogenoformans is an extremely thermophilic, gram-positive bacterium growing on carbon monoxide (co) as single carbon and energy source and producing only h(2) and co(2). carbon monoxide dehydrogenase is a key enzyme for co metabolism. the carbon monoxide dehydrogenase genes coof and coos from c. hydrogenoformans were cloned and sequenced. these genes showed the highest similarity to the coof genes from the archaeon archaeoglobus fulgidus and the coos gene from the bacterium r ...200011024270
unprecedented proximal binding of nitric oxide to heme: implications for guanylate cyclase.microbial cytochromes c' contain a 5-coordinate his-ligated heme that forms stable adducts with nitric oxide (no) and carbon monoxide (co), but not with dioxygen. we report the 1.95 and 1.35 a resolution crystal structures of the co- and no-bound forms of the reduced protein from alcaligenes xylosoxidans. no disrupts the his-fe bond and binds in a novel mode to the proximal face of the heme, giving a 5-coordinate species. in contrast, co binds 6-coordinate on the distal side. a second co molecul ...200011060017
interactions of the nadp(h)-binding domain iii of proton-translocating transhydrogenase from escherichia coli with nadp(h) and the nad(h)-binding domain i studied by nmr and site-directed mutagenesis.using the purified nadp(h)-binding domain of proton-translocating escherichia coli transhydrogenase (eciii) overexpressed in (15)n- and (2)h-labeled medium, together with the purified nad(h)-binding domain from e. coli (eci), the interface between eciii and eci, the nadp(h)-binding site and the influence on the interface by nad(p)(h) was investigated in solution by nmr chemical shift mapping. mapping of the nadp(h)-binding site showed that the nadp(h) substrate is bound to eciii in an extended c ...200011027139
a new appraisal of the prokaryotic origin of eukaryotic phytochromes.the evolutionary origin of the phytochromes of eukaryotes is controversial. three cyanobacterial proteins have been described as "phytochrome-like" and have been suggested to be potential ancestors of these essential photoreceptors: cph1 from synechocystis pcc 6803, showing homology to phytochromes along its entire length and known to attach a chromophore; and plpa from synechocystis pcc 6803 and rcae from fremyella diplosiphon, both showing homology to phytochromes most strongly only in the c-t ...200011029065
role of the dinitrogenase reductase arginine 101 residue in dinitrogenase reductase adp-ribosyltransferase binding, nad binding, and cleavage.dinitrogenase reductase is posttranslationally regulated by dinitrogenase reductase adp-ribosyltransferase (drat) via adp-ribosylation of the arginine 101 residue in some bacteria. rhodospirillum rubrum strains in which the arginine 101 of dinitrogenase reductase was replaced by tyrosine, phenylalanine, or leucine were constructed by site-directed mutagenesis of the nifh gene. the strain containing the r101f form of dinitrogenase reductase retains 91%, the strain containing the r101y form retain ...200111114923
energy transfer and charge separation in the purple non-sulfur bacterium roseospirillum parvum.the antenna reaction centre system of the recently described purple non-sulfur bacterium roseospirillum parvum strain 930i was studied with various spectroscopic techniques. the bacterium contains bacteriochlorophyll (bchl) a, 20% of which was esterified with tetrahydrogeranylgeraniol. in the near-infrared, the antenna showed absorption bands at 805 and 909 nm (929 nm at 6 k). fluorescence bands were located at 925 and 954 nm, at 300 and 6 k, respectively. fluorescence excitation spectra and tim ...200011106774
the dimerization of folded monomers of ribulose 1,5-bisphosphate carboxylase/oxygenase.spontaneous refolding and reconstitution processes of dimeric ribulose 1,5-bisphosphate carboxylase/oxygenase (rubisco) from rhodospirillum rubrum have been investigated using size-exclusion high performance liquid chromatography (hplc), spectroscopic, and activity measurements. when the unfolded rubisco in guanidine hydrochloride is diluted at 4 degrees c, a folding intermediate (rubisco-i) is rapidly formed, which remains in an unstable monomeric state and gradually develops into folded monome ...200111092881
component of the rhodospirillum centenum photosensory apparatus with structural and functional similarity to methyl-accepting chemotaxis protein chemoreceptors.photosynthetic bacteria respond to alterations in light conditions by migrating to locations that allows optimal use of light as an energy source. studies have indicated that photosynthesis-driven electron transport functions as an attractant signal for motility among purple photosynthetic bacteria. however, it is unclear just how the motility-based signal transduction system monitors electron flow through photosynthesis-driven electron transport. recently, we have demonstrated that the purple p ...200111114914
co sensing and regulation of gene expression by the transcriptional activator cooa.the transcriptional activator cooa from rhodospirillum rubrum contains a six-coordinate protoheme that acts as a co sensor in vivo. co is a physiological effector of cooa and replaces one of the axial ligands of the ferrous heme to form the co-bound cooa that is active as the transcriptional activator. cys75 or his77 is coordinated to the ferric and ferrous hemes in cooa, respectively. the redox-controlled ligand exchange between cys75 and his77 proceeds during the change in the redox state of t ...200011132638
maintenance and control of redox poise in rhodobacter capsulatus strains deficient in the calvin-benson-bassham pathway.carbon dioxide serves as the preferred electron acceptor during photoheterotrophic growth of nonsulfur purple photosynthetic bacteria such as rhodobacter capsulatus and rhodobacter sphaeroides. this co2, produced as a result of the oxidation of preferred organic carbon sources, is reduced through reactions of the calvin-benson-bassham reductive pentose phosphate pathway. this pathway is thus crucial to maintain a balanced intracellular oxidation-reduction potential (or redox poise) under photohe ...200011131022
effect of p(ii) and its homolog glnk on reversible adp-ribosylation of dinitrogenase reductase by heterologous expression of the rhodospirillum rubrum dinitrogenase reductase adp-ribosyl transferase-dinitrogenase reductase-activating glycohydrolase regulatory system in klebsiella pneumoniae.reversible adp-ribosylation of dinitrogenase reductase, catalyzed by the dinitrogenase reductase adp-ribosyl transferase-dinitrogenase reductase-activating glycohydrolase (drat-drag) regulatory system, has been characterized in rhodospirillum rubrum and other nitrogen-fixing bacteria. to investigate the mechanisms for the regulation of drat and drag activities, we studied the heterologous expression of r. rubrum dratg in klebsiella pneumoniae glnb and glnk mutants. in k. pneumoniae wild type, th ...200111160092
isolation and characterization of drat mutants that have altered regulatory properties of dinitrogenase reductase adp-ribosyltransferase in rhodospirillum rubrum.in rhodospirillum rubrum, dinitrogenase reductase adp-ribosyltransferase (drat) is responsible for the adp-ribosylation of dinitrogenase reductase in response to the addition of nh(+)(4) or removal from light, resulting in a decrease in nitrogenase activity. drat is itself subject to post-translational regulation; to investigate the mechanism for the regulation of drat activity, random pcr mutagenesis of drat (encoding drat) was performed and mutants with altered drat regulation were screened. t ...200111160813
n2-dependent growth and nitrogenase activity in the metal-metabolizing bacteria, geobacter and magnetospirillum species.cells of geobacter metallireducens, magnetospirillum strain amb-1, magnetospirillum magnetotacticum and magnetospirillum gryphiswaldense showed n2-dependent growth, the first anaerobically with fe(iii) as the electron acceptor, and the latter three species microaerobically in semi-solid oxygen gradient cultures. cells of the magnetospirillum species grown with n2 under microaerobic conditions were magnetotactic and therefore produced magnetosomes. cells of geobacter metallireducens reduced acety ...200011200427
nickel-binding proteins.nickel enzymes are a relatively new class of metalloenzymes. the seven known nickel enzymes are urease, hydrogenase, co-dehydrogenase, methyl-coenzyme m reductase, ni-superoxide dismutase, glyoxalase i and cis-trans isomerase. the requirement for nickel implies the presence of a nickel-processing system, since free transition metals are harmful to the cell. a nickel-processing system involves the recognition and transport of nickel into the cell and the handling of the nickel once it enters the ...199911212309
the crystal structure of an asymmetric complex of the two nucleotide binding components of proton-translocating transhydrogenase.membrane-bound ion translocators have important functions in biology, but their mechanisms of action are often poorly understood. transhydrogenase, found in animal mitochondria and bacteria, links the redox reaction between nad(h) and nadp(h) to proton translocation across a membrane. linkage is achieved through changes in protein conformation at the nucleotide binding sites. the redox reaction takes place between two protein components located on the membrane surface: di, which binds nad(h), an ...200111250201
an unusual pathway of excitation energy deactivation in carotenoids: singlet-to-triplet conversion on an ultrafast timescale in a photosynthetic antenna.carotenoids are important biomolecules that are ubiquitous in nature and find widespread application in medicine. in photosynthesis, they have a large role in light harvesting (lh) and photoprotection. they exert their lh function by donating their excited singlet state to nearby (bacterio)chlorophyll molecules. in photosynthetic bacteria, the efficiency of this energy transfer process can be as low as 30%. here, we present evidence that an unusual pathway of excited state relaxation in caroteno ...200111226245
a change in ionization of the nadp(h)-binding component (diii) of proton-translocating transhydrogenase regulates both hydride transfer and nucleotide release.transhydrogenase couples the transfer of hydride-ion equivalents between nad(h) and nadp(h) to proton translocation across a membrane. the enzyme has three components: di binds nad(h), diii binds nadp(h) and dii spans the membrane. coupling between transhydrogenation and proton translocation involves changes in the binding of nadp(h). mixtures of isolated di and diii from rhodospirillum rubrum transhydrogenase catalyse a rapid, single-turnover burst of hydride transfer between bound nucleotides; ...200111231296
assembled f1-(alpha beta ) and hybrid f1-alpha 3beta 3gamma -atpases from rhodospirillum rubrum alpha, wild type or mutant beta, and chloroplast gamma subunits. demonstration of mg2+versus ca2+-induced differences in catalytic site structure and function.refolding together the expressed alpha and beta subunits of the rhodospirillum rubrum f(1)(rf(1))-atpase led to assembly of only alpha(1)beta(1) dimers, showing a stable low mgatpase activity. when incubated in the presence of alcl(3), naf and either mgad(t)p or caad(t)p, all dimers associated into closed alpha(3)beta(3) hexamers, which also gained a low caatpase activity. both hexamer atpase activities exhibited identical rates and properties to the open dimer mgatpase. these results indicate t ...200111278351
ribulose-1,5-bisphosphate carboxylase/oxygenase from thermococcus kodakaraensis kod1. 200111265476
formation and properties of hybrid photosynthetic f1-atpases. demonstration of different structural requirements for stimulation and inhibition by tentoxin.a hybrid atpase composed of cloned chloroplast atp synthase beta and gamma subunits (betac and gammac) and the cloned alpha subunit from the rhodospirillum rubrum atp synthase (alphar) was assembled using solubilized inclusion bodies and a simple single-step folding procedure. the catalytic properties of the assembled alpha3rbeta3cgammac were compared to those of the core alpha3cbeta3cgammac complex of the native chloroplast coupling factor 1 (cf1) and to another recently described hybrid enzyme ...200111277942
binding of co at the pro2 side is crucial for the activation of co-sensing transcriptional activator cooa. (1)h nmr spectroscopic studies.cooa is a heme-containing transcriptional activator that anaerobically binds to dna at co atmosphere. to obtain information on the conformational transition of cooa induced by co binding to the heme, we assigned ring current-shifted (1)h nmr signals of cooa using two mutants whose axial ligands of the heme were replaced. in the absence of co, the nmr spectral pattern of h77y cooa, in which the axial histidine (his(77)) was replaced with tyrosine, was similar to that of wild-type cooa. in contras ...200111278259
regulation of nitrogenase in the photosynthetic bacterium rhodobacter sphaeroides containing dratg and nifhdk genes from rhodobacter capsulatus.the photosynthetic bacteria rhodobacter capsulatus and rhodospirillum rubrum regulate their nitrogenase activity by the reversible adp-ribosylation of nitrogenase fe-protein in response to ammonium addition or darkness. this regulation is mediated by two enzymes, dinitrogenase reductase adp-ribosyl transferase (drat) and dinitrogenase reductase activating glycohydrolase (drag). recently, we demonstrated that another photosynthetic bacterium, rhodobacter sphaeroides, appears to have no dratg gene ...200111315111
excitation trap approach to analyze size and pigment-pigment coupling: reconstitution of lh1 antenna of rhodobacter sphaeroides with ni-substituted bacteriochlorophyll.replacement of the central mg in chlorophylls by ni opens an ultrafast (tens of femtoseconds time range) radiationless de-excitation path, while the principal ground-state absorption and coordination properties of the pigment are retained. a method has been developed for substituting the native bacteriochlorophyll a by ni-bacteriochlorophyll a ([ni]-bchl) in the light harvesting antenna of the core complex (lh1) from the purple bacterium, rhodobacter (rb.) sphaeroides, to investigate its unit si ...200111297443
rhodocista centenaria vs rhodospirillum centenum: a reply to gest and favinger. 200111321119
resonance raman studies of cytochrome c' support the binding of no and co to opposite sides of the heme: implications for ligand discrimination in heme-based sensors.resonance raman (rr) studies have been conducted on alcaligenes xylosoxidans cytochrome c', a mono-his ligated hemoprotein which reversibly binds no and co but not o(2). recent crystallographic characterization of this protein has revealed the first example of a hemoprotein which can utilize both sides of its heme (distal and proximal) for binding exogenous ligands to its fe center. the present rr investigation of the fe coordination and heme pocket environments of ferrous, carbonyl, and nitrosy ...200111300792
taxonomic ambiguities: a case history. 200111321118
evidence for protein dielectric relaxations in reaction centers associated with the primary charge separation detected from rhodospirillum rubrum chromatophores by combined photovoltage and absorption measurements in the 1-15 ns time range.fast photovoltage measurements in rhodospirillum rubrum chromatophores in the nanosecond time range, escorted by time-resolved absorption measurements, are described. under reducing conditions, the photovoltage decayed significantly faster than the spectroscopically detected charge recombination of the radical pair p(+)h(a)(-). this indicates the occurrence of considerable dielectric relaxations. our data and data from the literature were analyzed by means of a reaction scheme consisting of thre ...200111318653
role of the core region of the pufx protein in inhibition of reconstitution of the core light-harvesting complexes of rhodobacter sphaeroides and rhodobacter capsulatus.pufx, the protein encoded by the pufx gene of rhodobacter capsulatus and rhodobacter sphaeroides, has been further characterized. the mature forms of these proteins contain 9 and 12 fewer amino acids, respectively, at the c-terminal end of the protein than are encoded by their pufx genes. to identify the portion of pufx responsible for inhibition of lh1 formation in reconstitution experiments, different regions (n-terminus and several core regions containing different lengths of the c-terminus) ...200111341824
equilibrium and kinetic parameters for the binding of inhibitors to the qb pocket in bacterial chromatophores: dependence on the state of qa.the equilibrium and kinetic parameters for the binding of various inhibitors to the q(b) pocket of the bacterial reaction center were investigated in chromatophores from rhodobacter capsulatus and rhodobacter sphaeroides. by monitoring the near-ir absorption changes specific to q(a)(-) and q(b)(-), we measured the fraction of inhibited centers in the dark and the kinetics and extent of inhibitor displacement after one flash due to the formation of the q(a)q(b)(-) state. the inhibitor release rat ...200111327844
phosphorylation of lhi beta during membrane synthesis in the photosynthetic bacterium rhodovulum sulfidophilum.cells of rhv. sulfidophilum were grown under different conditions in the presence of 32p-phosphate and the corresponding h and l membrane fractions obtained and fractionated by sds-page. both membranes showed almost identical polypeptide composition. the bacteriochlorophyll (bchl) specific content in h was always lower that in l. as described before, oxygen did not regulate gene expression. under high light, an almost two- to threefold decrease of the cellular specific bchl content was observed. ...200111400052
a bacteriochlorophyll a antenna complex from purple bacteria absorbing at 963 nm.a recently isolated species of the photosynthetic purple sulfur bacteria, provisionally called strain 970, was investigated with respect to its antenna function by means of various spectroscopic techniques, including fluorescence and pump-probe absorption difference spectroscopy. the bacterium contains bacteriochlorophyll a and an as yet unidentified carotenoid, perhaps 3,4,3',4'-tetrahydrospirilloxanthin. it has a single antenna complex of the lh1 type, with a q(y) absorption band situated at t ...200111331023
the heterotrimer of the membrane-peripheral components of transhydrogenase and the alternating-site mechanism of proton translocation.transhydrogenase undergoes conformational changes to couple the redox reaction between nad(h) and nadp(h) to proton translocation across a membrane. the protein comprises three components: di, which binds nad(h); diii, which binds nadp(h); and dii, which spans the membrane. experiments using isothermal titration calorimetry, analytical ultracentrifugation, and small angle x-ray scattering show that, as in the crystalline state, a mixture of recombinant di and diii from rhodospirillum rubrum tran ...200111399770
redox-mediated transcriptional activation in a cooa variant.cooa, the carbon monoxide-sensing transcription factor from rhodospirillum rubrum, binds co at a reduced (fe(ii)) heme moiety with resulting conformational changes that promote dna binding. in this study, we report a variant of cooa, m124r, that is active in transcriptional activation in a redox-dependent manner. where wild-type cooa is active only in the fe(ii) + co form, m124r cooa is active in both fe(ii) + co and fe(iii) forms. analysis of the ph dependence of the activity of fe(iii) m124r c ...200111359778
soret-excited raman spectroscopy of the spinach cytochrome b6f complex. structures of the b- and c-type hemes, chlorophyll a, and beta-carotene.soret-excited resonance raman (rr) spectra of the spinach cytochrome b6f complex (cyt b6f) are reported for the oxidized, native, ascorbate-reduced, and dithionite-reduced forms. using excitations at 441.6, 413.1, and 406.7 nm, rr contributions of chlorophyll a, beta-carotene, the c-type heme of cytochrome f, and the b-type hemes of cytochrome b6 of the b6f complex were identified and the data compared to those previously obtained for the rhodospirillum rubrum bc1 complex [le moigne, c., schoepp ...200111401579
redox-dependent co2 reduction activity of co dehydrogenase from rhodospirillum rubrum.carbon monoxide dehydrogenase (codh) from rhodospirillum rubrum catalyzes both the oxidation of co and the reduction of co(2). studies of the redox dependence of co(2) reduction by r. rubrum codh show that (1) codh is unable to catalyze co(2) reduction at potentials greater than -300 mv; (2) the maximum activity is observed at potentials less than -480 mv; and (3) the midpoint potential (e(m)) of the transition from minimum to maximum co(2) reduction activity occurs at approximately -339 mv. the ...200111412114
role of the atp synthase alpha-subunit in conferring sensitivity to tentoxin.tentoxin, produced by phytopathogenic fungi, selectively affects the function of the atp synthase enzymes of certain sensitive plant species. binding of tentoxin to a high affinity (k(i) approximately 10 nm) site on the chloroplast f(1) (cf(1)) strongly inhibits catalytic function, whereas binding to a second, lower affinity site (k(d) > 10 microm) leads to restoration and even stimulation of catalytic activity. sensitivity to tentoxin has been shown to be due, in part, to the nature of the amin ...200111412108
ligand-switching intermediates for the co-sensing transcriptional activator cooa measured by pulse radiolysis.cooa is a heme-containing and co-sensing transcriptional activator whose activity is regulated by co. the protoheme that acts as a co sensor in cooa shows unique properties for its coordination structure. the cys75 axial ligand of the ferric heme is replaced by his77 upon the reduction of the heme iron and vice versa. in this work, the ligand-switching process induced by the reduction of the heme was investigated by the technique of pulse radiolysis. hydrated electron reduced the heme iron in fe ...200111487580
redox-dependent activation of co dehydrogenase from rhodospirillum rubrum.studies of initial activities of carbon monoxide dehydrogenase (codh) from rhodospirillum rubrum show that codh is mostly inactive at redox potentials higher than -300 mv. initial activities measured at a wide range of redox potentials (0--500 mv) fit a function corresponding to the nernst equation with a midpoint potential of -316 mv. previously, extensive epr studies of codh have suggested that codh has three distinct redox states: (i) a spin-coupled state at -60 to -300 mv that gives rise to ...200111416171
methionine oxidation and its effect on the stability of a reconstituted subunit of the light-harvesting complex from rhodospirillum rubrum.an additional component in the purified core light-harvesting complex (lh1) from wild-type purple photosynthetic bacterium rhodospirillum rubrum has been identified as an oxidized species of alpha-polypeptide by maldi-tof mass spectrometry. this component appears as a slightly earlier-eluting peak in the rp-hplc chromatogram compared with the authentic alpha-polypeptide. the oxidation site has been determined to be the n-terminal methionine residue by high-resolution nmr spectroscopy, where the ...200111422366
purification and characterization of membrane-associated cooc protein and its functional role in the insertion of nickel into carbon monoxide dehydrogenase from rhodospirillum rubrum.the accessory protein cooc, which contains a nucleotide-binding domain (p-loop) near the n terminus, participates in the maturation of the nickel center of carbon monoxide dehydrogenase (codh). in this study, cooc was purified from the chromatophore membranes of rhodospirillum rubrum with a 3,464-fold purification and a 0.8% recovery, and its biochemical properties were characterized. cooc is a homodimer with a molecular mass of 61-63 kda, contains less than 0.1 atom of ni(2+) or fe(2+) per dime ...200111507093
mapping cooa.rna polymerase interactions. identification of activating regions 2 and 3 in cooa, the co-sensing transcriptional activator.cooa is a co-sensing protein that activates the transcription of genes encoding the co-oxidation (coo) regulon, whose polypeptide products are required for utilizing co as an energy source in rhodospirillum rubrum. cooa binds to a position overlapping the -35 element of the p(coof) promoter, similar to the arrangement of class ii crp (camp receptor protein)- and fnr (fumarate and nitrate reductase activator protein)-dependent promoters when expressed in escherichia coli. gain-of-function cooa va ...200111522788
the evolution of glutathione metabolism in phototrophic microorganisms.of the many roles ascribed to glutathione (gsh) the one most clearly established is its role in the protection of higher eucaryotes against oxygen toxicity through destruction of thiol-reactive oxygen byproducts. if this is the primary function of gsh then gsh metabolism should have evolved during or after the evolution of oxygenic photosynthesis. that many bacteria do not produce gsh is consistent with this view. in the present study we have examined the low-molecular-weight thiol compositio ...198711542078
cooa: a heme-containing regulatory protein that serves as a specific sensor of both carbon monoxide and redox state.cooa, the heme-containing carbon monoxide (co) sensor from the bacterium rhodospirillum rubrum, is a transcriptional factor that activates expression of certain genes in response to co. as with other heme proteins, cooa is unable to bind co when the fe heme is oxidized, consistent with the fact that some of the regulated gene products are oxygen-labile. upon reduction, there is an unusual switch of protein ligands to the six-coordinate heme and the reduced heme is able to bind co. co binding sta ...200111525385
the phylogeny of purple bacteria: the alpha subdivision.the technique of oligonucleotide cataloging shows the purple photosynthetic eubacteria to comprise three major subdivisions, temporarily called alpha, beta, and gamma--previously designated groups i-iii by gibson et al. (1979). each subdivision contains a number of non-photosynthetic genera in addition to the photosynthetic ones. the alpha subdivision, the subject of the present report, contains most but not all of the species that fall into the classically defined genera rhodospirillum, rhodo ...198411541974
melissa: a potential experiment for a precursor mission to the moon.melissa (micro-ecological life support system alternative) has been conceived as a micro-organism based ecosystem intended as a tool for developing the technology for a future artificial ecosystem for long term space missions, as for example a lunar base. the driving element of melissa is the recovering of edible biomass from waste, co2, and minerals with the use of sun light as energy source. in this publication, we focus our attention on the potential applications of melissa for a precursor mi ...199611543311
effects of specific amino acid substitutions on activities of dinitrogenase reductase-activating glycohydrolase from rhodospirillum rubrum.site-directed mutagenesis of the drag gene was used to generate altered forms of dinitrogenase reductase-activating glycohydrolase (drag) with d123a, h142l, h158n, d243g, and e279r substitutions. the amino acid residues h142 and e279 are not required either for the coordination to the metal center or for catalysis since the variants h142l and e279r retained both catalytic and electron paramagnetic resonance spectral properties similar to those of the wild-type enzyme. since drag-h158n and drag-d ...200111544238
phylogenetic origins of the plant mitochondrion based on a comparative analysis of 5s ribosomal rna sequences.the complete nucleotide sequences of 5s ribosomal rnas from rhodocyclus gelatinosa, rhodobacter sphaeroides, and pseudomonas cepacia were determined. comparisons of these 5s rna sequences show that rather than being phylogenetically related to one another, the two photosynthetic bacterial 5s rna sequences show that rather than being phylogenetically related to one another, the two photosynthetic bacterial 5s rnas share more sequence and signature homology with the rnas of two nonphotosynthetic ...198511542018
detection of bacteria in environmental samples by direct pcr without dna extraction.cultured cells and environmental samples were used directly in pcrs without the isolation of dna. serial dilution was used to eliminate the inhibitory effect of materials in natural samples. primers specific for pmoa, which encodes a subunit of the particulate methane monooxygenase, were used to detect and quantify methanotrophic bacteria by direct most probable number pcr. phototrophic bacteria were detected in environmental samples by direct pcr with primers specific for pufm, and members of t ...200111570503
fast hydride transfer in proton-translocating transhydrogenase revealed in a rapid mixing continuous flow device.transhydrogenase couples the redox reaction between nad(h) and nadp(h) to proton translocation across a membrane. coupling is achieved through changes in protein conformation. upon mixing, the isolated nucleotide-binding components of transhydrogenase (di, which binds nad(h), and diii, which binds nadp(h)) form a catalytic di(2).diii(1) complex, the structure of which was recently solved by x-ray crystallography. the fluorescence from an engineered trp in diii changes when bound nadp(+) is reduc ...200111577115
the heme pocket afforded by gly117 is crucial for proper heme ligation and activity of cooa.cooa, a co-sensing homodimeric transcription activator from rhodospirillum rubrum, undergoes a conformational change in response to co binding to its heme prosthetic group that allows it to bind specific dna sequences. in a recent structural study (lanzilotta, w. n., schuller, d. j., thorsteinsson, m. v., kerby, r. l., roberts, g. p., and poulos, t. l. (2000) nat. struct. biol. 7, 876-880), it was suggested that co binding to cooa results in a modest repositioning of the c-helices that serve as ...200111551932
conformational dynamics of the transcriptional regulator cooa protein studied by subpicosecond mid-infrared vibrational spectroscopy.cooa, which is a transcriptional regulator heme protein allosterically triggered by co, is studied by femtosecond visible-pump mid-ir-probe spectroscopy. transient bleaching upon excitation of the heme in the soret band is detected at approximately 1979 cm(-1), which is the absorption region of the co bound to the heme. the bleach signal shows a nonexponential decay with time constants of 56 and 290 ps, caused by the rebinding of the co to the heme. about 98% of dissociated co recombines geminat ...200111592884
life on carbon monoxide: x-ray structure of rhodospirillum rubrum ni-fe-s carbon monoxide dehydrogenase.a crystal structure of the anaerobic ni-fe-s carbon monoxide dehydrogenase (codh) from rhodospirillum rubrum has been determined to 2.8-a resolution. the codh family, for which the r. rubrum enzyme is the prototype, catalyzes the biological oxidation of co at an unusual ni-fe-s cluster called the c-cluster. the ni-fe-s c-cluster contains a mononuclear site and a four-metal cubane. surprisingly, anomalous dispersion data suggest that the mononuclear site contains fe and not ni, and the four-metal ...200111593006
functional characterization of three glnb homologs in the photosynthetic bacterium rhodospirillum rubrum: roles in sensing ammonium and energy status.the glnb (p(ii)) protein, the product of glnb, has been characterized previously in the photosynthetic bacterium rhodospirillum rubrum. here we describe identification of two other p(ii) homologs in this organism, glnk and glnj. although the sequences of these three homologs are very similar, the molecules have both distinct and overlapping functions in the cell. while glnb is required for activation of nifa activity in r. rubrum, glnk and glnj do not appear to be involved in this process. in co ...200111591658
supramolecular arrangement of rhodospirillum rubrum b880 holochrome as studied by radiation inactivation and electron paramagnetic resonance.oxidation of the b880 antenna holochrome gives rise to a 3.8-g linewidth electron paramagnetic resonance (epr) signal that is considerably narrower than the 13-g signal of monomeric bacteriochlorophyll (bchl) cation. radiation inactivation was used to verify a model according to which this linewidth narrowing is due to delocalization over several bchl molecules. chromatophores of the photoreaction centerless mutant f24 of rhodospirillum rubrum were subjected to different doses of gamma-radiation ...199011607076
oligomerization of light-harvesting i antenna peptides of rhodospirillum rubrum.we investigated the oligomerization of the core light-harvesting complex (lh1) of rhodospirillum rubrum from the separated alpha beta bchl(2) subunits (b820) and the oligomerization of the b820 subunit from its monomeric peptides. the full lh1 complex was reversibly associated from b820 subunits by either varying the temperature in the range 277-300 k or by varying the detergent concentration in the buffer from 0.36 to 0.52% n-octyl-beta-d-glucopyranoside. temperature-induced transition measurem ...200111669628
time-dependent changes in the carotenoid composition and preferential binding of spirilloxanthin to the reaction center and anhydrorhodovibrin to the lh1 antenna complex in rhodobium marinum.carotenoids were isolated from the cells of rhodobium marinum, and their structures were determined by mass spectrometry and 1h nuclear magnetic resonance spectroscopy; the carotenoids include lycopene, rhodopin, anhydrorhodovibrin, rhodovibrin and spirilloxanthin. time-dependent changes in the carotenoid composition in the reaction center (rc) and the light-harvesting complex 1 (lh1) were traced by high-performance liquid chromatography analysis of the extracts. the carotenoid composition chang ...200111594059
plastome-encoded bacterial ribulose-1,5-bisphosphate carboxylase/oxygenase (rubisco) supports photosynthesis and growth in tobacco.the efficiency with which crop plants use their resources of light, water, and fertilizer nitrogen could be enhanced by replacing their co(2)-fixing enzyme, d-ribulose-1,5-bisphosphate carboxylase-oxygenase (rubisco), with more efficient forms, such as those found in some algae, for example. this important challenge has been frustrated by failure of all previous attempts to substitute a fully functional, foreign rubisco (efficient or inefficient) into higher plants. this failure could be caused ...200111724961
photophysical characterization of natural cis-carotenoids.by means of steady-state fluorescence spectroscopy we explore the photophysics of two lowest lying singlet excited states in two natural 15-cis-carotenoids, namely phytoene and phytofluene, possessing three and five conjugated double bonds (n), respectively. the results are interpreted in relation to the photophysics of all-transcarotenoids with varying n. the fluorescence of phytofluene is more stokes-shifted relative to that of phytoene, and is ascribed to the forbidden s1-->s0 transition, wit ...200111683034
evidence for three distinct hydrogenase activities in rhodospirillum rubrum.inducer, inhibitor, and mutant studies on three hydrogenase activities of rhodospirillum rubrum indicate that they are mediated by three distinct hydrogenase enzymes. uptake hydrogenase mediates h2 uptake to an unknown physiological acceptor or methylene blue and is maximally synthesized during autotrophic growth in light. formate-linked hydrogenase is synthesized primarily during growth in darkness or when light becomes limiting, and links formate oxidation to h2 production. carbon-monoxide-lin ...200111778889
structure of cytochrome c2 from rhodospirillum centenum.cytochrome c(2) from the purple photosynthetic bacterium rhodospirillum centenum has been crystallized by the sitting-drop vapour-diffusion method. the crystals belong to the orthorhombic space group p2(1)2(1)2(1), with unit-cell parameters a = 29.7, b = 59.9, c = 65.4 a, and diffract to a resolution limit of 1.7 a. the fe-atom position was determined from its anomalous scattering contribution and a molecular-replacement solution was calculated. the correctness of the solution was confirmed by p ...200111679712
the 7.5-a electron density and spectroscopic properties of a novel low-light b800 lh2 from rhodopseudomonas palustris.a novel low-light (ll) adapted light-harvesting complex ii has been isolated from rhodopseudomonas palustris. previous work has identified a ll b800-850 complex with a heterogeneous peptide composition and reduced absorption at 850 nm. the work presented here shows the 850 nm absorption to be contamination from a high-light b800-850 complex and that the true ll light-harvesting complex ii is a novel b800 complex composed of eight alpha beta(d) peptide pairs that exhibits unique absorption and ci ...200211806936
spectroscopic studies of nickel-deficient carbon monoxide dehydrogenase from rhodospirillum rubrum: nature of the iron-sulfur clusters.carbon monoxide dehydrogenase (codh) from rhodospirillum rubrum utilizes three types of fe-s clusters to catalyze the reversible oxidation of co to co(2): a novel [ni4fe5s] active site (c cluster) and two distinct [4fe4s] electron-transfer sites (b and d clusters). while recent x-ray data show the geometric arrangement of the five metal centers at the c cluster, electronic structures of the various [ni4fe5s] oxidation states remain ambiguous. these studies report magnetic circular dichroism (mcd ...200211814363
hydrophobicity and prediction of the secondary structure of membrane proteins and peptides.reliability of the hydropathy method to predict the formation of membrane-spanning alpha-helices by integral membrane proteins and peptides whose structure is known from x-ray crystallography is analysed. it is shown that kyte-doolittle hydropathy plots do not predict accurately 22 transmembrane alpha-helices in the reaction centres (rc) of the photosynthetic bacteria rhodopseudomonas viridis and rhodobacter sphaeroides (r-26). the accuracy of prediction for these proteins was improved using an ...200111699870
structure of the rhodobacter sphaeroides light-harvesting 1 beta subunit in detergent micelles.the light harvesting 1 antenna (lh1) complex from rhodobacter sphaeroides funnels excitation energy to the photosynthetic reaction center. our ultimate goal is to build up the structure of lh1 from structures of its individual subunits, much as the antenna can self-assemble from its components in membrane-mimicking detergent micelles. the beta subunit adopts a nativelike conformation in zwittergent 3:12 micelles as demonstrated by its ability to take the first step of assembly, binding bchl a. m ...200211772000
effect of carotenoids on the interaction between pigment-protein complexes in membranes of the sulfur photosynthetic bacterium chromatium minutissimum. 200111813560
unsuspected diversity among marine aerobic anoxygenic phototrophs.aerobic, anoxygenic, phototrophic bacteria containing bacteriochlorophyll a (bchla) require oxygen for both growth and bchla synthesis. recent reports suggest that these bacteria are widely distributed in marine plankton, and that they may account for up to 5% of surface ocean photosynthetic electron transport and 11% of the total microbial community. known planktonic anoxygenic phototrophs belong to only a few restricted groups within the proteobacteria alpha-subclass. here we report genomic an ...200211832943
excitons in a photosynthetic light-harvesting system: a combined molecular dynamics, quantum chemistry, and polaron model study.the dynamics of pigment-pigment and pigment-protein interactions in light-harvesting complexes is studied with an approach that combines molecular dynamics simulations with quantum chemistry calculations and a polaron model analysis. the molecular dynamics simulation of light-harvesting (lh) complexes was performed on an 87 055 atom system comprised of a lh-ii complex of rhodospirillum molischianum embedded in a lipid bilayer and surrounded with appropriate water layers. for each of the 16 b850 ...200211909121
calculability analysis in underdetermined metabolic networks illustrated by a model of the central metabolism in purple nonsulfur bacteria.metabolite balancing has turned out to be a powerful computational tool in metabolic engineering. however, the linear equation systems occurring in this analysis are often underdetermined. if it is difficult or impossible to find the missing constraints, it is nevertheless feasible in some cases to determine the values of a subset of the unknown rates. here, a procedure for finding out which reaction rates can be uniquely calculated in underdetermined metabolic networks and computing these rates ...200211835134
general random matrix approach to account for the effect of static disorder on the spectral properties of light harvesting systems.we develop a random matrix model approach to study static disorder in pigment-protein complexes in photosynthetic organisms. as a case study, we examine the ring of b850 bacteriochlorophylls in the peripheral light-harvesting complex of rhodospirillum molischianum, formulated in terms of an effective hamiltonian describing the collective electronic excitations of the system. we numerically examine and compare various models of disorder and observe that both the density of states and the absorpti ...200211909118
photosynthetic apparatus in roseateles depolymerans 61a is transcriptionally induced by carbon limitation.production of a photosynthetic apparatus in roseateles depolymerans 61a, a recently discovered freshwater beta-proteobacterium showing characteristics of aerobic phototrophic bacteria, was observed when the cells were subjected to a sudden decrease in carbon sources (e.g., when cells grown with 0.1 to 0.4% casamino acids were diluted or transferred into medium containing <or=0.04% casamino acids). accumulation of bacteriochlorophyll (bchl) a was observed in the presence of oxygen and was enhance ...200211916683
absorption and cd spectroscopy and modeling of various lh2 complexes from purple bacteria.the absorption (od) and circular dichroism (cd) spectra of lh2 complexes from various purple bacteria have been measured and modeled. based on the lineshapes of the spectra we can sort the lh2 complexes into two distinguishable groups: "acidophila"-like (type 1) and "molischianum"-like (type 2). starting from the known geometric structures of rhodopseudomonas (rps.) acidophila and rhodospirillum (rsp.) molischianum we can model the od and cd spectra of all species by just slightly varying some k ...200211916874
relationship between light-induced quenching of atebrin fluorescence and atp formation in rhodospirillum rubrum chromatophores. 197111945647
energy-linked changes of the membrane of rhodospirillum rubrum chromatophores detected by the fluorescent probe 8-anilinonaphthalene-1-sulfonic acid. 197111945994
rhodospirillum rubrum possesses a variant of the bchp gene, encoding geranylgeranyl-bacteriopheophytin reductase.the bchp gene product of rhodobacter sphaeroides is responsible for the reduction of the isoprenoid moiety of bacteriochlorophyll (bchl) from geranylgeraniol (gg) to phytol; here, we show that this enzyme also catalyzes the reduction of the isoprenoid moiety of bacteriopheophytin (bphe). in contrast, we demonstrate that a newly identified homolog of this gene in rhodospirillum rubrum encodes an enzyme, gg-bphe reductase, capable of reducing the isoprenoid moiety of bphe only. we propose that rho ...200211872709
isolation of a "basic membrane" fraction enriched in an ornithine-containing lipid, from a blue-green mutant of rhodospirillum rubrum. 197011945444
dissociation-produced loss of regulatory control of homoserine dehydrogenase of rhodospirillum rubrum. 196911946964
on the nature of the light-induced bacteriochlorophyll absorbance changes in chromatophores of rhodospirillum rubrum. 197311947102
bacterial photosynthesis begins with quantum-mechanical coherence.in the antenna system of photosynthetic bacteria, pigments form circular aggregates whose excitations are excitons with quantum-mechanical coherence extending over many pigments. these excitons play crucial roles in light harvesting, storage, and excitation-energy transfer (eet). eet takes place rapidly to and/or from optically forbidden exciton states, without total transition dipole, within the antenna system and to the reaction center. such eets cannot be rationalized by förster's formula, th ...200111933253
a coupling factor from rhodospirillum rubrum chromatophores. 197211946452
rhodospirillum centenum utilizes separate motor and switch components to control lateral and polar flagellum rotation.rhodospirillum centenum is a purple photosynthetic bacterium that is capable of differentiating from vibrioid swimming cells that contain a single polar flagellum into rod-shaped swarming cells that have a polar flagellum plus numerous lateral flagella. microscopic studies have demonstrated that the polar flagellum is constitutively present and that the lateral flagella are found only when the cells are grown on solidified or viscous medium. in this study, we demonstrated that r. centenum contai ...200211948156
partial resolution of energy-linked reactions in rhodospirillum rubrum chromatophores. 196911946959
the occurrence of hydroxy-derivatives of phytoene and phytofluene in diphenylamine-inhibited cultures of rhodospirillum rubrum. 197011947408
h+-pyrophosphatase of rhodospirillum rubrum. high yield expression in escherichia coli and identification of the cys residues responsible for inactivation my mersalyl.h(+)-translocating pyrophosphatase (h(+)-ppase) of the photosynthetic bacterium rhodospirillum rubrum was expressed in escherichia coli c43(de3) cells. recombinant h(+)-ppase was observed in inner membrane vesicles, where it catalyzed both pp(i) hydrolysis coupled with h(+) transport into the vesicles and pp(i) synthesis. the hydrolytic activity of h(+)-ppase in e. coli vesicles was eight times greater than that in r. rubrum chromatophores but exhibited similar sensitivity to the h(+)-ppase inhi ...200211956221
formation of glyoxylate from alpha-hydroxyglutarate by rhodospirillum rubrum. 197011947489
photosynthetic apparatus of purple bacteria. 200211997980
[modeling of energy migration and trapping in purple bacteria. analysis of extreme formulae].homogeneous pigment ensembles similar to those of purple bacteria rhodospirillum rubrum were studied. two formulae were advanced for the limiting values of excitation lifetime and quantum yield of excitation trapping in these ensembles, provided all reaction centers are in an active state. it was demonstrated by mathematical modeling that these limiting values strictly depend on three parameters of molecular ensembles: the numbers of core-bacteriochlorophyll molecules per reaction center, the va ...200211969165
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