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fluorescence studies with malate dehydrogenase from bradyrhizobium japonicum 3i1b-143 bacteroids: a two-tryptophan containing protein.a number of fluorescence studies, both of trp residues and bound nadh, have been reported for porcine malate dehydrogenase (mdh). the large number of trp residues (six) complicates the interpretation of some studies. to circumvent this we have performed studies with a two-tryptophan (per subunit) mdh from bradyrhizobium japonicum 3i1b-143 bacteroids. we have performed phase/modulation fluorescence lifetime measurements, as a function of temperature and added quencher ki, in order to resolved the ...19902241162
ammonia regulation of nod genes in bradyrhizobium japonicum.the expression of the nodd and nodyabc operons of bradyrhizobium japonicum is repressed by the addition of ammonia. repression of nodyabc expression is probably due to the effect on nodd since nodd positively regulates itself, as well as other nod operons. the effect of ammonia is independent of the known nitrogen regulatory protein, ntrc, and another regulatory protein for nitrogen fixation, nifa.19902250656
cloning and sequence of the salmonella typhimurium heml gene and identification of the missing enzyme in heml mutants as glutamate-1-semialdehyde aminotransferase.salmonella typhimurium forms the heme precursor delta-aminolevulinic acid (ala) exclusively from glutamate via the five-carbon pathway, which also occurs in plants and some bacteria including escherichia coli, rather than by ala synthase-catalyzed condensation of glycine and succinyl-coenzyme a, which occurs in yeasts, fungi, animal cells, and some bacteria including bradyrhizobium japonicum and rhodobacter capsulatus. ala-auxotrophic heml mutant s. typhimurium cells are deficient in glutamate-1 ...19902254275
the nifen genes participating in femo cofactor biosynthesis and genes encoding dinitrogenase are part of the same operon in bradyrhizobium species.the nucleotide sequences of genes homologous to the klebsiella pneumoniae nifen genes have been determined in bradyrhizobium japonicum 110. the coding regions for the nife and nifn consist, respectively, of 1641 and 1407 nucleotides. the nifd gene (coding for the beta-subunit of dinitrogenase) and nife are linked, and separated by 95 nucleotides. in the region of 12 nucleotides that separates nife from nifn the stop codon for nife overlaps the putative ribosome binding site for nifn. in contrast ...19902266945
cell-associated oligosaccharides of bradyrhizobium spp.we report the initial characterization of the cell-associated oligosaccharides produced by four bradyrhizobium strains: bradyrhizobium japonicum usda 110, usda 94, and atcc 10324 and bradyrhizobium sp. strain 32h1. the cell-associated oligosaccharides of these strains were found to be composed solely of glucose and were predominantly smaller than the cyclic beta-1,2-glucans produced by agrobacterium and rhizobium species. linkage studies and nuclear magnetic resonance analyses demonstrated that ...19902294083
isolation and characterization of the lipopolysaccharides from bradyrhizobium japonicum.the lipopolysaccharide (lps) of bradyrhizobium japonicum 61a123 was isolated and partially characterized. phenol-water extraction of strain 61a123 yielded lps exclusively in the phenol phase. the water phase contained low-molecular-weight glucans and extracellular or capsular polysaccharides. the lpss from b. japonicum 61a76, 61a135, and 61a101c were also extracted exclusively into the phenol phase. the lpss from strain usda 110 and its nod- mutant hs123 were found in both the phenol and water p ...19902318801
proposed regulatory pathway encoded by the nodv and nodw genes, determinants of host specificity in bradyrhizobium japonicum.bradyrhizobium japonicum is the root nodule endosymbiont of soybean (glycine max), mung bean (vigna radiata), cowpea (vigna unguiculata), and siratro (macroptilium atropurpureum). we report the characteristics of a nodulation-gene region of b. japonicum that contributes only marginally to the bacterium's ability to nodulate soybean but is essential for the nodulation of the three alternative hosts. this dna region consists of two open reading frames designated nodv and nodw. the predicted amino ...19902320582
cloning and sequencing the genes encoding uptake-hydrogenase subunits of rhodocyclus gelatinosus.rhodocyclus gelatinosus grew photosynthetically in the light and consumed h2 at a rate of about 665 nmol/min per mg protein. the uptake-hydrogenase (h2ase) was found to be membrane bound and insensitive to inhibition by co. the structural genes of r. gelatinosus uptake-h2ase were isolated from a 40 kb cosmid gene library of r. gelatinosus dna by hybridization with the structural genes of uptake-h2ase of bradyrhizobium japonicum and rhodobacter capsulatus. the r. gelatinosus genes were localized ...19902325631
a model of nitrogen flow by malonamate in rhizobium japonicum-soybean symbiosis.two types of novel malonamidases were found in soybean nodules. one (e1) catalyzes the formation of malonamate from malonate and its hydrolysis to ammonia, whereas the other (e2) acts mainly on the hydrolysis of malonamate. e1 and e2 were found in bacteroids, but only e2 was found in the plant cytosol of the nodule. the substrate requirements of e1 and e2 were highly specific for malonate and malonamate, respectively. from these and other results reported previously, we propose that malonamate p ...19902357226
effects of the photobleaching herbicide, acifluorfen-methyl, on protoporphyrinogen oxidation in barley organelles, soybean root mitochondria, soybean root nodules, and bacteria.the photobleaching herbicide, acifluorfen-methyl (afm), has been reported to be an inhibitor of the heme and chlorophyll biosynthetic enzyme protoporphyrinogen oxidase (protox) in several plant species. however, afm had no effect on the levels of protox activity measured in a mitochondrial fraction from soybean roots. in contrast, afm inhibited protox activity in etioplasts from barley leaves and in mitochondria from barley roots, but the extent of inhibition varied depending upon the assay cond ...19902369128
genome analysis of bradyrhizobium japonicum serocluster 123 field isolates by using field inversion gel electrophoresis.the genomes of 11 bradyrhizobium japonicum serocluster 123 field isolates were analyzed by using field inversion gel electrophoresis. genomic fingerprints produced by digestion of intact genomic dna in agarose plugs with the rare-cutting restriction enzymes asei, drai, spei, and xbai showed that the isolates were genetically diverse. few (30 to 50%) isolates exhibited the same fingerprint as the usda serogroup strain with which they are antigenically related. southern hybridization with a nifhd ...19902383015
genetic organization of the hydrogen uptake (hup) cluster from rhizobium leguminosarum.in symbiosis with peas, rhizobium leguminosarum upm791 induces the synthesis of a hydrogen uptake (hup) system that recycles hydrogen generated in nodules by nitrogenase. a cosmid (pal618) containing hup genes from this strain on a 20-kilobase-pair (kb) dna insert has previously been isolated in our laboratory (a. leyva, j. m. palacios, t. mozo, and t.ruiz-argüeso, j. bacteriol. 169:4929-4934, 1987). here we show that cosmid pal618 contains all of the genetic information required to confer high ...19902407728
differential transcription of the two glutamine synthetase genes of bradyrhizobium japonicum.bradyrhizobium japonicum induces the formation of nitrogen-fixing symbiotic root nodules on soybean plants. the b. japonicum genome encodes two isoforms of glutamine synthetase (gs). one form, gsi, encoded by the gene glna, is similar in structure and activity to the enzyme found in all other bacteria. the second form, gsii, encoded by glnii, is structurally related to the eucaryotic enzyme. genetic analyses indicate that glna or glnii alone is sufficient to provide glutamine prototrophy, wherea ...19872445733
two host-inducible genes of rhizobium fredii and characterization of the inducing compound.random transcription fusions with mu d1(kan lac) generated three mutants in rhizobium fredii (strain usda 201) which showed induction of beta-galactosidase when grown in root exudate of the host plants glycine max, phaseolus vulgaris, and vigna ungliculata. two genes were isolated from a library of total plasmid dna of one of the mutants, 3f1. these genes, present in tandem on a 4.2-kilobase hindiii fragment, appear in one copy each on the symbiotic plasmid and do not hybridize to the rhizobium ...19882447061
immunological relationship among hydrogenases.we examined the immunological cross-reactions of 11 different hydrogenase antigens with 9 different hydrogenase antibodies. included were antibodies and antigens of both subunits of the hydrogenases of bradyrhizobium japonicum and thiocapsa roseopersicina. the results showed a strong relationship among the ni-fe dimeric hydrogenases. the two subunits of ni-fe dimeric hydrogenases appeared immunologically distinct: specific interactions occurred only when antibodies to the 60- and 30-kilodalton s ...19892464579
o-antigen from bradyrhizobium japonicum lipopolysaccharide inhibits intercellular (symplast) communication between soybean (glycine max) cells.the technique of fluorescence redistribution after photobleaching was utilized to measure intercellular movement of low molecular weight fluorescent hydrophilic substances across the cell wall/membrane interface between contiguous soybean (glycine max (l.) merr. cv. mandarin) root cells (sb-1 cell line) in tissue culture. lipopolysaccharide (lps) purified from bradyrhizobium japonicum r110d, a gram-negative bacterium that normally infects and induces nodulation in soybean roots in vivo, inhibits ...19892473070
expression of bradyrhizobium japonicum nodulation gene in rhizobium fredii nod mutants.the b. japonicum usda 110 plafr1 gene library was transferred to tn5-induced rhizobium fredii usda 191-4 nod- mutants with helper plasmid prk2013. smr tcr transconjugants occurred with a frequency of 5 x 10(-4). the transconjugants were purified and used to inoculate germinated soybean seeds. seven nodules were obtained in the nodulation experiment. the fast-growing nod+ smr tcr rhizobium fredii strain was isolated from all nodules. each isolate had acquired a new plasmid with a molecular mass o ...19892491321
characterization of hydrogen-uptake activity in the hyperthermophile pyrodictium brockii.pyrodictium brockii is a hyperthermophilic archaebacterium with an optimal growth temperature of 105 degrees c. p. brockii is also a chemolithotroph, requiring h2 and co2 for growth. we have characterized p. brockii hydrogen-uptake activity with regard to temperature, ability to couple hydrogen oxidation to artificial electron acceptor reduction, sensitivity to o2, and cellular localization. the hydrogen-uptake activity was localized predominantly in a particulate fraction, was reversibly inhibi ...19892492097
protein phosphorylation in bradyrhizobium japonicum bacteroids and cultures.protein phosphorylation was demonstrated in bradyrhizobium japonicum bacteroids in vivo and in cultures in vivo and in vitro. comparison of in vivo-labeled phosphoproteins of bacteroids and of cultured cells showed differences in both the pattern and intensity of labeling. in cultured cells, comparison of the labeling patterns and intensities of in vivo- and in vitro-labeled phosphoproteins showed a number of similarities; however, several phosphoproteins were found only after one of the two lab ...19892498290
the bradyrhizobium japonicum fixbcx operon: identification of fixx and of a 5' mrna region affecting the level of the fixbcx transcript.the bradyrhizobium japonicum fixx gene was identified and shown to be essential for symbiotic and free-living, microaerobic nitrogen fixation. the fixx gene encodes a ferredoxin-like protein which may be involved in a redox process (electron transport?) essential for nitrogenase activity. this gene was localized downstream of fixc and its expression was dependent on the fixb promoter, providing evidence for the existence of a fixbcx operon. mutagenesis and sequence analysis of the unusually long ...19892503674
fine-tuning of nif and fix gene expression by upstream activator sequences in bradyrhizobium japonicum.the significance of bradyrhizobium japonicum upstream activator sequences (uass) for differential nifa-mediated fix and nif gene expression was investigated by two means: (i) hybrid fixa- and fixb-lacz fusions were constructed by transposing a nifh-uas cartridge in front of their promoters; and (ii) b. japonicum mutants were generated carrying specific chromosomal deletions or uas cartridge insertions within the fixa, fixb or nifh promoter-upstream regions. expression of fixa was not affected, a ...19892503675
production of extracellular nucleic acids by genetically altered bacteria in aquatic-environment microcosms.the factors which affect the production of extracellular dna by genetically altered strains of escherichia coli, pseudomonas aeruginosa, pseudomonas cepacia, and bradyrhizobium japonicum in aquatic environments were investigated. cellular nucleic acids were labeled in vivo by incubation with [3h]thymidine or [3h]adenine, and production of extracellular dna in marine waters, artificial seawater, or minimal salts media was determined by detecting radiolabeled macromolecules in incubation filtrates ...19892506807
identification, cloning, and sequence analysis of the nitrogen regulation gene ntrc of agrobacterium tumefaciens c58.we describe the cloning of an ntrc gene of agrobacterium tumefaciens c58 by interspecific complementation of an escherichia coli ntrc mutant. restriction mapping and southern blot analysis of the complementing clone identified a 1.7-kb ecori-pvuii dna fragment whose sequence was determined. analysis of this sequence revealed coding regions corresponding to a complete ntrc gene and the c-terminal region of an ntrb gene. amino acid sequence comparisons of a. tumefaciens ntrc protein with ntrc sequ ...19892520824
nodule-specific kinases phosphorylating nuclear factors in isolated nuclei.in vitro phosphorylation of total nuclear proteins from soybean (glycine max l) nodules formed by bradyrhizobium japonicum 61a76 showed several differences in comparison with those from uninfected roots or embryonic-axes nuclei. three types of protein phosphorylations were observed in nodule nuclei: ca(2+)- and calmodulin-independent, ca(2+)- and calmodulin-dependent, and ca(2+)-dependent but calmodulin-independent. in addition, ca(2+)-dependent dephosphorylation of some nuclear proteins was obs ...19892535508
identification and isolation of genes essential for h2 oxidation in rhodobacter capsulatus.mutants of rhodobacter capsulatus unable to grow photoautotrophically with h2 and co2 were isolated. those lacking uptake hydrogenase activity as measured by h2-dependent methylene blue reduction were analyzed genetically and used in complementation studies for the isolation of the wild-type genes. results of further subcloning and transposon tn5 mutagenesis suggest the involvement of a minimum of five genes. hybridization to the 2.2-kilobase-pair ssti fragment that lies within the coding region ...19892536678
an unusual gene cluster for the cytochrome bc1 complex in bradyrhizobium japonicum and its requirement for effective root nodule symbiosis.two adjacent genes in bradyrhizobium japonicum, fbcf and fbch, encode the rieske iron sulfur protein and cytochromes b and c1, characteristic constituents of the respiratory complex iii. remarkably, fbch is a single gene of which the 5' half codes for cytochrome b and the 3' half codes for cytochrome c1. experimental evidence suggests that a large fbch precursor is posttranslationally processed into the two proteins. b. japonicum fbcf and fbch insertion mutants grow aerobically but are unable to ...19892541921
identification of bradyrhizobium nod genes involved in host-specific nodulation.three loci important for soybean nodulation by bradyrhizobium japonicum were delimited by tn5 mutagenesis on a 5.3-kilobase ecori fragment adjacent to the nodabc genes. results of hybridization studies suggested that this region is conserved in bradyrhizobium species but absent in all rhizobium species. lacz translational fusions of two of the loci contained in this region were found to be inducible by host-produced flavonoid chemicals via a mechanism requiring a functional nodd gene product. a ...19892542223
hydrogenase synthesis in bradyrhizobium japonicum hupc mutants is altered in sensitivity to dna gyrase inhibitors.in the hupc mutants of bradyrhizobium japonicum sr, regulation of expression of hydrogenase is altered; the mutants synthesize hydrogenase constitutively in the presence of atmospheric levels of oxygen. the dna gyrase inhibitors nalidixic acid, novobiocin, and coumermycin were used to inhibit growth of wild-type and mutant cells. for each inhibitor tested, growth of mutant and wild-type strains was equally sensitive. however, in contrast to the wild type, the hupc mutants synthesized hydrogenase ...19892547335
adenylate cyclase and cyclic amp phosphodiesterase in bradyrhizobium japonicum bacteroids.adenylate cyclase and cyclic amp (camp) phosphodiesterase have been identified and partially characterized in bacteroids of bradyrhizobium japonicum 3i1b-143. adenylate cyclase activity was found in the bacteroid membrane fraction, whereas camp phosphodiesterase activity was located in both the membrane and the cytosol. in contrast to other microorganisms, b. japonicum adenylate cyclase remained firmly bound to the membrane during treatment with detergents. adenylate cyclase was activated four- ...19892548992
the nifa gene product from rhizobium leguminosarum biovar trifolii lacks the n-terminal domain found in other nifa proteins.the nifa gene has been identified between the fixx and nifb genes in the clover microsymbiont rhizobium leguminosarum biovar trifolii (r.i. bv. trifolii) strain anu843. expression of the nifa gene is induced in the symbiotic state and site-directed mutagenesis experiments indicate that nifa expression is essential for symbiotic nitrogen fixation. interestingly, the predicted r.i. bv. trifolii nifa protein lacks an n-terminal domain that is present in the homologous proteins from r.i. bv. viciae, ...19892552256
expression of the adenyl cyclase-encoding gene (cya) of rhizobium meliloti f34: existence of two cya genes?to gain insight into the role of cyclic amp (camp) in gram-negative soil bacteria, we have studied the expression of an adenyl cyclase-encoding gene 'cya' of rhizobium meliloti f34. in both escherichia coli and bradyrhizobium japonicum, the gene is expressed from a promoter(s) contained on a 2.6-kb fragment of the cloned insert, which may indicate the presence of a functional 'cya' promoter or the coincidental presence of sequences which can function as promoters in these two species. the study ...19892555267
identification of a regulatory nifa type gene and physical mapping of cloned new nif regions of azospirillum brasilense.three new tn5-mutagenized nif genes of azospirillum brasilense were characterized. the sizes of the restriction fragments and the restriction maps of the cloned nif dna regions showed that these nif genes are distinct from those reported earlier, e.g. nifhdk, nife, nifus, fixabc. the nif27 mutant was identified as a nifa type regulatory gene of a. brasilense (a) by genetic complementation with nifa of klebsiella pneumoniae, (b) by the absence of nitrogenase iron protein in western protein blots ...19892559312
rhizobium meliloti 1021 has three differentially regulated loci involved in glutamine biosynthesis, none of which is essential for symbiotic nitrogen fixation.we have cloned and characterized three distinct rhizobium meliloti loci involved in glutamine biosynthesis (glna, glnii, and glnt). the glna locus shares dna homology with the glna gene of klebsiella pneumoniae, encodes a 55,000-dalton monomer subunit of the heat-stable glutamine synthetase (gs) protein (gsi), and complemented an escherichia coli glna mutation. the glnii locus shares dna homology with the glnii gene of bradyrhizobium japonicum and encodes a 36,000-dalton monomer subunit of the h ...19892563998
isolation, characterization, and complementation of rhizobium meliloti 104a14 mutants that lack glutamine synthetase ii activity.the glutamine synthetase (gs)-glutamate synthase pathway is the primary route used by members of the family rhizobiaceae to assimilate ammonia. two forms of glutamine synthetase, gsi and gsii, are found in rhizobium and bradyrhizobium species. these are encoded by the glna and glnii genes, respectively. starting with a rhizobium meliloti glna mutant as the parent strain, we isolated mutants unable to grow on minimal medium with ammonia as the sole nitrogen source. for two auxotrophs that lacked ...19892570058
glutamine synthetase ii in rhizobium: reexamination of the proposed horizontal transfer of dna from eukaryotes to prokaryotes.we have determined the dna sequence of a rhizobium meliloti gene that encodes glutamine synthetase ii (gsii). the deduced amino acid sequence was compared to that of bradyrhizobium japonicum gsii and those of various plant and mammalian glutamine synthetases (gs) in order to evaluate a proposal that the gene for this enzyme was recently transferred from plants to their symbiotic bacteria. there is 83.6% identity between the r. meliloti and b. japonicum proteins. the bacterial gsii proteins avera ...19892575672
fast-growing root nodule bacteria produce a novel polyamine, aminobutylhomospermidine.polyamines in various root nodule bacteria including bradyrhizobium japonicum, rhizobium fredii, r. leguminosarum, r. meliloti and r. loti were identified by capillary gas chromatography. homospermidine was the polyamine present in highest concentration in all the rhizobia tested. in addition to putrescine and homospermidine, fast-growing type of rhizobial cells contained a novel polyamine, aminobutylhomospermidine, nh2(ch2)4nh(ch2)4nh(ch2)4nh2. the unusual tetraamine was not found in the cells ...19892597153
common nodabc genes in nod locus 1 of azorhizobium caulinodans: nucleotide sequence and plant-inducible expression.azorhizobium caulinodans strain ors571 induces nitrogen-fixing nodules on roots and stem-located root primordia of sesbania rostrata. two essential nod loci have been previously identified in the bacterial genome, one of which (nod locus 1) shows weak homology with the common nodc gene of rhizobium meliloti. here we present the nucleotide sequence of this region and show that it contains three contiguous open reading frames (orfa, orfb and orfc) that are related to the nodabc genes of rhizobium ...19892615763
immunological identification and distribution of dissimilatory heme cd1 and nonheme copper nitrite reductases in denitrifying bacteria.polyclonal antibodies were used to identify heme or copper nitrite reductases in the following groups: 23 taxonomically diverse denitrifiers from culture collections, 100 numerically dominant denitrifiers from geographically diverse environments, and 51 denitrifiers from a culture collection not selected for denitrification. antisera were raised against heme nitrite reductases from pseudomonas aeruginosa and pseudomonas stutzeri and against copper nitrite reductase from achromobacter cycloclaste ...19892624465
molecular cloning of a gene for indole-3-acetamide hydrolase from bradyrhizobium japonicum.a plafr1 cosmid genomic library of wild-type bradyrhizobium japonicum j1063 was constructed. a cosmid clone designated pbjj4, containing a 26-kilobase (kb) dna insert, was identified as being able to confer the ability to convert alpha-naphthaleneacetamide acid on b. japonicum j1b7 rifr, which cannot perform this conversion. the gene coding for the enzyme that converts alpha-naphthaleneacetamide to alpha-naphthaleneacetic acid was localized in the 3.5-kb region of pbjj4 by recloning in plasmid p ...19892646294
estimation of nitrogenase activity in the presence of ethylene biosynthesis by use of deuterated acetylene as a substrate.nitrogenase reduces deuterated acetylene primarily to cis dideuterated ethylene. this can be distinguished from undeuterated ethylene by the use of fourier transform infrared spectroscopy. characteristic bands in the region from 800 to 3,500 cm-1 can be used to identify and quantitate levels of these products. this technique is applicable to field studies of nitrogen fixation where ethylene biosynthesis by plants or bacteria is occurring. we have verified the reaction stoichiometry by using kleb ...19892655535
construction of chimaeric promoter regions by exchange of the upstream regulatory sequences from fdhf and nif genes.hybrid 5' regulatory regions were constructed in which the upstream activator sequence (uas) and promoter of various nif genes were exchanged with the upstream regulatory sequence (urs) of the fdhf gene from escherichia coli. they were analysed for their regulatory response under different growth conditions with the aid of fdhf'-'lacz or nif'-'lacz fusions. placement of the uas from the bradyrhizobium japonicum nifh gene in front of the spacer (dna region between urs and promoter) plus promoter ...19892664422
the azorhizobium caulinodans nitrogen-fixation regulatory gene, nifa, is controlled by the cellular nitrogen and oxygen status.the nucleotide sequence of the azorhizobium caulinodans ors571 nifa locus was determined and the deduced nifa amino acid sequence compared with that of nifa from other nitrogen-fixing species. highly conserved domains, including helix-turn-helix and atp-binding motifs, and specific conserved residues, such as a cluster of cysteines, were identified. the nifa 5' upstream region was found to contain dna sequence motifs highly homologous to promoter elements involved in nifa/ntr-mediated control an ...19892664425
identification of dna regions homologous to nitrogen fixation genes nife, nifus and fixabc in azospirillum brasilense sp7.a 30 kb dna region from azospirillum brasilense sp7, containing the nitrogenase structural genes (nifhdk), has been cloned. the presence of nif genes, in the 20 kb located next to nifhdk, was explored by tn5 mutagenesis after subcloning various restriction fragments in the broad-host-range suicide vehicle psup202. over 25 mutations due to tn5 random insertions were obtained in the 20 kb and each recombined into the genome of strain sp7. four new nif loci were identified, located at about 4, 9, 1 ...19892695597
mutational analysis of the bradyrhizobium japonicum common nod genes and further nod box-linked genomic dna regions.by insertional and deletional marker replacement mutagenesis the common nod region of bradyrhizobium japonicum was examined for the presence of additional, essential nodulation genes. an open reading frame located in the 800 bp large intergenic region between nodd1 and noda did not appear to be essential for nodulation of soybean. furthermore, a strain with a deletion of the nodi- and nodj-like genes downstream of nodc had a nod+ phenotype. a mutant with a 1.7 kb deletion immediately downstream ...19892710106
lipid a with 2,3-diamino-2,3-dideoxy-glucose in lipopolysaccharides from slow-growing members of rhizobiaceae and from "pseudomonas carboxydovorans".lipid a's from two bradyrhizobium species and from the phylogenetically closely related species "pseudomonas carboxydovorans" were found to contain 2,3-diamino-2,3-dideoxy-glucose as lipid a backbone sugar. in contrast, three representatives of the genus rhizobium, as well as the phylogenetically related species agrobacterium tumefaciens, contain solely glucosamine as lipid a backbone sugar. these findings support independent studies on the phylogenetical relatedness based on 16s rrna-data of th ...19892719525
purification and partial characterization of the rhizobium leguminosarum biovar viciae ca2+-dependent adhesin, which mediates the first step in attachment of cells of the family rhizobiaceae to plant root hair tips.the ca2+-dependent adhesin which mediates the first step in attachment of bacteria of the family rhizobiaceae to plant root hair tips was isolated from the surface of rhizobium leguminosarum biovar viciae cells; its ability to inhibit attachment of r. leguminosarum to pea root hair tips was used as a bioassay. isolated adhesin was found to be able to inhibit attachment of both carbon-limited and manganese-limited r. leguminosarum cells. a multicolumn purification procedure was developed which re ...19892738027
structural and functional analysis of nitrogenase genes from the broad-host-range rhizobium strain anu240.the genes encoding the structural components of nitrogenase, nifh, nifd and nifk, from the fast-growing, broad-host-range rhizobium strain anu240 have been identified and characterized. they are duplicated and linked in an operon nifhdk in both copies. sequence analysis of the nifh gene from each copy, together with partial sequence analysis of the nifd and nifk genes, and restriction endonuclease analysis suggested that the duplication is precise. comparison of the fe-protein sequence from stra ...19892744485
dual control of the bradyrhizobium japonicum symbiotic nitrogen fixation regulatory operon fixr nifa: analysis of cis- and trans-acting elements.aerobic expression of the fixr nifa operon in bradyrhizobium japonicum was shown to depend on a cis-acting, promoter-upstream dna sequence located between the -24/-12 promoter and position -86 relative to the transcription start site. an adenine at position -66 was essential for maximal expression. a chromosomal deletion of the upstream activator sequence (uas) led to a symbiotically defective phenotype which was typical of nifa mutants. b. japonicum crude extracts contained a protein that bound ...19892753853
cytoplasmic membrane systems involved in bacterium release into soybean nodule cells as studied with two bradyrhizobium japonicum mutant strains.two bradyrhizobium japonicum, tn5-induced, mutant strains, ml126 and ml150, were studied. both induce host cell division to form normal-sized nodules that do not fix nitrogen and whose cells have very few bacteroids (bar-). early-infection (15 days post infection) cells have much endoplasmic reticulum (er), numerous golgi bodies, and large vacuoles that are probably secondary lysosomes. later the cytoplasm of the host cells of both are dominated by hundreds of vesicles containing only finely fib ...19892759102
nucleotide sequence of a gene for indole-3-acetamide hydrolase from bradyrhizobium japonicum. 19892771653
critical spacing between two essential cysteine residues in the interdomain linker of the bradyrhizobium japonicum nifa protein.a special sequence motif in the bradyrhizobium japonicum nifa protein, consisting of two functionally essential cysteines separated by four other amino acids (cys-aa4-cys), has been proposed to be part of a potential metal-binding site [(1988) nucleic acids res. 16, 2207-2224]. using the techniques of oligonucleotide-directed mutagenesis, we report here that several of the four intervening amino acids can be replaced by others without loss of nifa function. the deletion of one amino acid to give ...19892792368
bradyrhizobium japonicum glnb, a putative nitrogen-regulatory gene, is regulated by ntrc at tandem promoters.the glnb gene from bradyrhizobium japonicum, the endosymbiont of soybeans (glycine max), was isolated and sequenced, and its expression was examined under various culture conditions and in soybean nodules. the b. japonicum glnb gene encodes a 12,237-dalton polypeptide that is highly homologous to the glnb gene products from klebsiella pneumoniae and escherichia coli. the gene is located directly upstream from glna (encoding glutamine synthetase), a linkage not observed in enteric bacteria. the g ...19892793830
sequence of nifl from klebsiella pneumoniae: mode of action and relationship to two families of regulatory proteins.we present the nucleotide sequence of k. pneumoniae nifl, which negatively regulates nif transcription in response to oxygen and fixed nitrogen. it shows partial sequence homology to the general nitrogen regulatory proteins ntrb of k. pneumoniae and bradyrhizobium parasponiae. this homology is weaker than that shown between the nifa and ntrc activator components of the nif and general nitrogen control systems. the n-terminal section of the nifl protein includes a structural duplication sharing s ...19872838726
characterization of the gene encoding glutamine synthetase i (glna) from bradyrhizobium japonicum.we have isolated the bradyrhizobium japonicum gene encoding glutamine synthetase i (glna) from a phage lambda library by using a fragment of the escherichia coli glna gene as a hybridization probe. the rhizobial glna gene has homology to the e. coli glna gene throughout the entire length of the gene and can complement an e. coli glna mutant when borne on an expression plasmid in the proper orientation to be transcribed from the e. coli lac promoter. high levels of glutamine synthetase activity c ...19852859270
involvement of glutamate in the respiratory metabolism of bradyrhizobium japonicum bacteroids.bradyrhizobium japonicum bacteroids were isolated anaerobically and supplied with 14c-labeled succinate, malate, aspartate, or glutamate for periods of up to 60 min in the presence of myoglobin to control the o2 concentration. succinate and malate were absorbed about twice as rapidly as glutamate and aspartate. conversion of substrate to co2 was most rapid for malate, followed by succinate, glutamate, and aspartate. when co2 production was expressed as a proportion of total carbon taken up, mala ...19872879829
identification and characterization of the rhizobium meliloti ntrc gene: r. meliloti has separate regulatory pathways for activation of nitrogen fixation genes in free-living and symbiotic cells.we show here that rhizobium meliloti, the nitrogen-fixing endosymbiont of alfalfa (medicago sativa), has a regulatory gene that is structurally homologous to previously characterized ntrc genes in enteric bacteria. dna sequence analysis showed that r. meliloti ntrc is homologous to previously sequenced ntrc genes from klebsiella pneumoniae and bradyrhizobium sp. (parasponia) and that an ntrb-like gene is situated directly upstream from r. meliloti ntrc. similar to its counterparts in k. pneumoni ...19872881918
acetoacetyl-coa thiolase of bradyrhizobium japonicum bacteroids: purification and properties.acetoacetyl-coa thiolase of bradyrhizobium japonicum bacteroids has been purified greater than 130-fold. the enzyme has a molecular weight of 180,000 +/- 15,000 and consists of four identical subunits of 44,000 +/- 2,000. the enzyme was specific for acetoacetyl-coa; ketodecanoyl-coa did not serve as a substrate. catalysis proceeds via a ping-pong mechanism. iodoacetamide effectively inhibited the enzyme but acetoacetyl-coa provided considerable protection against this compound. magnesium was fou ...19872883931
role of the bradyrhizobium japonicum ntrc gene product in differential regulation of the glutamine synthetase ii gene (glnii).we isolated the ntrc gene from bradyrhizobium japonicum, the endosymbiont of soybean (glycine max), and examined its role in regulating nitrogen assimilation. two independent ntrc mutants were constructed by gene replacement techniques. one mutant was unable to produce ntrc protein, while the other constitutively produced a stable, truncated ntrc protein. both ntrc mutants were unable to utilize potassium nitrate as a sole nitrogen source. in contrast to wild-type b. japonicum, the ntrc null mut ...19882903856
identification of the klebsiella pneumoniae glnb gene: nucleotide sequence of wild-type and mutant alleles.the glnb gene of klebsiella pneumoniae, which encodes the nitrogen regulation protein pii, has been cloned and sequenced. the gene encodes a 12429 dalton polypeptide and is highly homologous to the escherichia coli glnb gene. the sequences of a glnb mutation which causes glutamine auxotrophy and of a tn5 induced gln+ suppressor of this mutation were also determined. the glutamine auxotrophy was deduced to be the result of a modification of the uridylylation site of pii, and the suppression was s ...19882907369
protoporphyrinogen oxidation, a step in heme synthesis in soybean root nodules and free-living rhizobia.extracts of the crude bacteroid fraction of symbiotically grown bradyrhizobium japonicum were much more active in oxidizing protoporphyrinogen to protoporphyrin than were extracts of cells grown under free-living conditions, especially when assayed in atmospheres containing only traces of oxygen. this correlates with the higher heme content of the microaerophilic nodules. furthermore, the high level of oxidative activity in the crude bacteroid fraction was associated with an uncharacterized memb ...19892914857
isolation and characterization of the dna region encoding nodulation functions in bradyrhizobium japonicum.the dna region encoding early nodulation functions of bradyrhizobium japonicum 3i1b110 (i110) was isolated by its homology to the functionally similar region from rhizobium meliloti. isolation of a number of overlapping recombinant clones from this region allowed the construction of a restriction map of the region. the identified nodulation region of b. japonicum shows homology exclusively to those regions of r. meliloti and rhizobium leguminosarum dna known to encode early nodulation functions. ...19852999080
effects of k+ on the proton motive force of bradyrhizobium sp. strain 32h1.in previous studies, respiring bradyrhizobium sp. strain 32h1 cells grown under 0.2% o2, conditions that derepress n2 fixation, were found to have a low proton motive force of less than -121 mv, because of a low membrane potential (delta psi). in contrast, cells grown under 21% o2, which do not fix n2, had high proton motive force values of -175 mv or more, which are typical of respiring bacteria, because of high delta psi values. in the present study, we found that a delta psi of 0 mv in respir ...19863009414
genetic locus in rhizobium japonicum (fredii) affecting soybean root nodule differentiation.a genetic locus in fast-growing rhizobium japonicum (fredii) usda 191 (fix+ on several contemporary soybean cultivars) was identified by random tn5 mutagenesis as affecting the development and differentiation of root nodules. this mutant (mu042) is prototrophic and shows no apparent alterations in its surface properties. it induces aberrant nodules, arrested at the same early level of differentiation, on all its host plants. an 8.1-kilobase ecori fragment containing tn5 was cloned from mu042. in ...19863009416
two-component regulatory systems responsive to environmental stimuli share strongly conserved domains with the nitrogen assimilation regulatory genes ntrb and ntrc.we report that the ntrb and ntrc proteins of bradyrhizobium sp. [parasponia] strain rp501 share homology with other regulatory proteins. there is extensive conservation of c-terminal regions between products of rp501 ntrb; klebsiella pneumoniae ntrb; escherichia coli envz, cpxa, and phor; agrobacterium tumefaciens vira; and, to a lesser extent, e. coli chea. there is also extensive conservation of n-terminal regions between products of rp501 ntrc; k. pneumoniae ntrc; e. coli ompr, sfra, phob, ch ...19863020561
tn5-induced cytochrome mutants of bradyrhizobium japonicum: effects of the mutations on cells grown symbiotically and in culture.two bradyrhizobium japonicum cytochrome mutants were obtained by tn5 mutagenesis of strain lo and were characterized in free-living cultures and in symbiosis in soybean root nodules. one mutant strain, lo501, expressed no cytochrome aa3 in culture; it had wild-type levels of succinate oxidase activity but could not oxidize nadh or n,n,n',n'-tetramethyl-p-phenylenediamine (tmpd). the cytochrome content of lo501 root nodule bacteroids was nearly identical to that of the wild type, but the mutant e ...19873029019
genetic and structural analysis of the rhizobium meliloti fixa, fixb, fixc, and fixx genes.the fixa, fixb, fixc, and fixx genes of rhizobium meliloti 1021 constitute an operon and are required for nitrogen fixation in alfalfa nodules. dna homologous to the r. meliloti fixabc genes is present in all other rhizobium and bradyrhizobium species examined, but fixabc-homologous sequences were found in only one free-living diazotroph, azotobacter vinelandii. to determine whether the fixabcx genes share sequence homology with any of the 17 klebsiella pneumoniae nif genes, we determined the en ...19873029021
isolation and characterization of symbiotic mutants of bradyrhizobium sp. (arachis) strain nc92: mutants with host-specific defects in nodulation and nitrogen fixation.random transposon tn5 mutagenesis of bradyrhizobium sp. (arachis) strain nc92, a member of the cowpea cross-inoculation group, was carried out, and kanamycin-resistant transconjugants were tested for their symbiotic phenotype on three host plants: groundnut, siratro, and pigeonpea. two nodulation (nod- phenotype) mutants were isolated. one is unable to nodulate all three hosts and appears to contain an insertion in one of the common nodulation genes (nodabcd); the other is a host-specific nodula ...19873032910
inhibition of hydrogenase synthesis by dna gyrase inhibitors in bradyrhizobium japonicum.derepression of an uptake hydrogenase in bradyrhizobium japonicum is dependent on a microaerophilic environment. addition of dna gyrase inhibitors during depression of hydrogenase specifically prevented expression of the hydrogenase enzyme. antibodies to individual hydrogenase subunits failed to detect the protein after derepression in the presence of inhibitors, although there was no general inhibition of protein synthesis. the general pattern of proteins synthesized from 14c-labeled amino acid ...19873034865
a genetic locus essential for formate-dependent growth of bradyrhizobium japonicum.a genetic locus essential for the formate-dependent growth of bradyrhizobium japonicum was isolated by complementation of ethyl methanesulfonate-induced mutants with a cosmid gene library of b. japonicum dna. three related cosmids containing 18.7 kilobase pairs of b. japonicum dna in common were identified as being able to restore formate-dependent growth capability to mutants lacking either ribulosebisphosphate carboxylase or both ribulosebisphosphate carboxylase and phosphoribulokinase activit ...19873036781
transposon-induced symbiotic mutants of bradyrhizobium japonicum: isolation of two gene regions essential for nodulation.two strains of the soybean endosymbiont bradyrhizobium japonicum, usda 110 and 61 a101 c, were mutagenized with transposon tn5. after plant infection tests of a total of 6,926 kanamycin and streptomycin resistant transconjugants, 25 mutants were identified that are defective in nodule formation (nod-) or nitrogen fixation (fix-). seven nod- mutants were isolated from strain usda110 and from strain 61 a101 c, 4 nod- mutants and 14 fix- mutants were identified. subsequent auxotrophic tests on thes ...19873037278
conformational changes in the membrane-bound hydrogenase of bradyrhizobium japonicum. evidence that the redox state of the enzyme affects its accessibility to protease and membrane-impermeant reagents.the sensitivity of the membrane-bound hydrogenase of bradyrhizobium japonicum to inactivation by proteases and membrane-impermeant protein modification reagents was compared under hydrogen versus oxygen. in membrane vesicles, the half-life of enzyme inactivation by trypsin of the h2-reduced enzyme was approximately 10 min, whereas o2-oxidized enzyme was much less sensitive to trypsin inactivation (half-life of over 90 min). diazobenzene sulfonate (dabs) affected the enzyme activity in a manner s ...19883053719
nucleotide sequence of the genetic loci encoding subunits of bradyrhizobium japonicum uptake hydrogenase.an indispensable part of the hydrogen-recycling system in bradyrhizobium japonicum is the uptake hydrogenase, which is composed of 34.5- and 65.9-kda subunits. the gene encoding the large subunit is located on a 5.9-kilobase fragment of the h2-uptake-complementing cosmid phu52 [zuber, m., harker, a.r., sultana, m.a. & evans, h.j. (1986) proc. natl. acad. sci. usa 83, 7668-7672]. we have now determined that the structural genes for both subunits are present on this fragment. two open reading fram ...19883054886
selenium increases hydrogenase expression in autotrophically cultured bradyrhizobium japonicum and is a constituent of the purified enzyme.we have investigated the effect of added selenite on autotrophic growth and the time course of hydrogen oxidation derepression in bradyrhizobium japonicum 122des cultured in a medium purified to remove selenium compounds. in addition, hydrogenase was purified to near homogeneity and examined for the specific incorporation of se into the enzyme. the addition of se at 0.1 microm significantly increased total cell protein and hydrogenase specific activity of harvested cells. also, the addition of s ...19883056905
cloning and sequencing of the genes encoding the large and the small subunits of the h2 uptake hydrogenase (hup) of rhodobacter capsulatus.the structural genes (hup) of the h2 uptake hydrogenase of rhodobacter capsulatus were isolated from a cosmid gene library of r. capsulatus dna by hybridization of bradyrhizobium japonicum. the r. capsulatus genes were localized on a 3.5 kb hindiii fragment. the fragment, cloned onto plasmid pac76, restored hydrogenase activity and autotrophic growth of the r. capsulatus mutant jp91, deficient in hydrogenase activity (hup-). the nucleotide sequence, determined by the dideoxy chain termination me ...19883067084
nickel as a micronutrient element for plants.the detrimental effects of excessive ni on plant growth have been well known for many years. more recent evidence indicates that ni is required in small amounts for normal plant growth and development. ni is an essential component of urease in plants and microorganisms. a deficiency of ni in plants is reported to result in necrotic lesions in leaves in response to toxic accumulations of urea. urease plays an essential role in mobilization of nitrogenous compounds in plants, a process that is esp ...19883076427
activation of the bradyrhizobium japonicum nifh and nifdk operons is dependent on promoter-upstream dna sequences.previous analysis of b. japonicum nifh'- and nifd'-'lacz translational fusions showed that these promoters could be activated by the k. pneumoniae nifa plus the e. coli ntra gene products. to study the functions of the dna 5' to these promoters, plasmids carrying deletions in this region were constructed and analyzed in vivo in a heterologous system consisting of an e. coli (ntra+) background with a plasmid that constitutively expresses the k. pneumoniae nifa gene. activation of the b. japonicum ...19863086837
expression of uptake hydrogenase and hydrogen oxidation during heterotrophic growth of bradyrhizobium japonicum.strains i-110 ars, sr, usda 136, usda 137, and ak13 1c of bradyrhizobium japonicum induced hup activity when growing heterotrophically in medium with carbon substrate and nh4cl in the presence of 2% h2 and 2% o2. hup activity was induced during heterotrophic growth in the presence of carbon substrates, which were assimilated during the time of h2 oxidation. strains i-110 ars and sr grown heterotrophically or chemoautotrophically for 3 days had similar rates of h2 oxidation. similar rates of hup ...19873115959
analysis of lectin binding by bradyrhizobium japonicum strains grown on nitrocellulose filters using peroxidase-labeled lectin.a procedure was developed to assess the ability of wild-type and mutant strains of bradyrhizobium japonicum to bind soybean lectin. the lectin-binding ability of bacteria grown on nitrocellulose filters was determined using peroxidase-labeled soybean lectin. the assay produced clear differences between strains known to be unable to bind soybean lectin and those which can. the assay gave results identical to those of the fluorescein isothiocyanate-soybean lectin-binding assay of t. v. bhuvaneswar ...19873118739
characterization of the rhizobium leguminosarum genes nodlmn involved in efficient host-specific nodulation.three nodulation genes, nodl, nodm and nodn, were isolated from rhizobium leguminosarum and their dna sequences were determined. the three genes are in the same orientation as the previously described nodfe genes and the predicted molecular weights of their products are 20,105 (nodl), 65,795 (nodm) and 18,031 (nodn). analysis of gene regulation using operon fusions showed that nodl, nodm and nodn are induced in response to flavanone molecules and that this induction is nodd-dependent. in additio ...19883132583
regulation of nod gene expression in bradyrhizobium japonicum.the best inducers of nod::lacz translational fusions in bradyrhizobium japonicum are isoflavones, primarily genistein and daidzein. upstream of the nodabc genes in b. japonicum is a novel gene, nody, which is coregulated with nodabc. measurements of the activity of lacz fusions to the nodd gene of b. japonicum show that this gene is inducible by soybean seed extract and selected flavonoid chemicals. the induction of the nody abc and nodd operons appears to require a functional nodd gene, indicat ...19883146016
molybdate transport by bradyrhizobium japonicum bacteroids.bacteroid suspensions of bradyrhizobium japonicum usda 136 isolated from soybeans grown in mo-deficient conditions were able to transport molybdate at a nearly constant rate for up to 1 min. the apparent km for molybdate was 0.1 microm, and the vmax was about 5 pmol/min per mg (dry weight) of bacteroid. supplementation of bacteroid suspensions with oxidizable carbon sources did not markedly increase molybdate uptake rates. anaerobically isolated bacteroids accumulated twice as much mo in 1 h as ...19883192511
the symbiotic nitrogen fixation regulatory operon (fixrnifa) of bradyrhizobium japonicum is expressed aerobically and is subject to a novel, nifa-independent type of activation.the bradyrhizobium japonicum n2 fixation regulatory gene, nifa, was sequenced and its transcription start site determined. between the start of transcription and the nifa gene an open reading frame of 278 codons was found and named fixr. a deletion in fixr which allowed transcription into nifa resulted in a 50% reduced fix activity. the fixrnifa operon was expressed in soybean root nodules, in cultures grown anaerobically with nitrate as terminal electron acceptor, in microaerobic cultures, and ...19873313281
effect of ph on tritium exchange and hydrogen production and uptake in free-living cells and in bacteroids of bradyrhizobium japonicum.soybean nodule bacteroids and bradyrhizobium japonicum free-living cells induced for h2-uptake hydrogenase, actively catalyze the evolution of h2 in a reaction highly dependent on the ph. the optimal phs for the evolution and uptake reactions were 4.0 and 7.5-8.0, respectively. no differences were found between free-living cells and bacteroids with respect to hydrogen acceptor specificity, although absolute rates of h2 uptake were higher for free-living cells. both types of cells were able to ev ...19873322198
regulation of the fixa gene and fixbc operon in bradyrhizobium japonicum.the transcriptional start site of the bradyrhizobium japonicum fixbc operon was identified by nuclease s1 mapping. it was located approximately 700 base pairs upstream of fixb and was preceded by a promoter sequence that showed strong homology to the b. japonicum fixa promoter and thus to the general nif consensus promoter sequence. further transcript mapping experiments revealed that fixa and fixbc transcription in b. japonicum strictly depended on the presence of the regulatory gene nifa and o ...19883343218
lack of carbon substrate repression of uptake hydrogenase activity in bradyrhizobium japonicum sr.the expression of ex planta uptake hydrogenase (hup) activity in bradyrhizobium japonicum sr induced in the absence or presence of carbon substrates was compared. hup activity was influenced by ph, indicating that acidification of induction medium with low buffering capacity resulting from carbon substrate metabolism inhibited hup activity. cell suspensions in medium with adequate buffering capacity and carbon substrate were limited in o2; increasing o2 availability to cells during induction sti ...19883350794
identification of a new bradyrhizobium japonicum gene (frxa) encoding a ferredoxinlike protein.an open reading frame of 74 codons was identified downstream of the nifb gene of bradyrhizobium japonicum 110. the predicted amino acid sequence shared 63% similarity with the rhodopseudomonas palustris ferredoxin i sequence. we propose to name the gene frxa. the frxa gene was found to be cotranscribed with the nifb gene. an insertion mutation within frxa hardly affected nitrogen fixation activity.19883350797
essential and non-essential domains in the bradyrhizobium japonicum nifa protein: identification of indispensable cysteine residues potentially involved in redox reactivity and/or metal binding.the amino acid sequence of the bradyrhizobium japonicum nitrogen fixation regulatory protein nifa, as derived from the nucleotide sequence of the nifa gene, was aligned to the corresponding protein sequences from klebsiella pneumoniae, rhizobium meliloti and rhizobium leguminosarum biovar viciae. high conservation was found in the central domain and in the cooh-terminal, putative dna binding domain, whereas very little homology was present within the first 250 amino acids from the nh2-terminus. ...19883357773
quantitative assay for binding of bradyrhizobium japonicum to cultured soybean cells.incubation of bradyrhizobium japonicum with the cultured soybean cell line sb-1 resulted in the adhesion of the bacteria to the plant cells. an antiserum was raised against b. japonicum, and the 125i-labeled immunoglobulin fraction was used to quantitate the number of bacteria bound to the soybean cells. the measurement of 125i-labeled antibody binding correlated well with parallel assays by microscopic observation. using this quantitation, we have optimized the parameters of the assay in terms ...19883410819
dicarboxylic acid transport in bradyrhizobium japonicum: use of rhizobium meliloti dct gene(s) to enhance nitrogen fixation.a recombinant plasmid encoding rhizobium meliloti sequences involved in dicarboxylic acid transport (plasmid prk290:4:46) (e. bolton, b. higgisson, a. harrington, and f. o'gara, arch. microbiol. 144:142-146, 1986) was used to study the relationship between dicarboxylic acid transport and nitrogen fixation in bradyrhizobium japonicum. the expression of the dct sequences on plasmid prk290:4:46 in b. japonicum cj1 resulted in increased growth rates in media containing dicarboxylic acids as the sole ...19883422072
organization and characterization of genes essential for symbiotic nitrogen fixation from bradyrhizobium japonicum i110.a total of 96 independent tn5 insertions within a 39-kilobase-pair (kbp) segment of chromosomal dna containing the three structural genes for nitrogenase (nifh, nifd, and nifk) from bradyhizobium japonicum i110 were obtained in escherichia coli and transferred to the wild-type strain by marker exchange. individual transconjugants containing a tn5 insertion were inoculated onto glycine max cv. wilkin (soybeans) and analyzed for their effect on symbiotic nitrogen fixation. in addition to the three ...19863462181
nitrogenase promoter-lacz fusion studies of essential nitrogen fixation genes in bradyrhizobium japonicum i110.dna fragments containing either the nifd or nifh promoter and 5' structural gene sequences from bradyrhizobium japonicum i110 were fused in frame to the lacz gene. stable integration of these nif promoter-lacz fusions by homologous double reciprocal crossover into a symbiotically nonessential region of the b. japonicum chromosome provided an easy assay for the effects of potential nif regulatory mutants. the level of beta-galactosidase activity expressed from these two nif promoter-lacz fusions ...19863462182
bradyrhizobium japonicum mutants defective in nitrogen fixation and molybdenum metabolism.bradyrhizobium japonicum jh mutants deficient in molybdenum metabolism into the enzymes nitrogenase and nitrate reductase were isolated by using the vector psup1011, which carries transposon tn5 (streptomycin and kanamycin resistance). mutants in mo metabolism were obtained at a frequency of 3.6 x 10(-3) (per kan strr colony). the mutants were detected by their poor ability to grow in nitrate-containing medium without added mo. one of the mutant types required 10(5) times more molybdate than the ...19873473063
variability in molybdenum uptake activity in bradyrhizobium japonicum strains.twenty naturally occurring strains of bradyrhizobium japonicum in 11 serogroups were screened for the ability to take up mo as bacteroids from soybean root nodules. the strains varied greatly in their ability to take up mo in a 1-min period. the best strain was usda 136, which had an mo uptake activity of almost 3.0 pmol/min per mg of bacteroid (dry weight). in contrast, the poorest strain, usda 62, had an mo uptake activity of 0.35 pmol of mo per min per mg of bacteroid. there were similarities ...19873473064
bacterial heme synthesis is required for expression of the leghemoglobin holoprotein but not the apoprotein in soybean root nodules.in bradyrhizobium japonicum/soybean symbiosis, the leghemoglobin (legume hemoglobin) apoprotein is a plant product, but the origin of the heme prosthetic group is not known. b. japonicum strain lo505 is a transposon tn5-induced cytochrome-deficient mutant; it excreted the oxidized heme precursor coproporphyrin iii into the growth medium. mutant strain lo505 was specifically deficient in protoporphyrinogen oxidase (protoporphyrinogen-ix:oxygen oxidoreductase, ec 1.3.3.4) activity, and thus it cou ...19873479799
cloning and expression of bradyrhizobium japonicum uptake hydrogenase structural genes in escherichia coli.to identify the structural genes for the components of bradyrhizobium japonicum uptake hydrogenase (mr 60,000 and 30,000), we have expressed these genes in escherichia coli and shown that the products cross-react with antibodies to the respective hydrogenase subunits. we constructed subclones of overlapping dna fragments from an uptake hydrogenase-complementing cosmid, phu52 [lambert, g. r., cantrell, m. a., hanus, f. j., russell, s. a., haddad, k. r. & evans, h. j. (1985) proc. natl. acad. sci. ...19863532119
nickel uptake in bradyrhizobium japonicum.free-living bradyrhizobium japonicum grown heterotrophically with 1 microm 63ni2+ accumulated label. strain sr470, a hupc mutant, accumulated almost 10-fold more 63ni2+ on a per-cell basis than did strain sr, the wild type. nongrowing cells were also able to accumulate nickel over a 2-h period, with the hupc mutant strain sr470 again accumulating significantly more 63ni2+ than strain sr. these results suggest that this mutant is constitutive for nickel uptake as well as for hydrogenase expressio ...19873558318
a locus encoding host range is linked to the common nodulation genes of bradyrhizobium japonicum.by using cloned rhizobium meliloti, rhizobium leguminosarum, and rhizobium sp. strain mpik3030 nodulation (nod) genes as hybridization probes, homologous regions were detected in the slow-growing soybean symbiont bradyrhizobium japonicum usda 110. these regions were found to cluster within a 25-kilobase (kb) region. specific nod probes from r. meliloti were used to identify noda-, nodb-, nodc-, and nodd-like sequences clustered on two adjacent hindiii restriction fragments of 3.9 and 5.6 kb. a 7 ...19873584066
structure of the bradyrhizobium japonicum gene hema encoding 5-aminolevulinic acid synthase.the nucleotide (nt) sequence of the hema gene, which encodes 5-aminolevulinic acid synthase (alas) from the bacterium bradyrhizobium japonicum, is presented. this sequence predicts a protein of 408 amino acids (aa) with an mr of 44,599. this predicted amino acid sequence is highly homologous to that of the chicken embryonic liver alas, exhibiting a 48.8% identical amino acid sequence over the entire length of the bacterial protein. a single mrna start point was demonstrated by s1 protection anal ...19873609750
mapping and nucleotide sequence of the nifs promoter of bradyrhizobium japonicum. 19873684606
effect of lectin on nodulation by wild-type bradyrhizobium japonicum and a nodulation-defective mutant.the nodulation characteristics of wild-type bradyrhizobium japonicum usda 110 and mutant strain hs111 were examined. mutant strain hs111 exhibits a delayed-nodulation phenotype, a result of its inability to initiate successful nodulation promptly following inoculation of the soybean root. previously, we showed that the defect in initiation of infection leading to subsequent nodulation which is found in hs111 can be phenotypically reversed by pretreatment with soybean root exudate or soybean seed ...19863707122
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