Purification and characterization of homodimeric methylmalonyl-CoA mutase from Sinorhizobium meliloti
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Sinorhizobium meliloti
Sinorhizobium
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Sinorhizobium meliloti is a soil bacterium able to induce the formation of nodules on the root of specific legumes, including alfalfa (Medicago sativa). Bacteria colonize nodules through infection threads, invade the plant intracellularly, and ultimately differentiate into bacteroids capable of reducing atmospheric nitrogen to ammonia, which is directly assimilated by the plant. As a first step to describe global changes in gene expression of S. meliloti during the symbiotic process, we used whole genome microarrays to establish the transcriptome profile of bacteria from nodules induced by a bacterial mutant blocked at the infection stage and from wild-type nodules harvested at various timepoints after inoculation. Comparison of these profiles to those of cultured bacteria grown either to log or stationary phase as well as examination of a number of genes with known symbiotic transcription patterns allowed us to correlate global gene-expression patterns to three known steps of symbiotic bacteria bacteroid differentiation, i.e., invading bacteria inside infection threads, young differentiating bacteroids, and fully differentiated, nitrogen-fixing bacteroids. Finally, analysis of individual gene transcription profiles revealed a number of new potential symbiotic genes.
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Homologues to Sinorhizobium meliloti insertion sequences ISRm3 and ISRm4 were found within Rhizobium leguminosarum bv. viciae field populations. Similarly, homologous sequences to S. meliloti ISRm4 were found in S. fredii strains. Based on the polymerase chain reaction, we cloned and partially sequenced some of these putative insertion sequence elements. DNA sequence comparisons indicate that S. meliloti and R. leguminosarum ISRm3-type elements are closely related whereas the ISRm4 homologues show distinct relationships. In addition, specific primers for PCR recognition of ISRm3 and ISRm4 elements in rhizobia species and S. meliloti ISRm6 were designed.
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Rhizobium leguminosarum
Insertion sequence
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Nitrogen-fixing symbiotic bacteria known as rhizobia can exist in different soils and adapt to different environmental conditions. The aim of this study was to determine the impact of pH on the growth of Rhizobium trifolii and Sinorhizobium meliloti. Rhizobial species were isolated using yeast extract mannitol agar medium) in which the pH values were adjusted to 5.0, 6.0, 8.0 and 9.0 by adding HCl and NaOH. The optimum pH for rhizobia is neutral or slightly alkaline (pH 8) and they are more sensitive to acidity. Sinorhizobium meliloti developed better in an acid medium compared to Rhizobium trifolii.
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The symbiotic interaction between Medicago sativa and Sinorhizobium meliloti RmkatB(++) overexpressing the housekeeping catalase katB is delayed, and this delay is combined with an enlargement of infection threads. This result provides evidence that H(2)O(2) is required for optimal progression of infection threads through the root hairs and plant cell layers.
Sinorhizobium meliloti
Medicago sativa
Medicago
Sinorhizobium
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ABSTRACT To improve symbiotic nitrogen fixation on alfalfa plants, Sinorhizobium meliloti strains containing different average copy numbers of a symbiotic DNA region were constructed by specific DNA amplification (SDA). A DNA fragment containing a regulatory gene ( nodD1 ), the common nodulation genes ( nodABC ), and an operon essential for nitrogen fixation ( nifN ) from the nod regulon region of the symbiotic plasmid pSyma of S. meliloti was cloned into a plasmid unable to replicate in this organism. The plasmid then was integrated into the homologous DNA region of S. meliloti strains 41 and 1021, which resulted in a duplication of the symbiotic region. Sinorhizobium derivatives carrying further amplification were selected by growing the bacteria in increased concentrations of an antibiotic marker present in the integrated vector. Derivatives of strain 41 containing averages of 3 and 6 copies and a derivative of strain 1021 containing an average of 2.5 copies of the symbiotic region were obtained. In addition, the same region was introduced into both strains as a multicopy plasmid, yielding derivatives with an average of seven copies per cell. Nodulation, nitrogenase activity, plant nitrogen content, and plant growth were analyzed in alfalfa plants inoculated with the different strains. The copy number of the symbiotic region was critical in determining the plant phenotype. In the case of the strains with a moderate increase in copy number, symbiotic properties were improved significantly. The inoculation of alfalfa with these strains resulted in an enhancement of plant growth.
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Sinorhizobium meliloti
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In Sinorhizobium meliloti the NolR repressor displays differential negative regulation of nodulation genes and is required for optimal nodulation. Here, we demonstrate that the NolR function is not unique to S. meliloti but is also present in other species of the Rhizobiaceae family. DNA hybridization indicates the presence of nolR homologous sequences in species belonging to the Rhizobium and Sinorhizobium genera while no hybridization signal was detected in species from the Mesorhizobium, Bradyrhizo-bium, Azorhizobium, and Agrobacterium genera. We isolated the nolR gene from the Rhizobium leguminosarum bv. viciae strain TOM and showed that the TOM nolR gene acts similarly to S. meliloti nolR by repressing the expression of both the nodABCIJ and the nodD genes, resulting in decreased Nod factor production. The presence of a functional nolR gene in R. leguminosarum is correlated with an increased rate and extent of nodulation of pea. The conserved primary structure, the location of the DNA-binding domain, and the similar size of NolR proteins, compared with a family of small bacterial regulatory proteins including HlyU, SmtB, and the ArsR-type regulators, revealed that NolR belongs to this family.
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Sinorhizobium
Rhizobium leguminosarum
Nod factor
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ABSTRACT Rhizobium bacteria synthesize signal molecules called Nod factors that elicit responses in the legume root during nodulation. Nod factors, modified N-acylated β-(1,4)- N -acetylglucosamine, are synthesized by the nodulation ( nod ) gene products. We tested the ability of three Sinorhizobium meliloti nod gene products to modify Nod factor analogs with thio linkages instead of O-glycosidic bonds in the oligosaccharide backbone.
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Nod factor
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Glycosidic bond
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Sinorhizobium meliloti
Sinorhizobium
Medicago
Medicago sativa
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ABSTRACTSinorhizobium meliloti is an α-proteobacteria of the family Rhizobiaceae that alternates between a free-living phase in soil and a symbiotic phase within the host plant cells, where the bacteria ultimately differentiate into nitrogen-fixing organelle-like cells, called bacteroids. The present study was designed to compare the difference in responses to nitrogen and carbon limitation in free living Sinorhizobium meliloti strains and their ability to form symbiotic association with alfalfa (Medicago sativa) plants. The effectiveness of observed symbiotic associations was evaluated by number of formed nodules, nitrogen fixing activity, and plant biomass production in control environmental conditions of a growth chamber. A wild type strain Sinorhizobium meliloti 1021 and two mutants—Sinorhizobium meliloti NitR and Sinorhizobium meliloti TspO were used in this research. Both mutant strains were previously created to study the general stress response in Sinorhizobium meliloti and its regulation mechanisms. The products of the genes named tspO and nitR, act either as direct or indirect regulators of gene expression in response to various stresses, including starvation. The up- and down regulated genes under conditions of nitrogen and carbon limitation were identified in free living forms of the three strains. Comparison of the genes differentially expressed in the wild type strain to those found in the tspO and nitR mutants showed no significant difference. In both starvation conditions the most effective symbiotic system was established between alfalfa and Sinorhizobium meliloti TspO, concerning nitrogen fixing capacity and plant biomass production.Keywords: nitrogen fixationstarvationstrain effect
Sinorhizobium meliloti
Sinorhizobium
Medicago sativa
Medicago
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