Physiology, genetics, and biochemistry of carbon metabolism in the alphaproteobacterium<i>Sinorhizobium meliloti</i>

Geddes, Barney A.; Oresnik, Ivan J.

doi:10.1139/cjm-2014-0306

Cited by 51 publications

(53 citation statements)

References 162 publications

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“…(i) The ED pathway is needed for optimal growth on both glucose and succinate. The Entner-Doudoroff (ED) pathway was shown to be essential for growth on glucose, as expected (26), but surprisingly, its mutation caused growth deficiency on succinate ( Fig. 2 and Table 2).…”

supporting

confidence: 69%

“…2 and Table 2). Carbon cycling has been reported for a number of other bacterial species, including Sinorhizobium meliloti (26)(27)(28). RL0753 and RL1315 encode putative glucose-6-phosphate-1-dehydrogenases ( Fig.…”

mentioning

confidence: 98%

“…2 and Table 2). Most free-living rhizobia utilize the ED pathway for the catabolism of glucose to pyruvate (26,27), with the pathway converting glucose-6-phosphate into pyruvate in a 1:2 molar ratio and generating one molecule of ATP (27). Its requirement for growth on succinate suggests that sugars made by gluconeogenesis may, at least partly, cycle back through the ED pathway ( Fig.…”

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confidence: 99%

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Role of O ₂ in the Growth of Rhizobium leguminosarum bv. viciae 3841 on Glucose and Succinate

et al. 2017

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Insertion sequencing (INSeq) analysis of Rhizobium leguminosarum bv. viciae 3841 (Rlv3841) grown on glucose or succinate at both 21% and 1% O 2 was used to understand how O 2 concentration alters metabolism. Two transcriptional regulators were required for growth on glucose (pRL120207 [eryD] and RL0547[phoB]), five were required on succinate (pRL100388, RL1641, RL1642, RL3427, and RL4524 [ecfL]), and three were required on 1% O 2 (pRL110072, RL0545 [phoU], and RL4042). A novel toxin-antitoxin system was identified that could be important for generation of new plasmidless rhizobial strains. Rlv3841 appears to use the methylglyoxal pathway alongside the Entner-Doudoroff (ED) pathway and tricarboxylic acid (TCA) cycle for optimal growth on glucose. Surprisingly, the ED pathway was required for growth on succinate, suggesting that sugars made by gluconeogenesis must undergo recycling. Altered amino acid metabolism was specifically needed for growth on glucose, including RL2082 (gatB) and pRL120419 (opaA, encoding omegaamino acid:pyruvate transaminase). Growth on succinate specifically required enzymes of nucleobase synthesis, including ribose-phosphate pyrophosphokinase (RL3468 [prs]) and a cytosine deaminase (pRL90208 [codA]). Succinate growth was particularly dependent on cell surface factors, including the PrsD-PrsE type I secretion system and UDP-galactose production. Only RL2393 (glnB, encoding nitrogen regulatory protein PII) was specifically essential for growth on succinate at 1% O 2 , conditions similar to those experienced by N 2 -fixing bacteroids. Glutamate synthesis is constitutively activated in glnB mutants, suggesting that consumption of 2-ketoglutarate may increase flux through the TCA cycle, leading to excess reductant that cannot be reoxidized at 1% O 2 and cell death.IMPORTANCE Rhizobium leguminosarum, a soil bacterium that forms N 2 -fixing symbioses with several agriculturally important leguminous plants (including pea, vetch, and lentil), has been widely utilized as a model to study Rhizobium-legume symbioses. Insertion sequencing (INSeq) has been used to identify factors needed for its growth on different carbon sources and O 2 levels. Identification of these factors is fundamental to a better understanding of the cell physiology and core metabolism of this bacterium, which adapts to a variety of different carbon sources and O 2 tensions during growth in soil and N 2 fixation in symbiosis with legumes.KEYWORDS INSeq, nodules, Rhizobium leguminosarum, legumes, nitrogen fixation, Pisum sativum R hizobia are alphaproteobacteria able to form symbioses with legumes, on which they induce development of root nodules. Nodules provide an environment with both a low O 2 concentration and the correct nutrients to enable differentiation of rhizobia into nitrogen (N 2 )-fixing bacteroids (1). Bacteroids fix atmospheric N 2 into ammonium (NH 4 ϩ ), which is secreted to the plant host, and are, in turn, provided with

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confidence: 69%

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confidence: 98%

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confidence: 99%

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Role of O ₂ in the Growth of Rhizobium leguminosarum bv. viciae 3841 on Glucose and Succinate

et al. 2017

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“…For example, S. meliloti produces two types of EPS, succinoglycan and galactoglucan, depending on environmental conditions (Reuber and Walker 1993;Rüberg et al 1999) and R. leguminosarum produces additional types of surface polysaccharides (neutral glucomannan and gel-forming polysaccharide) (Laus et al 2006). This shows the complexity of the outer surface of rhizobial cells and suggests a complex regulatory network engaged in the synthesis of polysaccharides (Janczarek 2011;Geddes and Oresnik 2014).…”

Section: Discussionmentioning

confidence: 93%

Production of exopolysaccharide by Rhizobium leguminosarum bv. trifolii and its role in bacterial attachment and surface properties

et al. 2014

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Background and aims The acidic exopolysaccharide (EPS) produced by Rhizobium leguminosarum bv. trifolii is required for the establishment of effective symbiosis with compatible host plants (Trifolium spp.). In the rhizobium-legume interaction, early stages of root infection and nodule development have been well studied from a genetic standpoint. However, factors important for colonization of several surfaces by rhizobia, including soil particles and roots, have not yet been thoroughly investigated. The aim of this study was establishing of environmental factors affecting production of EPS by R. leguminosarum bv. trifolii strain 24.2 and the role of this polysaccharide in bacterial surface properties and attachment ability. Methods Besides the wild-type strain, its derivatives differing in the level of EPS produced were used to these analyses. The ability of attachment to abiotic and biotic surfaces of these strains were established using CFU counting experiments. Three-dimensional structure and other parameters of biofilms formed were characterized in confocal laser scanning microscopy. Electrokinetic (zeta) potential of rhizobial cells were determined using Laser Doppler Velocimetry. Results It was evidenced that the ability of R. leguminosarum bv. trifolii to produce EPS significantly affected bacterial attachment and biofilm formation on both abiotic and biotic surfaces. In addition, the presence of this polysaccharide influenced the zeta potential of rhizobial cells. Mutant strains having a mutation in genes involved in EPS synthesis were significantly impaired in attachment, whereas strains overproducing this polysaccharide showed higher adhesion efficiency to all of the tested materials. EPS facilitated attachment of bacterial cells to the tested surfaces most probably due to hydrophobic interactions and heterogeneity of the envelope surface. Conclusions EPS produced by R. leguminosarum bv. trifolii plays a significant role in attachment and biofilm formation to both abiotic and biotic surfaces as well as bacterial surface properties.

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“…The CUT1 class maltose transporter from Escherichia coli has served as a model for Gramnegative ABC-type importers (8,9,(11)(12)(13)(14). Unlike with the genetic and biochemical evidence that exists for the CUT1 class of transporters (6, 10), relatively little evidence is available for the CUT2 class of ABC transporters.The catabolism of carbon and the transport of substrates are important aspects that come into play during the interaction of bacteria with plants (15,16). In Rhizobium leguminosarum, the catabolism of rhamnose was shown to be necessary for competition for nodule occupancy (17).…”

mentioning

confidence: 99%

The Sugar Kinase That Is Necessary for the Catabolism of Rhamnose in Rhizobium leguminosarum Directly Interacts with the ABC Transporter Necessary for Rhamnose Transport

Rivers

Oresnik

2015

J Bacteriol

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Rhamnose catabolism in Rhizobium leguminosarum was found to be necessary for the ability of the organism to compete for nodule occupancy. Characterization of the locus necessary for the catabolism of rhamnose showed that the transport of rhamnose was dependent upon a carbohydrate uptake transporter 2 (CUT2) ABC transporter encoded by rhaSTPQ and on the presence of RhaK, a protein known to have sugar kinase activity. A linker-scanning mutagenesis analysis of rhaK showed that the kinase and transport activities of RhaK could be separated genetically. More specifically, two pentapeptide insertions defined by the alleles rhaK72 and rhaK73 were able to uncouple the transport and kinase activities of RhaK, such that the kinase activity was retained, but cells carrying these alleles did not have measurable rhamnose transport rates. These linker-scanning alleles were localized to the C terminus and N terminus of RhaK, respectively. Taken together, the data led to the hypothesis that RhaK might interact either directly or indirectly with the ABC transporter defined by rhaSTPQ. In this work, we show that both N-and C-terminal fragments of RhaK are capable of interacting with the N-terminal fragment of the ABC protein RhaT using a 2-hybrid system. Moreover, if RhaK fragments carrying either the rhaK72 or rhaK73 allele were used, this interaction was abolished. Phylogenetic and bioinformatic analysis of the RhaK fragments suggested that a conserved region in the N terminus of RhaK may represent a putative binding domain. Alanine-scanning mutagenesis of this region followed by 2-hybrid analysis revealed that a substitution of any of the conserved residues greatly affected the interaction between RhaT and RhaK fragments, suggesting that the sugar kinase RhaK and the ABC protein RhaT interact directly. IMPORTANCEABC transporters involved in the transport of carbohydrates help define the overall physiological fitness of bacteria. The two largest groups of transporters are the carbohydrate uptake transporter classes 1 and 2 (CUT1 and CUT2, respectively). This work provides the first evidence that a kinase that is necessary for the catabolism of a sugar can directly interact with a domain from the ABC protein that is necessary for its transport. C ells are separated from the external environment by a selectively permeable membrane. A means to transport physiologically relevant substrates across these membranes is required (1-3). The largest and most widely distributed family of transporters is the ATP binding cassette (ABC) transporters (1). ABC transporters are responsible for the transport of a very diverse set of substrates and can be broken into two main classes, importers and exporters (1, 3-5). Much of our understanding of the structure, mechanism, and kinetics of these systems has been derived from the study of relatively few model transporters (6).Gram-negative ABC importers typically consist of a permease component that is made up of two membrane-spanning proteins (encoded by either one or two genes), a periplasm...

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Physiology, genetics, and biochemistry of carbon metabolism in the alphaproteobacteriumSinorhizobium meliloti

Cited by 51 publications

References 162 publications

Role of O ₂ in the Growth of Rhizobium leguminosarum bv. viciae 3841 on Glucose and Succinate

Role of O ₂ in the Growth of Rhizobium leguminosarum bv. viciae 3841 on Glucose and Succinate

Production of exopolysaccharide by Rhizobium leguminosarum bv. trifolii and its role in bacterial attachment and surface properties

The Sugar Kinase That Is Necessary for the Catabolism of Rhamnose in Rhizobium leguminosarum Directly Interacts with the ABC Transporter Necessary for Rhamnose Transport

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Physiology, genetics, and biochemistry of carbon metabolism in the alphaproteobacteriumSinorhizobium meliloti

Cited by 51 publications

References 162 publications

Role of O 2 in the Growth of Rhizobium leguminosarum bv. viciae 3841 on Glucose and Succinate

Role of O 2 in the Growth of Rhizobium leguminosarum bv. viciae 3841 on Glucose and Succinate

Production of exopolysaccharide by Rhizobium leguminosarum bv. trifolii and its role in bacterial attachment and surface properties

The Sugar Kinase That Is Necessary for the Catabolism of Rhamnose in Rhizobium leguminosarum Directly Interacts with the ABC Transporter Necessary for Rhamnose Transport

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Role of O ₂ in the Growth of Rhizobium leguminosarum bv. viciae 3841 on Glucose and Succinate

Role of O ₂ in the Growth of Rhizobium leguminosarum bv. viciae 3841 on Glucose and Succinate