Dataset Information

RhaU of Rhizobium leguminosarum is a rhamnose mutarotase.

ABSTRACT: Of the nine genes comprising the L-rhamnose operon of Rhizobium leguminosarum, rhaU has not been assigned a function. The construction of a Delta rhaU strain revealed a growth phenotype that was slower than that of the wild-type strain, although the ultimate cell yields were equivalent. The transport of L-rhamnose into the cell and the rate of its phosphorylation were unaffected by the mutation. RhaU exhibits weak sequence similarity to the formerly hypothetical protein YiiL of Escherichia coli that has recently been characterized as an L-rhamnose mutarotase. To characterize RhaU further, a His-tagged variant of the protein was prepared and subjected to mass spectrometry analysis, confirming the subunit size and demonstrating its dimeric structure. After crystallization, the structure was refined to a 1.6-A resolution to reveal a dimer in the asymmetric unit with a very similar structure to that of YiiL. Soaking a RhaU crystal with L-rhamnose resulted in the appearance of beta-L-rhamnose in the active site.

SUBMITTER: Richardson JS

PROVIDER: S-EPMC2293258 | biostudies-literature | 2008 Apr

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

RhaU of Rhizobium leguminosarum is a rhamnose mutarotase.

Richardson Jason S JS Carpena Xavi X Switala Jack J Perez-Luque Rosa R Donald Lynda J LJ Loewen Peter C PC Oresnik Ivan J IJ

Journal of bacteriology 20071221 8

Of the nine genes comprising the L-rhamnose operon of Rhizobium leguminosarum, rhaU has not been assigned a function. The construction of a Delta rhaU strain revealed a growth phenotype that was slower than that of the wild-type strain, although the ultimate cell yields were equivalent. The transport of L-rhamnose into the cell and the rate of its phosphorylation were unaffected by the mutation. RhaU exhibits weak sequence similarity to the formerly hypothetical protein YiiL of Escherichia coli ...[more]

PMID: 18156270

Similar Datasets

Project description: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.ABC 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.

Project description:Establishment of the symbiotic relationship that develops between rhizobia and their legume hosts is contingent upon an interkingdom signal exchange. In response to host legume flavonoids, NodD proteins from compatible rhizobia activate expression of nodulation genes that produce lipochitin oligosaccharide signaling molecules known as Nod factors. Root nodule formation commences upon legume recognition of compatible Nod factor. Rhizobium leguminosarum was previously considered to contain one copy of nodD; here, we show that some strains of the Trifolium (clover) microsymbiont R. leguminosarum bv. trifolii contain a second copy designated nodD2. nodD2 genes were present in 8 out of 13 strains of R. leguminosarum bv. trifolii, but were absent from the genomes of 16 R. leguminosarum bv. viciae strains. Analysis of single and double nodD1 and nodD2 mutants in R. leguminosarum bv. trifolii strain TA1 revealed that NodD2 was functional and enhanced nodule colonization competitiveness. However, NodD1 showed significantly greater capacity to induce nod gene expression and infection thread formation. Clover species are either annual or perennial and this phenological distinction is rarely crossed by individual R. leguminosarum bv. trifolii microsymbionts for effective symbiosis. Of 13 strains with genome sequences available, 7 of the 8 effective microsymbionts of perennial hosts contained nodD2, whereas the 3 microsymbionts of annual hosts did not. We hypothesize that NodD2 inducer recognition differs from NodD1, and NodD2 functions to enhance competition and effective symbiosis, which may discriminate in favor of perennial hosts.IMPORTANCE Establishment of the rhizobium-legume symbiosis requires a highly specific and complex signal exchange between both participants. Rhizobia perceive legume flavonoid compounds through LysR-type NodD regulators. Often, rhizobia encode multiple copies of nodD, which is one determinant of host specificity. In some species of rhizobia, the presence of multiple copies of NodD extends their symbiotic host-range. Here, we identified and characterized a second copy of nodD present in some strains of the clover microsymbiont Rhizobium leguminosarum bv. trifolii. The second nodD gene contributed to the competitive ability of the strain on white clover, an important forage legume. A screen for strains containing nodD2 could be utilized as one criterion to select strains with enhanced competitive ability for use as inoculants for pasture production.

Dataset Information

RhaU of Rhizobium leguminosarum is a rhamnose mutarotase.

Publications

RhaU of Rhizobium leguminosarum is a rhamnose mutarotase.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets