The mannitol utilization genes of Pseudomonas fluorescens are regulated by an activator: Cloning, nucleotide sequence and expression of the mtlR gene

Brünker, Peter; Hils, Martin; Altenbuchner, Josef; Mattes, Ralf

doi:10.1016/s0378-1119(98)00274-1

Cited by 8 publications

(8 citation statements)

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“…We found PSPTO2708 was related to sugar metabolism. In fact, PSPTO2708 has recently been characterized as a regulator of genes responsible for the internalisation of mannitol, MltR (Brunker et al 1998). Finally, PSPTO4528 is in the cluster with EutR, which is a transcriptional activator of the ethanolamine operon, suggesting its probable participation in the regulation of genes related to ethanolamine.…”

Section: Identification and Distribution Of Arac/xyls Tfsmentioning

confidence: 99%

The DNA-binding domain as a functional indicator: the case of the AraC/XylS family of transcription factors

et al. 2007

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The AraC/XylS family of transcription factors, which include proteins that are involved in the regulation of diverse biological processes, has been of considerable interest recently and has been constantly expanding by means of in silico predictions and experimental analysis. In this work, using a HMM based on the DNA binding domain of 58 experimentally characterized proteins from the AraC/XylS (A/X), 1974 A/X proteins were found in 149 out of 212 bacterial genomes. This domain was used as a template to generate a phylogenetic tree and as a tool to predict the putative regulatory role of the new members of this family based on their proximity to a particular functional cluster in the tree. Based on this approach we assigned a functional regulatory role for 75% of the TFs dataset. Of these, 33.7% regulate genes involved in carbon-source catabolism, 9.6% global metabolism, 8.3% nitrogen metabolism, 2.9% adaptation responses, 8.9% stress responses, and 11.7% virulence. The abundance of TFs involved in the regulation of metabolic processes indicates that bacteria have optimized their regulatory systems to control energy uptake. In contrast, the lower percentage of TFs required for stress, adaptation and virulence regulation reflects the specialization acquired by each subset of TFs associated with those processes. This approach would be useful in assigning regulatory roles to uncharacterized members of other transcriptional factor families and it might facilitate their experimental analysis.

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Section: Identification and Distribution Of Arac/xyls Tfsmentioning

confidence: 99%

The DNA-binding domain as a functional indicator: the case of the AraC/XylS family of transcription factors

et al. 2007

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“…For instance, in P. fluorescens DSM 50156, the gene coding for an activator protein (MtlR) belonging to the Xyl/AraC family and involved in the regulation of the mannitol catabolic pathway is not localized in the region containing all the genes involved in this catabolism (29). The genome of Z. galactanivorans contains several putative AraC-type transcriptional regulators, but none of them are located close to the mannitol utilization cluster.…”

Section: Discussionmentioning

confidence: 99%

“…Other bacteria, such as Pseudomonas fluorescens, are known to contain a mannitol-2-dehydrogenase, an enzyme that oxidizes mannitol into fructose; mannitol is transported by ATP binding cassette (ABC) transporters (28). In this organism, fructose is believed to be phosphory-lated into fructose-6-phosphate by a kinase coded for by mtlZ, a gene of the mannitol catabolic operon (28,29). In the same way, Phaeobacter inhibens DSM 17395 imports mannitol via a specific ABC transporter whose corresponding genes are located next to the mtlK gene, coding for the mannitol-2-dehydrogenase; furthermore, the frk gene, encoding a fructokinase, colocalizes with genes corresponding to another ABC transporter (30).…”

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confidence: 99%

The Mannitol Utilization System of the Marine Bacterium Zobellia galactanivorans

Groisillier

Labourel

Michel

et al. 2015

Appl Environ Microbiol

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Mannitol is a polyol that occurs in a wide range of living organisms, where it fulfills different physiological roles. In particular, mannitol can account for as much as 20 to 30% of the dry weight of brown algae and is likely to be an important source of carbon for marine heterotrophic bacteria. Zobellia galactanivorans (Flavobacteriia) is a model for the study of pathways involved in the degradation of seaweed carbohydrates. Annotation of its genome revealed the presence of genes potentially involved in mannitol catabolism, and we describe here the biochemical characterization of a recombinant mannitol-2-dehydrogenase (M2DH) and a fructokinase (FK). Among the observations, the M2DH of Z. galactanivorans was active as a monomer, did not require metal ions for catalysis, and featured a narrow substrate specificity. The FK characterized was active on fructose and mannose in the presence of a monocation, preferentially K ؉ . Furthermore, the genes coding for these two proteins were adjacent in the genome and were located directly downstream of three loci likely to encode an ATP binding cassette (ABC) transporter complex, suggesting organization into an operon. Gene expression analysis supported this hypothesis and showed the induction of these five genes after culture of Z. galactanivorans in the presence of mannitol as the sole source of carbon. This operon for mannitol catabolism was identified in only 6 genomes of Flavobacteriaceae among the 76 publicly available at the time of the analysis. It is not conserved in all Bacteroidetes; some species contain a predicted mannitol permease instead of a putative ABC transporter complex upstream of M2DH and FK ortholog genes. Brown algae (Phaeophyceae) are the dominant macroalgae in temperate and polar regions and thus play a crucial role in the primary production of coastal ecosystems (1). They contain large amounts of different structural and storage carbohydrates. For instance, their extracellular matrices are formed by the accumulation of cellulose, fucanes, and alginates (2-4), while they store carbon by accumulating laminarin and mannitol (5). The potential of this biomass resource for the production of liquid biofuels (6), including ethanol from alginate and mannitol (7,8), and for the implementation of biorefineries (9) has been highlighted recently.Depending on the species, mannitol can represent as much as 20 to 30% of the dry weight of brown seaweed (10). In the genomic and genetic model of brown algae Ectocarpus sp., formerly included in the species Ectocarpus siliculosus (11), it has been observed that the content of this polyol differs according to the diurnal cycle (12) and that it is likely to act as an osmoprotectant or a local compatible osmolyte (13). Mannitol is localized in the cytosol and is also present at the reducing ends of vacuolar laminarin molecules of the M series (in contrast to the G series, which contain only glucose residues) (14). Mannitol in brown algae is produced directly from the photoassimilate fructose-6-phosphate (F6P) by two st...

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“…As for many members of the AraC family, previous attempts to purify the complete XylS protein were unsuccessful and produced protein aggregates (3,7,49,61). To investigate the basis for this insolubility, we analyzed the two protein domains independently.…”

Section: Xyls-n-terminal Domain Is Responsible Of Xyls Insolubilitymentioning

confidence: 99%

Sequential XylS-CTD Binding to the Pm Promoter Induces DNA Bending Prior to Activation

2010

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XylS protein, a member of the AraC family of transcriptional regulators, comprises a C-terminal domain (CTD) involved in DNA binding and an N-terminal domain required for effector binding and protein dimerization. In the absence of benzoate effectors, the N-terminal domain behaves as an intramolecular repressor of the DNA binding domain. To date, the poor solubility properties of the full-length protein have restricted XylS analysis to genetic approaches in vivo. To characterize the molecular consequences of XylS binding to its operator, we used a recombinant XylS-CTD variant devoid of the N-terminal domain. The resulting protein was soluble and monomeric in solution and activated transcription from its cognate promoter in an effectorindependent manner. XylS binding sites in the Pm promoter present an intrinsic curvature of 35°centered at position ؊42 within the proximal site. Gel retardation and DNase footprint analysis showed XylS-CTD binding to Pm occurred sequentially: first a XylS-CTD monomer binds to the proximal site overlapping the RNA polymerase binding sequence to form complex I. This first event increased Pm bending to 50°and was followed by the binding of the second monomer, which further increased the observed global curvature to 98°. This generated a concomitant shift in the bending center to a region centered at position ؊51 when the two sites were occupied (complex II). We propose a model in which DNA structure and binding sequences strongly influence XylS binding events previous to transcription activation.

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The mannitol utilization genes of Pseudomonas fluorescens are regulated by an activator: Cloning, nucleotide sequence and expression of the mtlR gene

Cited by 8 publications

References 32 publications

The DNA-binding domain as a functional indicator: the case of the AraC/XylS family of transcription factors

The DNA-binding domain as a functional indicator: the case of the AraC/XylS family of transcription factors

The Mannitol Utilization System of the Marine Bacterium Zobellia galactanivorans

Sequential XylS-CTD Binding to the Pm Promoter Induces DNA Bending Prior to Activation

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