Role of the aminotransferase domain in <scp><i>B</i></scp><i>acillus subtilis</i> <scp>GabR</scp>, a pyridoxal 5′‐phosphate‐dependent transcriptional regulator

Okuda, Keita; Kato, Shiro; Ito, Tomokazu; Shiraki, Shunsuke; Kawase, Yumiko; Goto, Masaru; Kawashima, Susumu; Hemmi, Hisashi; Fukada, Harumi; Yoshimura, Teizo

doi:10.1111/mmi.12861

Cited by 34 publications

(72 citation statements)

References 24 publications

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“…The DB‐(linker included) and the AAT‐like domains were separately expressed and purified as soluble proteins, suggesting that each domain is an independent folding unit . Differential scanning calorimetry, sedimentation velocity and gel filtration analyses showed that GabR is a dimer . The shape of GabR in solution, determined by small angle X‐ray scattering (SAXS), is also consistent with its dimeric crystal structure .…”

Section: The Chimeric Structure Of Mocr‐tfsmentioning

confidence: 71%

The MocR‐like transcription factors: pyridoxal 5′‐phosphate‐dependent regulators of bacterial metabolism

et al. 2018

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Many biological functions played by current proteins were not created by evolution from scratch, rather they were obtained combining already available protein scaffolds. This is the case of MocR-like bacterial transcription factors (MocR-TFs), a subclass of GntR transcription regulators, whose structure is the outcome of the fusion between DNA-binding proteins and pyridoxal 5'-phosphate (PLP)-dependent enzymes. The resultant chimeras can count on the properties of both protein classes, i.e. the capability to recognize specific DNA sequences and to bind PLP and amino-compounds; it is the modulation of such binding properties to confer to MocR-TFs chimeras the ability to interact with effector molecules and DNA so as to regulate transcription. MocR-TFs control different metabolic processes involving vitamin B and amino acids, which are canonical ligands of PLP-dependent enzymes. However, MocR-TFs are also implicated in the metabolism of compounds that are not substrates of PLP-dependent enzymes, such as rhizopine and ectoine. Genomic analyses show that MocR-TFs are widespread among eubacteria, implying an essential role in their metabolism and highlighting the scarcity of our knowledge on these important players in microbial metabolism. Although MocR-TFs have been discovered 15 years ago, the research activity on these transcriptional regulators has only recently intensified, producing a wealth of information that needs to be brought back to general principles. This is the main task of this review, which reports and analyses the available information concerning MocR-TFs functional role, structural features, interaction with effector molecules and the characteristics of DNA transcriptional factor-binding sites of MocR-based regulatory systems.

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Section: The Chimeric Structure Of Mocr‐tfsmentioning

confidence: 71%

The MocR‐like transcription factors: pyridoxal 5′‐phosphate‐dependent regulators of bacterial metabolism

et al. 2018

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“…1A). A search of the Conserved Domain Database (CDD) (http://www.ncbi.nlm.nih.gov/Structure/cdd/cdd.shtml) revealed that the predicted gene product, EhuR, belongs to the MocR/GntR subfamily, members of which have been reported to be transcriptional regulators (3,4,7,8). Similar to other MocR proteins described previously, EhuR is predicted to consist of an N-terminal helix-turn-helix DNA-binding domain and a C-terminal domain with homology to members of the aspartate aminotransferase superfamily.…”

Section: Resultsmentioning

confidence: 99%

“…Bioinformatic analysis revealed that ehuR belongs to the MocR family, which has a characteristic structure, i.e., an N-terminal HTH DNA-binding domain and a C-terminal domain with homology to members of the aspartate aminotransferase superfamily (3). These proteins require the cofactor PLP to catalyze the reversible transfer of an amino group from an amino acid to an acceptor (8,13,14). Members of the MocR family can be found in both Gram-negative and Gram-positive bacteria.…”

Section: Discussionmentioning

confidence: 99%

Negative Regulation of Ectoine Uptake and Catabolism in Sinorhizobium meliloti: Characterization of the EhuR Gene

Cai

Zhang

et al. 2017

J Bacteriol

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Ectoine has osmoprotective effects on Sinorhizobium meliloti that differ from its effects in other bacteria. Ectoine does not accumulate in S. meliloti cells; instead, it is degraded. The products of the ehuABCD-eutABCDE operon were previously discovered to be responsible for the uptake and catabolism of ectoine in S. meliloti. However, the mechanism by which ectoine is involved in the regulation of the ehuABCD-eutABCDE operon remains unclear. The ehuR gene, which is upstream of and oriented in the same direction as the ehuABCD-eutABCDE operon, encodes a member of the MocR/GntR family of transcriptional regulators. Quantitative reverse transcription-PCR and promoter-lacZ reporter fusion experiments revealed that EhuR represses transcription of the ehuABCD-eutABCDE operon, but this repression is inhibited in the presence of ectoine. Electrophoretic mobility shift assays and DNase I footprinting assays revealed that EhuR bound specifically to the DNA regions overlapping the Ϫ35 region of the ehuA promoter and the ϩ1 region of the ehuR promoter. Surface plasmon resonance assays further demonstrated direct interactions between EhuR and the two promoters, although EhuR was found to have higher affinity for the ehuA promoter than for the ehuR promoter. In vitro, DNA binding by EhuR could be directly inhibited by a degradation product of ectoine. Our work demonstrates that EhuR is an important negative transcriptional regulator involved in the regulation of ectoine uptake and catabolism and is likely regulated by one or more end products of ectoine catabolism.IMPORTANCE Sinorhizobium meliloti is an important soil bacterium that displays symbiotic interactions with legume hosts. Ectoine serves as a key osmoprotectant for S. meliloti. However, ectoine does not accumulate in the cells; rather, it is degraded. In this study, we characterized the transcriptional regulation of the operon responsible for ectoine uptake and catabolism in S. meliloti. We identified and characterized the transcription repressor EhuR, which is the first MocR/GntR family member found to be involved in the regulation of compatible solute uptake and catabolism. More importantly, we demonstrated for the first time that an ectoine catabolic end product could modulate EhuR DNA-binding activity. Therefore, this work provides new insights into the unique mechanism of ectoine-induced osmoprotection in S. meliloti.KEYWORDS Sinorhizobium meliloti, EhuR, MocR, ectoine, transcriptional regulator S inorhizobium meliloti is a soil bacterium that can be found either free-living or on the roots of leguminous plants. The ability to adapt to changes in the osmolality of the external soil environment is of vital importance for the growth and survival of soil bacteria, and ectoine serves as a key osmoprotectant for S. meliloti (1). Unlike many other bacteria, however, S. meliloti does not produce ectoine. Therefore, other soil

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“…The band at 425 nm may be attributed to the ketoenamine form of either PLP or PL covalently bound to bcPdxR as a Schiff base with a lysine residue [23]. The band at 335 nm arguably corresponds to the enolimine form of the same Schiff base, as indicated by the presence of emission bands at 400 and 500 nm when the protein was excited at 335 nm (data not shown) [24] and similarly to GabR from B. subtilis [25]. Treating the protein with hydroxylamine [26], we could obtain an apo-form of PdxR, in which the bands at 335 and 425 nm had disappeared (Fig.…”

Section: B Clausii Pdxr Activates Transcription Of the Pdxst Operonmentioning

confidence: 99%

Molecular mechanism of PdxR – a transcriptional activator involved in the regulation of vitamin B₆ biosynthesis in the probiotic bacterium Bacillus clausii

et al. 2015

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Pyridoxal 5′‐phosphate (PLP), the well‐known active form of vitamin B6, is an essential enzyme cofactor involved in a large number of metabolic processes. PLP levels need to be finely tuned in response to cell requirements; however, little is known about the regulation of PLP biosynthesis and recycling pathways. The transcriptional regulator PdxR activates transcription of the pdxST genes encoding PLP synthase. It is characterized by an N‐terminal helix‐turn‐helix motif that binds DNA and an effector‐binding C‐terminal domain homologous to PLP‐dependent enzymes. Although it is known that PLP acts as an anti‐activator, the mechanism of action of PdxR is unknown. In the present study, we analyzed the biochemical and DNA‐binding properties of PdxR from the probiotic Bacillus clausii. Spectroscopic measurements showed that PLP is the only B6 vitamer that acts as an effector molecule of PdxR. Binding of PLP to PdxR determines a protein conformational change, as detected by gel filtration chromatography and limited proteolysis experiments. We showed that two direct repeats and one inverted repeat are present in the DNA promoter region and PdxR is able to bind DNA fragments containing any combination of two of them. However, when PLP binds to PdxR, it modifies the DNA‐binding properties of the protein, making it selective for inverted repeats. A molecular mechanism is proposed in which the two different DNA binding modalities of PdxR determined by the presence or absence of PLP are responsible for the control of pdxST transcription.

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Role of the aminotransferase domain in Bacillus subtilis GabR, a pyridoxal 5′‐phosphate‐dependent transcriptional regulator

Cited by 34 publications

References 24 publications

The MocR‐like transcription factors: pyridoxal 5′‐phosphate‐dependent regulators of bacterial metabolism

The MocR‐like transcription factors: pyridoxal 5′‐phosphate‐dependent regulators of bacterial metabolism

Negative Regulation of Ectoine Uptake and Catabolism in Sinorhizobium meliloti: Characterization of the EhuR Gene

Molecular mechanism of PdxR – a transcriptional activator involved in the regulation of vitamin B₆ biosynthesis in the probiotic bacterium Bacillus clausii

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Role of the aminotransferase domain in Bacillus subtilis GabR, a pyridoxal 5′‐phosphate‐dependent transcriptional regulator

Cited by 34 publications

References 24 publications

The MocR‐like transcription factors: pyridoxal 5′‐phosphate‐dependent regulators of bacterial metabolism

The MocR‐like transcription factors: pyridoxal 5′‐phosphate‐dependent regulators of bacterial metabolism

Negative Regulation of Ectoine Uptake and Catabolism in Sinorhizobium meliloti: Characterization of the EhuR Gene

Molecular mechanism of PdxR – a transcriptional activator involved in the regulation of vitamin B6 biosynthesis in the probiotic bacterium Bacillus clausii

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Molecular mechanism of PdxR – a transcriptional activator involved in the regulation of vitamin B₆ biosynthesis in the probiotic bacterium Bacillus clausii