PLP and GABA trigger GabR-mediated transcription regulation in
            <i>Bacillus subtilis</i>
            via external aldimine formation

Wu, Rui; Sanishvili, Ruslan; Belitsky, Boris R.; Juncosa, Jose I.; Le, Hoang V.; Lehrer, Helaina J.; Farley, Michael; Silverman, Richard B.; Petsko, Gregory A.; Ringe, Dagmar; Liu, Dali

doi:10.1073/pnas.1703019114

Cited by 25 publications

(50 citation statements)

References 40 publications

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“…PLP is initially bound in this sequence of chemical reactions by the side‐chain of Lys‐302 present in the carboxy‐terminal aminotransferase domain of EnuR (Schulz et al ., ). This scheme was adapted from data reported for the MocR/GabR‐type regulators GabR and DdlR that respond to the availability of GABA and a di‐peptide (D‐alanyl‐D‐alanine), respectively (Edayathumangalam et al ., ; Okuda et al ., ; Takenaka et al ., ; Park et al ., ; Wu et al ., ).…”

Section: Resultsmentioning

confidence: 97%

“…Instead, a partial aminotransferase reaction occurs that initially involves the covalent binding of a system‐specific low molecular mass effector molecule to the protein‐bound PLP co‐factor (the internal aldimine) and the subsequent formation of an external aldimine between PLP and the effector molecule. These chemical reactions are transduced into a conformational change of the entire regulatory protein, which then dictates the DNA‐binding and functional properties of MocR/GabR‐type proteins to function either as activators or repressors (or both) of gene transcription (Belitsky and Sonenshein, ; Wiethaus et al ., ; Edayathumangalam et al ., ; Okuda et al ., ; Takenaka et al ., ; Park et al ., ; Tramonti et al ., ; Wu et al ., ). EnuR possesses such a covalently bound PLP molecule, and Lys‐302 present in its carboxy‐terminal aminotransferase domain has been identified through in silico modelling and mutant studies as the residue to which the co‐factor is attached (Schulz et al ., ).…”

Section: Introductionmentioning

confidence: 97%

“…This latter domain is structurally related to aminotransferases of type-I fold (Edayathumangalam et al, 2013;Milano et al, 2015;Suvorova and Rodionov, 2016). It frequently contains a covalently bound pyridoxal-5'-phosphate (PLP) co-factor (attached to a Lys-residue) but MocR/GabR-type regulators do not perform a full aminotransferase enzyme reaction (Edayathumangalam et al, 2013;Okuda et al, 2015a,b;Takenaka et al, 2015;Park et al, 2017;Wu et al, 2017). Instead, a partial aminotransferase reaction occurs that initially involves the covalent binding of a system-specific low molecular mass effector molecule to the protein-bound PLP co-factor (the internal aldimine) and the subsequent formation of an external aldimine between PLP and the effector molecule.…”

Section: Introductionmentioning

confidence: 99%

“…All enzymes involved in ectoine and hydroxyectoine biosynthesis have been enzymatically studied (Ono et al, 1999;Bursy et al, 2007;St€ oveken et al, 2011), while EutD (DeoA) is the only enzyme from the catabolic route whose function has been experimentally assessed. The information on the generation of both N-(a)-ADABA and N-(g)-ADABA by the recombinant EutD (DoeA) enzyme from H. elongata is based on data reported by Schwibbert et al (Schwibbert et al, 2011). regulatory protein, which then dictates the DNA-binding and functional properties of MocR/GabR-type proteins to function either as activators or repressors (or both) of gene transcription (Belitsky and Sonenshein, 2002;Wiethaus et al, 2008;Edayathumangalam et al, 2013;Okuda et al, 2015a,b;Takenaka et al, 2015;Park et al, 2017;Tramonti et al, 2017;Wu et al, 2017). EnuR possesses such a covalently bound PLP molecule, and Lys-302 present in its carboxy-terminal aminotransferase domain has been identified through in silico modelling and mutant studies as the residue to which the co-factor is attached (Schulz et al, 2017).…”

Section: Introductionmentioning

confidence: 99%

“…Although an exogenous supply of ectoines triggers a strong induction of the transcription of the hydroxyectoine/ ectoine uptake and catabolic genes in R. pomeroyi DSS-3 (Lecher et al, 2009;Schulz et al, 2017), they cannot be the true inducer molecules because the chemistry underlying the interaction of the system-specific effector molecule with the Lys-bound PLP co-factor requires a primary amino group (Bramucci et al, 2011;Edayathumangalam et al, 2013;Okuda et al, 2015a,b;Takenaka et al, 2015;Suvorova and Rodionov, 2016;Park et al, 2017;Wu et al, 2017). Since free amino groups are only generated after the first step of ectoine degradation (Fig.…”

Section: Introductionmentioning

confidence: 99%

See 4 more Smart Citations

Transcriptional regulation of ectoine catabolism in response to multiple metabolic and environmental cues

Schulz

Hermann

Freibert

et al. 2017

Environmental Microbiology

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Ectoine and hydroxyectoine are effective microbial osmostress protectants, but can also serve as versatile nutrients for bacteria. We have studied the genetic regulation of ectoine and hydroxyectoine import and catabolism in the marine Roseobacter species Ruegeria pomeroyi and identified three transcriptional regulators involved in these processes: the GabR/MocR-type repressor EnuR, the feast and famine-type regulator AsnC and the two-component system NtrYX. The corresponding genes are widely associated with ectoine and hydroxyectoine uptake and catabolic gene clusters (enuR, asnC), and with microorganisms predicted to consume ectoines (ntrYX). EnuR contains a covalently bound pyridoxal-5'-phosphate as a co-factor and the chemistry underlying the functioning of MocR/GabR-type regulators typically requires a system-specific low molecular mass effector molecule. Through ligand binding studies with purified EnuR, we identified N-(alpha)-L-acetyl-2,4-diaminobutyric acid and L-2,4-diaminobutyric acid as inducers for EnuR that are generated through ectoine catabolism. AsnC/Lrp-type proteins can wrap DNA into nucleosome-like structures, and we found that the asnC gene was essential for use of ectoines as nutrients. Furthermore, we discovered through transposon mutagenesis that the NtrYX two-component system is required for their catabolism. Database searches suggest that our findings have important ramifications for an understanding of the molecular biology of most microbial consumers of ectoines.

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Section: Resultsmentioning

confidence: 97%

Section: Introductionmentioning

confidence: 97%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

Transcriptional regulation of ectoine catabolism in response to multiple metabolic and environmental cues

Schulz

Hermann

Freibert

et al. 2017

Environmental Microbiology

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(S)‐4‐Amino‐5‐phenoxypentanoate designed as a potential selective agonist of the bacterial transcription factor GabR

et al. 2020

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Addressing molecular recognition in the context of evolution requires pursuing new molecular targets to enable the development of agonists or antagonists with new mechanisms of action. Disruption of transcriptional regulation through targeting transcription factors that regulate the expression of key enzymes in bacterial metabolism may provide a promising method for controlling the bacterial metabolic pathways. To this end, we have selectively targeted a bacterial transcription regulator through the design and synthesis of a series of γ‐aminobutyric acid (GABA) derivatives, including (S)‐4‐amino‐5‐phenoxypentanoate (4‐phenoxymethyl‐GABA), which are based on docking insights gained from a previously‐solved crystal structure of GabR from Bacillus subtilis. This target was selected because GabR strictly controls GABA metabolism by regulating the transcription of the gabT/D operon. These GabR transcription modulators are selective for the bacterial transcription factor GabR and are unable to bind to structural homologs of GabR due to distinct steric constraints. We have obtained a crystal structure of 4‐phenoxymethyl‐GABA bound as an external aldimine with PLP in the effector binding site of GabR, which suggests that this compound is capable of binding and reacting in the same manner as the native effector ligand. Inhibition assays demonstrate high selectivity of 4‐phenoxymethyl‐GABA for bacterial GabR versus several selected eukaryotic enzymes. Single‐molecule fluorescence resonance energy transfer (smFRET) experiments reveal a ligand‐induced DNA distortion that is very similar to that of the native effector GABA, suggesting that the compound functions as a potential selective agonist of GabR.

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Design, Docking Simulations, Synthesis, and in vitro and in vivo Behavioral Assessment of m‐Aminobenzoic Acid Analogues as GABA‐AT Inhibitors

et al. 2021

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γ-Aminobutyric acid (GABA) is an inhibitory neurotransmitter whose deficiency is related to affections involving overexcited neurons (e. g., anxiety and epilepsy); rise of GABA levels by inhibition of GABA-aminotransferase (GABA-AT) is a treatment option. The aim of this contribution was to assess a series of maminobenzoic acid derivatives (analogous to GABA) as GABA-AT inhibitors. Chosen compounds from docking simulations, were synthesized in good yields by green chemistry, tested in vitro for inhibition of GABA-AT and assayed for protection against pentylenetetrazole (PTZ) induced seizures, and ability to counter picrotoxin (PTX) induced anxiety. These compounds had a lower binding energy (higher affinity) than GABA and GABA-AT inhibitor vigabatrin (VGB) at the model active site. DABA_2 a (3,5-dimaleamilbenzoic acid) had excellent attributes in silico (ΔG = À 9.00 kcal/mol vs À 5.33 kcal/mol of VGB), good competitive inhibition of GABA-AT in vitro (K I = 3.6 × 10 À 6 M, ΔG = À 7.42 kcal/mol) and a low acute toxicity (� 1,000 mg/kg) with good protection against PTZ and PTX (100 mg/kg), leading to stablish the good modulation of GABAergic transmission in vivo.

show abstract

PLP and GABA trigger GabR-mediated transcription regulation in Bacillus subtilis via external aldimine formation

Cited by 25 publications

References 40 publications

Transcriptional regulation of ectoine catabolism in response to multiple metabolic and environmental cues

Transcriptional regulation of ectoine catabolism in response to multiple metabolic and environmental cues

(S)‐4‐Amino‐5‐phenoxypentanoate designed as a potential selective agonist of the bacterial transcription factor GabR

Design, Docking Simulations, Synthesis, and in vitro and in vivo Behavioral Assessment of m‐Aminobenzoic Acid Analogues as GABA‐AT Inhibitors

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