• Korean Journal of Microbiology
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• v.38 no.4
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• pp.260-266
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• 2002

In this study, chromosomal DNA fragment related to the regulation of phenol metabolism in Ralstonia eutropha JMP 134 was cloned and sequenced. The result has shown that two open reading frames (ORF1 and ORF2) exist on this regulatory region. ORF1, which initiates from 454 bp downstream of the stop codon of the phenol hydroxylase genes, was found to be composed of 501 amino acids. ORF2, whose start codon is overlapped with the stop codon of ORFl, was found to contain 232 amino acids. The comparison of amino acid sequences with other proteins has revealed that ORF1 belongs to the family of NtrC transcriptional activator, whereas ORF2 shares high homology with the family of GntR protein, which is known to be a negative regulator. ORF1 and ORF2 were designated as a putative positive regulator, phlR2 and a negative regulator phlA, respectively. Possible regulatory mechanisms of phenol metabolism in this strain was discussed.

• BMB Reports
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• v.34 no.4
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• pp.305-309
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• 2001

MatR in Rhizobium trifolii is a malonate-responsive transcription factor that regulates the expression of genes, matABC, enabling decarboxylation of malonyl-CoA into acetyl-CoA, synthesis of malonyl-CoA from malonate and CoA, and malonate transport. According to an analysis of the amino acid sequence homology, MatR belongs to the GntR family The proteins of this family have two-domain folds, the N-terminal helix-turn-helix DNA-binding domain and the C-terminal ligand-binding domain. In order to End the malonate binding site and amino acid residues that interact with RNA polymerase, a site-directed mutagenesis was performed. Analysis of the mutant MatR suggests that Arg-160 might be involved in malonate binding, whereas Arg-102 and Arg-174 are critical for the repression activity by interacting with RNA polymerase.

• Proceedings of the Microbiological Society of Korea Conference
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• 2002.10a
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• pp.27-30
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• 2002

Ralstonia eutrupha JMP134 is a well-known soil bacterium which can metabolite diverse aromatic compounds and xenobiotics, such as phenol, 2,4-dichlorophenoxy acetic acid (2, 4-D), and trichloroethylene (TCE), etc. Phenol is degraded through chromosomally encoded phenol degradation pathway. Phenol is first metabolized into catechol by a multicomponent phenol hydroxylase, which is further metabolized to TCA cycle intermediates via a meta-cleavage pathway. The nucleotide sequences of the genes for the phenol hydroxylase have previously been determined, and found to composed of eight genes phlKLMNOPRX in an operon structure. The phlR, whose gene product is a NtrC-like transcriptional activator, was found to be located at the internal region of the structural genes, which is not the case in most bacteria where the regulatory genes lie near the structural genes. In addition to this regulatory gene, we found other regulatory genes, the phlA and phlR2, downstream of the phlX. These genes were found to be overlapped and hence likely to be co-transcribed. The protein similarity analysis has revealed that the PhlA belongs to the GntR family, which are known to be negative regulators, whereas the PhlR2 shares high homology with the NtrC-type family of transcriptional activators like the PhlR. Disruption of the phlA by insertional mutation has led to the constitutive expression of the activity of phenol hydroxylase in JMP134, indicating that PhlA is a negative regulator. Possible regulatory mechanisms of phenol metabolism in R. eutropha JMP134 has been discussed.

• Journal of Microbiology and Biotechnology
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• v.33 no.10
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• pp.1370-1375
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• 2023

In this study, we aimed to enhance the accumulation of chorismate (CHR) and anthranilate (ANT), key intermediates in the shikimate pathway, by modifying a shikimate over-producing recombinant strain of Corynebacterium glutamicum [19]. To achieve this, we utilized a CRISPR-driven genome engineering approach to compensate for the deletion of shikimate kinase (AroK) as well as ANT synthases (TrpEG) and ANT phosphoribosyltransferase (TrpD). In addition, we inhibited the CHR metabolic pathway to induce CHR accumulation. Further, to optimize the shikimate pathway, we overexpressed feedback inhibition-resistant Escherichia coli AroG and AroH genes, as well as C. glutamicum AroF and AroB genes. We also overexpressed QsuC and substituted shikimate dehydrogenase (AroE). In parallel, we optimized the carbon metabolism pathway by deleting the gntR family transcriptional regulator (IolR) and overexpressing polyphosphate/ATP-dependent glucokinase (PpgK) and glucose kinase (Glk). Moreover, acetate kinase (Ack) and phosphotransacetylase (Pta) were eliminated. Through our CRISPR-driven genome re-design approach, we successfully generated C. glutamicum cell factories capable of producing up to 0.48 g/l and 0.9 g/l of CHR and ANT in 1.3 ml miniature culture systems, respectively. These findings highlight the efficacy of our rational cell factory design strategy in C. glutamicum, which provides a robust platform technology for developing high-producing strains that synthesize valuable aromatic compounds, particularly those derived from the shikimate pathway metabolites.

• Journal of Microbiology and Biotechnology
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• v.19 no.8
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• pp.765-773
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• 2009

Edwardsiella tarda is a pathogen with a broad host range that includes human and animals. The E. tarda hemolysin (Eth) system, which comprises EthA and EthB, is a noted virulence element that is widely distributed in pathogenic isolates of E. tarda. Previous study has shown that the expression of ethB is regulated by iron, which suggests the possibility that the ferric uptake regulator (Fur) is involved in the regulation of ethB. The work presented in this report supports the previous findings and demonstrates that ethB expression was decreased under conditions when the E. tarda Fur ($Fur_{Et}$) was overproduced, and enhanced when $Fur_{Et}$ was inactivated. We also identified a second ethB regulator, EthR, which is a transcription regulator of the GntR family. EthR represses ethB expression by direct interaction with the ethB promoter region. In addition to ethB, EthR also modulates, but positively, luxS expression and AI-2 production by binding to the luxS promoter region. The expression of ethR itself is subject to negative autoregulation; interference with this regulation by overexpressing ethR during the process of infection caused (i) drastic changes in ethB and luxS expressions, (ii) vitiation in the tissue dissemination and survival ability of the bacterium, and (iii) significant attenuation of the overall bacterial virulence. These results not only provide new insights into the regulation mechanisms of the Eth hemolysin and LuxS/AI-2 quorum sensing systems but also highlight the importance of these systems in bacterial virulence.

• Proceedings of the Korean Society of Crop Science Conference
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• 2017.06a
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• pp.131-131
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• 2017

Though the Platycodon grandiflorum, has a broad range of pharmacologic properties, but the mechanisms underlying these effects remain unclear. In order to profile proteins from the nodal segment, callus, root and shoot, high throughput proteome approach was executed in the present study. Two-dimensional gels stained with CBB, a total of 84 differential expressed proteins were confirmed out of 839 protein spots using image analysis by Progenesis SameSpot software. Out of total differential expressed spots, 58 differential expressed protein spots (${\geq}2-fold$) were analyzed using MASCOT search engine according to the similarity of sequences with previously characterized proteins along with the UniProt database. Out of 58 differential expressed protein, 32 protein spots were up-regulated such as ribulose-1,5-bisphosphate carboxylase, endoplasmic oxidoreductin-1, heat stress transcription factor A3, RNA pseudourine synthase 4, cysteine proteinase, GntR family transcriptional regulator, E3 xyloglucan 6-xylosyltransferase, while 26 differential protein spots were down-regulated such as L-ascorbate oxidase precursor, late embryogenesis abundant protein D-34, putative SCO1 protein, oxygen-evolving enhancer protein 3. However, the frequency distribution of identified proteins using iProClass databases, and assignment by function based on gene ontology revealed that the identified proteins from the explants were mainly associated with the nucleic acid binding (17%), transferase activity (14%) and ion binding (12%). Taken together, the protein profile may provide insight clues for better understanding the characteristics of proteins and its metabolic activities in various explants of this essential medicinal plant P. grandiflorum.

• KOREAN JOURNAL OF CROP SCIENCE
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• v.60 no.1
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• pp.97-106
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• 2015

Platycodon grandiflorum, commonly known as Doraji in Korea, has a wide range of pharmacologic properties, such as reducing adiposity and hyperlipidemia, and antiatherosclerotic effects. However, the mechanisms underlying these effects remain unclear. In order to profile proteins from the nodal segment, callus, root and shoot, high throughput proteome approach was executed in the present study. Two dimensional gels stained with CBB, a total of 84 differential expressed proteins were confirmed out of 839 protein spots using image analysis by Progenesis SameSpot software. Out of total differential expressed spots, 58 differential expressed protein spots (${\geq}$ 2-fold) were analyzed using MASCOT search engine according to the similarity of sequences with previously characterized proteins along with the UniProt database. Out of 58 differential expressed protein, 32 protein spots were up-regulated such as ribulose-1,5-bisphosphate carboxylase, endoplasmic oxidoreductin-1, heat stress transcription factor A3, RNA pseudourine synthase 4, cysteine proteinase, GntR family transcriptional regulator, E3 xyloglucan 6-xylosyltransferase, while 26 differential protein spots were down-regulated such as L-ascorbate oxidase precursor, late embryogenesis abundant protein D-34, putative SCO1 protein, oxygen-evolving enhancer protein 3. However, frequency distribution of identified proteins using iProClass databases, and assignment by function based on gene ontology revealed that the identified proteins from the explants were mainly associated with the nucleic acid binding (17%), transferase activity (14%) and ion binding (12%). In that way, the exclusive protein profile may provide insight clues for better understanding the characteristics of proteins and metabolic activity in various explants of the economically important medicinal plant Platycodon grandiflorum.