• Journal of Applied Biological Chemistry
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• v.48 no.4
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• pp.218-221
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• 2005

Nucleotide sequence of Brevibacterium flavum ptsG gene capable of complementing Escherichia coli ZSC113 mutations defective to glucose permease activity of phosphotransferase system was completely determined, and the gene product was compared with other glucose-specific enzyme II ($EII^{Glc}$). A ptsG gene of B. flavum consisted of open reading frame of 2,025 nucleotides putatively encoding polypeptide of 675 amino acid residues and TAA stop codon. Deduced amino acid sequence of B. flavum ($EII^{Glc}$) had high homology with ($EIIs^{Glc}$) of Corynebacterium glutamicum, C. efficiens, and B. lactofermentum. Arrangement of structural domains, IIBCA, of B. flanum ($EII^{Glc}$) protein was identical to that of EIIs belonging to glucose-phosphotransferase system.

• Journal of Microbiology and Biotechnology
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• v.32 no.8
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• pp.1047-1053
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• 2022

When ptsG, a glucose-specific phosphotransferase system (PTS) component, is deleted in Escherichia coli, growth can be severely poor because of the lack of efficient glucose transport. We discovered a new PTS transport system that could transport glucose through the growth-coupled experimental evolution of ptsG-deficient E. coli C strain under anaerobic conditions. Genome sequencing revealed mutations in agaR, which encodes a repressor of N-acetylgalactosamine (Aga) PTS expression in evolved progeny strains. RT-qPCR analysis showed that the expression of Aga PTS gene increased because of the loss-of-function of agaR. We confirmed the efficient Aga PTS-mediated glucose uptake by genetic complementation and anaerobic fermentation. We discussed the discovery of new glucose transporter in terms of different genetic backgrounds of E. coli strains, and the relationship between the pattern of mixed-acids fermentation and glucose transport rate.

• Journal of Microbiology and Biotechnology
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• v.3 no.1
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• pp.1-5
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• 1993

The gene for mannose enzyme II of phosphoenolpyruvate-dependent phosphotransferase system from Corynebacterium glutamicum KCTC 1445 was cloned into Escherichia coli ZSC113 using plasmid pBR 322. The recombinant plasmid, designated pCTS3, contained 2.2 kb DNA fragment, and the physical map of the cloned DNA fragment was determined. The E. coli ptsM ptsG mutant transformed with pCTS3 restored glucose and mannose fermentation ability, and grew well on these sugars as the sole carbon source in the minimal medium. The transform ant harboring pCTS3 showed a PTS-mediated repression of growth on maltose by mannose analogue, 2-deoxyglucose. The specificity of the response to 2DG therefore indicates that the cloned DNA fragment carries mannose enzyme II gene.

• Journal of Microbiology and Biotechnology
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• v.9 no.5
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• pp.582-588
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• 1999

A Brevibacterium lactofermentum gene coding for a glucose-specific permease of the phosphoenolpyruvate-dependent phosphotransferase system (PTS) was cloned, by complementing an Escherichia coli mutation affecting a ptsG gene with the B. lactofermentum genomic library, and completely sequenced. The gene was identified as a ptsG, which enables an E. coli transformant to transport non-metabolizable glucose analogue 2-deoxyglucose (2DG). The ptsG gene of B. lactofermentum consists of an open reading frame of 2,025 nucleotides encoding a polypeptide of 674 amino acid residues and a TAA stop codon. The 3' flanking region contains two stem-loop structures which may be involved in transcriptional termination. The deduced amino acid sequence of the B. lactofermentum enzyme $II^{GIe}$ specific to glucose ($EII^{GIe}$) has a high homology with the Corynebacterium glutamicum enzyme $II^{Man}$ specific to glucose and mannose ($EII^{Man}$), and the Brevibacterium ammoniagenes enzyme $II^{GIc}$ specific to glucose ($EII^{GIc}$). The 171-amino-acid C-terminal sequence of the $EII^{Glc}$ is also similar to the Escherichia coli enzyme $IIA^{GIc}$ specific to glucose ($IIA^{GIc}$). It is interesting that the arrangement of the structural domains, IIBCA, of the B. lactofermentum $EII^{GIc}$ protein is identical to that of EIIs specific to sucrose or $\beta$-glucoside. Several in vivo complementation studies indicated that the B. lactofermentum $EII^{Glc}$ protein could replace both $EII^{ Glc}$ and $EIIA^{Glc}$ in an E. coli ptsG mutant or crr mutant, respectively.

• Journal of Microbiology and Biotechnology
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• v.8 no.3
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• pp.214-221
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• 1998

A Brevibacterium ammoniagenes gene coding for glucose/mannose-specific enzyme II ($EII^{Glc}$) of the phosphoenolpyruvate-dependent phosphotransferase system (PTS) was cloned by complementing an Escherichia coli mutation affecting a ptsG gene, and the complete DNA nucleotide sequence was determined. The cloned gene was identified to be a ptsG, which enables the E. coli transportment to use glucose more efficiently than mannose as the sole carbon source in an M9 minimal medium. The ptsG gene of B. ammoniagenes consists of an open reading frame of 1,983 nucleotides putatively encoding a polypeptide of 661 amino acid residues and a TAA stop codon. The deduced amino acid sequence of the B. ammoniagenes $EII^{Glc}$ shows, at $46\%$, the highest degree of sequence similarity with the Corynebacterium glutamicum EII specific for both glucose and mannose. In addition, the $EII^{Glc}$ shares approximately $30\%$ sequence similarities with sucrose-specific and ${\beta}$-glucoside-specific EIIs of the several bacteria belonging to the glucose-PTS class. The 161-amino-acid C-terminal sequence of $EII^{Glc}$ is also similar to that of E. coli enzyme $IIA^{Glc}$, specific for glucose ($EIIA^{Glc}$). The B. ammoniagenes $EII^{Glc}$ consists of three domains; a hydrophobic region (EIIC) and two hydrophilic regions (EIIA, EIIB). The arrangement of structural domains, IIBCA, of the $EII^{Glc}$ is identical to those of EIIs specific for sucrose or ${\beta}$-glucoside. While the domain IIA was removed from the B. ammoniagenes $EII^{Glc}$ the remaining domains IIBC were found to restore the glucose and mannose-utilizing capacity of E. coli mutant lacking $EII^{Glc}$ activity with $EIIA^{Glc}$ of the E. coli mutant. $EII^{Glc}$ contains a histidine residue and a cysteine residue which are putative phosphorylation sites for the protein.

• Applied Biological Chemistry
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• v.42 no.4
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• pp.293-297
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• 1999

The pts operon, which encodes several factors in the phosphoenolpyruvate:carbohydrate phosphotransferase system (PTS) of Escherichia coli, has multiple promoters which respond to different signals to facilitate quick adaptation to changes in growth conditions. The influence of an 1 kbp DNA region upstream of the pts P0 promoter on pts expression was studied in vitro by employing the DNA templates containing both P0 and P1 promoter with or without the 1 kbp upstream DNA region for in vitro transcription assay. The 1 kbp DNA region upstream of the pts P0 promoter, however, had no effect on pts transcription in vitro. The intracellular concentration of cAMP was measured when cells were grown in the presence of glucose, mannose, or mannitol. The transcription of P0 was increased maximally in the presence of glucose even though the concentration of cAMP in the condition was lowest while the transcription from the P1b was highest when cells were grown in the presence of mannose or mannitol even though the intracellular concentration of cAMP was lower than cells grown in the absence of the sugar. These results suggest the possibility of the existence of a glucose inducible repressor specific for the P0 promoter and a second repressor that is inducible by glucose, mannose and mannitol specific for the P1 promoter.

• Journal of Microbiology and Biotechnology
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• v.5 no.4
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• pp.188-193
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• 1995

A Brevibacterium flavum gene coding for glucose permease of the phosphoenolpyruvate-dependent phosphotransferase system (PTS) was cloned by complementing the Escherichia coli ZSCl13 mutations affecting a ptsG gene with the B. flavum genomic library. From the E. coli clone grown as red colony on a MacConkey plate supplemented with glucose as an additional carbon source, a recombinant plasmid was isolated and named pBFT93. The plasmid pBFT93 was identified as carrying a 3.6-kb fragment of B. flavum chromosomal DNA which enables the E. coli transformant to use glucose or man nose as a sole carbon source in an M9 minimal medium. The non-metabolizable sugar analogues, 2-deoxy-D-glucose (2-DG) and methyl-$\alpha$-D-glucopyranoside (MeGlc) affected the growth of ZSCl13 cells carrying the plasmid pBFT93 on minimal medium supplemented with non-PTS carbohydrate, glycerol, as a sole cabon source, while the analogues did not repress the growth of ZSCl13 cells without pBFT93. It was also found that both $2-deoxy-D-[U-^{14}C]glucose{\;}and{\;}methyl-{\alpha}-D-[U-^{14}C]glucopyranoside$ could be effectively transported into ZSCl13 cells transformed with plasmid pBFT93. Several in vivo complementation studies suggested that the B. flavum DNA in pBFT93 encodes a glucose permease specific for glucose and mannose.

• Journal of Microbiology and Biotechnology
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• v.16 no.5
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• pp.795-798
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• 2006

The phosphoenolpyruvate-dependent carbohydrate phosphotransferase system (PTS) is responsible for the simultaneous transfer and phosphorylation of various carbon sources in Escherichia coli. The ptsG gene encoding the enzyme $IICB^{Glc}$, the membrane component of the glucose-specific PTS, is repressed by Mlc and activated by the CRP cAMP complex; various other factors, such as Fis, FruR, and ArcA, are also known to be involved in ptsG regulation. Thus, in an attempt to discover a novel gene affecting the regulation of ptsG, a mutant with a decreased ptsG transcription in the presence of glucose compared with the wild-type strain was screened using transposon random mutagenesis. The mutant was found to have a transposon insertion in yhjV, a putative gene encoding a transporter protein whose function is yet unknown.

• Korean Journal of Microbiology
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• v.27 no.3
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• pp.181-187
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• 1989

Plasmid DNA was isolated from Lactobacillus casei SW-M1($Lac^{+}$strain). The curing frequencies of pPLac plasmid from L. casei SW-M1 showed 43% for acriflavin treatment and 53% for ethidium bromide treatment after 3 times transfer. On the charaterization of pPLac plasmid, it was found that the plasmid contained gene encoding phospho-$\beta$-galactosidase for lactose utilization. Lactose-PTS(phosphotransferase system)was involved in membrane transport system in $Lac^{+}$ strain. Induction of phospho-$\beta$-galactosidase was specially effective by galactose, lower effect with lactose and glucose but not by IPTG(isopropyl-$\beta$-D-thiogalactoside). This result showed that induction of phospho-$\beta$-galactosidase by IPTG did not appeared. The catabolite repression of phospho-$\beta$-galactosidase synthesis by glucose was not found in L. casei.

• Korean Journal of Microbiology
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• v.39 no.3
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• pp.141-147
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• 2003

The simultaneous expression levels of antifungal activity related genes was analyzed by DNA microarray. We constructed DNA chips contained 2,000 randomly digested genome spots of the antifungal bacterium of Bacillus lentimorbus WJ5 and compared its quantitative aspect with 7 antifungal activity deficient mutants induced by gamma radiation ($^{60}Co$). From the analysis of microarray hybridization by the Gene Cluster (Michael Eisen, Stanford Univ.), totally 408 genes were expressed and 20 genes among them were significantly suppressed in mutants. pbuX (xanthine permease, K222), ywbA (phosphotransferase system enzyme II, K393), ptsG (PTS glucose specific enzyme II ABC component, K877), yufO (ABC transporter (ATP-binding protein), K130l), and ftsY (signal recognition particle (docking protein), K868) were simultaneously down-regulated in all mutants. It suggested that they were supposed to be related to the antifungal activity of B. lentimorbus WJ5.