• Journal of Microbiology and Biotechnology
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• 제17권2호
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• pp.244-252
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• 2007

Escherichia coli has a PhoR-PhoB two-component regulatory system to detect and respond to the changes of environmental phosphate concentration. For the E. coli W3110 strain growing under phosphate-limiting condition, the changes of global gene expression levels were investigated by using DNA microarray analysis. The expression levels of some genes that are involved in phosphate metabolism were increased as phosphate became limited, whereas those of the genes involved in ribosomal protein or amino acid metabolism were decreased, owing to the stationary phase response. The upregulated genes could be divided into temporarily and permanently inducible genes by phosphate starvation. At the peak point showing the highest expression levels of the phoB and phoR genes under phosphate-limiting condition, the phoB- and/or phoR-dependent regulatory mechanisms were investigated in detail by comparing the gene expression levels among the wild-type and phoB and/or phoR mutant strains. Overall, the phoB mutation was epistatic over the phoR mutation. It was found that PhoBR and PhoB were responsible for the upregulation of the phosphonate or glycerol phosphate metabolism and high-affinity phosphate transport system, respectively. These results show the complex regulation by the PhoR-PhoB two-component regulatory system in E. coli.

• 미생물학회지
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• 제46권2호
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• pp.113-121
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• 2010

인산 결핍기에 직면한 Bacillus subtilis는 PhoP-PhoR twocomponent system (TCS)를 통해 이러한 상황을 인식하고 생존을 유지하기 위해 Pho regulon으로 불리는 일련의 유전자들의 발현을 조절한다. 이때 histidine kinase인 PhoR은 자동 인산화되어, 인산을 response regulator인 PhoP에 전달한다. 인산화된 PhoP (PhoP~P)는 Pho regulon 유전자의 프로모터(promoter) 부위에 존재하는 반복되는 6 bp의 잘 보존된 PhoP 결합서열에 결합하여 해당 유전자의 발현을 활성화시키거나 억제한다. 이러한 Pho regulon 신호전달 시스템은 최소한 세 개의 TCS (PhoP-PhoR, ResD-ResE TCS, SpoOA phosphorelay), 광범위한 탄소대사 조절자(CcpA), 전위기 조절자(AbrB, ScoC) 등을 포함하는 신호전달 시스템과 밀접하게 상호 연결되어 있을 뿐만 아니라, 생육에 필수적인 YycF-YycG TCS와 상호조절을 통한 밀접한 관련을 가지고 있다. Pho regulon에 의한 인산결핍 스트레스 반응을 이해하는데 많은 진척이 있었으나, 많은 의문들은 여전히 남아있다. 이러한 의문들을 푸는 일은 B.subtilis의 응용연구에 중요한 정보를 제공할 것이다.

• Journal of Microbiology and Biotechnology
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• 제26권9호
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• pp.1620-1628
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• 2016

In this study, we investigated colistin resistance mechanisms associated with the regulation of the pbgP operon in Klebsiella pneumoniae, using four isogenic pairs of colistin-susceptible strains and their colistin-resistant derivatives and two colistin-resistant clinical isolates. Amino acid sequence alterations of PhoPQ, PmrAB, and MgrB were investigated, and mRNA expression levels of phoQ, pmrB, pmrD, and pbgP were measured using quantitative real-time PCR. The phoQ and pmrB genes were deleted from two colistin-resistant derivatives, 134R and 063R. We found that phoQ, pmrD, and pbgP were significantly upregulated in all colistin-resistant derivatives. However, pmrB was significantly upregulated in only two colistin-resistant derivatives and one clinical strain. pmrB was not overexpressed in the other strains. The minimum inhibitory concentration of colistin was drastically lower in both phoQ- and pmrB-deleted mutants from a colistin-resistant derivative (134R) that was overexpressing phoQ and pmrB. However, colistin susceptibility was restored only in a phoQ-deleted mutant from a colistin-resistant derivative (063R) without overexpression of pmrB. In conclusion, two different regulations of the pbgP operon may associate with the development of colistinresisant K. pneumoniae.

• 미생물학회지
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• 제48권2호
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• pp.57-65
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• 2012

인은 인지질, 탄수화물 및 핵산 등의 생분자 합성에 필요한 원소이다. 세균은 외부환경으로부터 인산이나 인산을 포함하는 영양소를 흡수하여 인을 얻고, 세포대사에 사용되고 남은 인산은 polyphosphate 형태로 저장한다. 현재까지 알려진 다섯 개의 인산 수송 시스템 중, 인산에 특이적으로 높은 친화력을 갖는 Pst 시스템이 가장 중요한 역할을 하며, 그 발현은 세포외부 인산 농도에 반응하는 PhoB-PhoR two component 신호전달 시스템에 의해 조절된다. 반응 조절 단백질 PhoB는 인산 대사뿐 아니라 이와 관계없는 유전자들의 전사를 조절하는 것으로 알려졌으며, 따라서 PhoB의 활성이 조절되지 않으면 많은 종류의 다른 표현형이 나타난다. 본 총설은 각 인산 수송 시스템의 기능이 결여된 세균의 표현형에 대한 최근 연구 결과를 토대로 다음과 같은 내용을 기술하였다. 첫째, 세포 내부 인산의 적정 농도 유지를 위한 인산 수송 시스템들의 역할, 둘째, 인산뿐 아니라 여타 환경 신호와 관련된 수송 시스템의 다양한 표현형, 그리고 마지막으로, 수송 시스템들 간 혹은 그 조절자들 간의 표현형 중복을 분류하여 제시하였다. 이러한 내용은 결국 세균의 대사, 적응반응 및 병원성 발현에 미치는 인산 항상성의 중요성을 강조한다.

• Journal of Microbiology and Biotechnology
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• 제33권9호
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• pp.1130-1140
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• 2023

Among the AAA+ proteases in bacteria, FtsH is a membrane-bound ATP-dependent metalloprotease, which is known to degrade many membrane proteins as well as some cytoplasmic proteins. In the intracellular pathogen Salmonella enterica serovar Typhimurium, FtsH is responsible for the proteolysis of several proteins including MgtC virulence factor and MgtA/MgtB Mg²⁺ transporters, the transcription of which is controlled by the PhoP/PhoQ two-component regulatory system. Given that PhoP response regulator itself is a cytoplasmic protein and also degraded by the cytoplasmic ClpAP protease, it seems unlikely that FtsH affects PhoP protein levels. Here we report an unexpected role of the FtsH protease protecting PhoP proteolysis from cytoplasmic ClpAP protease. In FtsH-depleted condition, PhoP protein levels decrease by ClpAP proteolysis, lowering protein levels of PhoP-controlled genes. This suggests that FtsH is required for normal activation of PhoP transcription factor. FtsH does not degrade PhoP protein but directly binds to PhoP, thus sequestering PhoP from ClpAP-mediated proteolysis. FtsH's protective effect on PhoP can be overcome by providing excess ClpP. Because PhoP is required for Salmonella's survival inside macrophages and mouse virulence, these data implicate that FtsH's sequestration of PhoP from ClpAP-mediated proteolysis is a mechanism ensuring the amount of PhoP protein during Salmonella infection.

• 미생물학회지
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• 제38권4호
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• pp.318-321
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• 2002

토양 등의 인 제한환경에서 특이적으로 발현하는 벡터를 구축하기 위해서 Enterobacter areogenes의 phoA 유전자의 promoter가 든 pEAAP를 구축하였다. pEAAP는 pET-22b(+)을 BglII와 XbaI으로 절단하여 T7 promoter와 lac operator를 제거하고pho box가 포함된 phoA promoter를 삽입하여 구축하였다. pEAAP가 인 제한 환경에서 특이적으로 발현되는지 조사하고자 Bacillus subtillis var. amyloliquefaciens (KCTC 8913P)의 Phytase유전자인 Bsa-phy1을 도입한 pEAPHY1을 구축하였다. CK-PHY1 (pEAPHY1을 도입한Escherichia coli JM109)는 인 제한 환경에서 41 kD)의 Bsa-Phy1을 발현하였다. 또한, CK-PHY1은 phytate를 유일한 인산원으로 첨가된 고체배지에서 phytate를 분해하여 투명대를 형성하였다.

• 생명과학회지
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• 제19권5호
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• pp.566-572
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• 2009

S. marcescens KCTC 2172로부터 유전자 은행을 작성하여 재조합 클론 pDH3를 얻었으며, pDH3 유래의 서브클론을 작성하였다. 플라스미드 pPH4의 전염기서열 5,137 bp 영역을 결정한 결과 3개의 ORF가 있음을 확인하였다. 이들은 pst 오페론의 pstC, pstA, 및 pstB, 세 유전자를 동일 전사방향으로 코드하고 있었다. 타 세균의 유전자와 비교한 결과 S. marcescens의 pst 오페론은 pstS와 phoU가 결손되어 있다. 조절영역에는 CRP 결합영역과 pho box 서열이 존재하였다. 보고된 유전자와 상동성 조사결과, PstC 단백질은 Yersinia sp., Vibrio sp. 및 Pseudomonas sp.와는 49, 37, 33%의 상동성을, PstA 단백질은 Yersinia sp., Vibrio sp. 및 Pseudomonas sp.와 64, 51, 47%의 상동성을, PstB 단백질은 Methanocaldococcus sp., E. coli 및 Mycoplasma sp.와 60, 50, 48%의 상동성을 나타내었다. Pst 유전자들은 조절영역의 cAMP-CRP 복합체에 의해 in vivo에서 양성적으로 발현됨을 확인하였다. Pst 오페론을 포함하는 플라스미드를 도입한 대장균은 인산운송에 관여하는 능력을 확인하였다.

• Journal of Microbiology and Biotechnology
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• 제5권3호
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• pp.125-131
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• 1995

A 3, 346 bp of the cdd upstream region in Bacillus subtilis was sequenced from the pSO1 (Song BH and J Neuhard. 1989. Mol. Gen. Genet 216: 462-468) and sequence homology was searched to the known genes in Genbank and European Molecular Biology Laboratory databanks. Five complete and one truncated putative coding sequences deduced from the nucleotide sequence were found through the ORF searching by Genetyx and Macvector software, and one of them was identified as the dgk (diacylglycerol kinase) gene and another, a truncated one, as the phoH (phosphate starvation-inducible gene) gene. The B. subtilis dgk gene, having a role for response to several environmental stress signals, revealed an open reading frame of 134 amino acids with 43.1% of sequence identity to the Streptococcus mutans dgk gene. The carboxy terminal 59 residues of the truncated phoH gene showed 52.7% and 34.5% of sequence identity in amino acids with the corresponding genes of Mycobacterium leprae and Escherichia coli. The four remaining coding sequences consisting of 115, 421, 91, and 91 residues were thought to be unknown ORFs because they have no significant similarity to known genes.

• Molecules and Cells
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• 제44권3호
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• pp.179-185
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• 2021

Vancomycin response regulator (VncR) is a pneumococcal response regulator of the VncRS two-component signal transduction system (TCS) of Streptococcus pneumoniae. VncRS regulates bacterial autolysis and vancomycin resistance. VncR contains two different functional domains, the N-terminal receiver domain and C-terminal effector domain. Here, we investigated VncR C-terminal DNA binding domain (VncRc) structure using a crystallization approach. Crystallization was performed using the micro-batch method. The crystals diffracted to a 1.964 Å resolution and belonged to space group P212121. The crystal unit-cell parameters were a = 25.71 Å, b = 52.97 Å, and c = 60.61 Å. The structure of VncRc had a helix-turn-helix motif highly similar to the response regulator PhoB of Escherichia coli. In isothermal titration calorimetry and size exclusion chromatography results, VncR formed a complex with VncS, a sensor histidine kinase of pneumococcal TCS. Determination of VncR structure will provide insight into the mechanism by how VncR binds to target genes.

• 한국초지조사료학회지
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• 제17권3호
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• pp.285-292
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• 1997

This study was conducted to obtain the transgenic Brnssica napus plants with tobacco Apase gene using the binary vector system of Agrobacteriurn fumefociens. The results obtained were summarized as follows: A repressible acid phosphatase gene of Saccharon~yces cerevisiae, pho105 was used for screening of tobacco Apase cDNA. In order to identify Apase gene in tobacco genome, Southern blot analysis was pcrformed and the Apase gcnc may be present as a single copy, or at most two or three copies, in tobacco genome. To isolate the tobacco Apase gene, tobacco cDNA library was constructed using purifed mRNA from -Pi treated tobacco root and the plaque forming unit of the library was 2.8 x $10^5$ pfu/m${\ell}$, therefore the library might cover all expressed mRNAs. Using pho5 as a probe. tobacco Apase cDNA was cloned, and restriction mapping and Southern blot analysis of cDNA insert were revealed that the 3.6 kb cDNA contained tobacco acid phosphatase cDNA. Plasmid pGA695 -tcAPl was constructed by subcloning tobacco Apase cDNA into the Hind site of pGA695 with 35s promoter which can be expressed constitutively in plants. The Brassica napus cotyledonary petioles were cocultivated with the ,4 grobacteriunz and transferred to the selection medium. The transformed and regenerated plants were transplanted to soil medium. Southern blot analysis was done on the transformed plants, and it was confirmed that a foregin gene was stably integrated into the genonies of B. nnpus plants.