• Title/Summary/Keyword: purE

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Identification of Potential Corynebacterium ammoniagenes Purine Gene Regulators Using the pur-lacZ Reporter in Escherichia coli

  • HAN , RI-NA;CHO, ICK-HYUN;CHUNG, SUNG-OH;HAN, JONG-KWON;LEE, JIN-HOO;KIM, SOO-KI;CHOI, KANG-YELL
    • Journal of Microbiology and Biotechnology
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    • v.14 no.6
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    • pp.1249-1255
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    • 2004
  • This study has developed Corynebacterium ammoniagenes (c. ammoniagenes) purine gene transcriptional reporters (purF-lacZ and purE-lacZ) that function in Escherichia coli (E. coli) DH5a. After transformation of a C. ammoniagenes gDNA library into E. coli cells harboring either purF-lacZ or purE-lacZ, C. ammoniagenes clones were obtained that repress purF-lacZ and purE-lacZ gene expression. The potential purE and purF regulatory genes are homologous to the genes encoding transcription regulators, the regulatory subunit of RNA polymerase, and genes for purine nucleotide biosynthesis of various bacteria. The C. ammoniagenes purE-lacZ and purF-lacZ reporters were repressed by adenine and guanine within E. coli, indicating similarity in the regulatory mechanism of purine biosynthesis in C. ammoniagenes and E. coli. Gene regulation of pur-lacZ by adenine and guanine was partly abolished in cells expressing potential purine regulatory genes, indicating functionality of the purine gene regulators in repression of purE-lacZ and purF-lacZ. The purE-lacZ and purF-lacZ reporters can be used for the screening of genes involved in the regulation of the de novo synthesis of the purine nucleotides.

Regulation of Corynebacterium ammoniagenes purF and Isolation of purF-Specific Regulatory Proteins (Corynebacterium ammoniagenes에서 purF 유전자의 조절 및 이에 특이적인 조절 단백질의 분리)

  • Lee, Seok-Myung;Kim, Youn-Hee;Lee, Heung-Shick
    • Korean Journal of Microbiology
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    • v.45 no.3
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    • pp.233-238
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    • 2009
  • The expression of Corynebacterium ammoniagenes purF was analyzed by utilizing a plasmid carrying a cat gene fused to the purF promoter region. Adenine and guanine repressed the expression of the purF gene by 20~30% but hypoxanthine did not exert such repressive effect. The expression purF was maximal at the late log phase and remained constant throughout the stationary phase. Promoter $P_{180}$ which was developed in C. glutamicum was also functional in C. ammoniagenes, achieving maximal activity at the late log phase. The promoter outperformed Escherichia coli $P_{tac}$ promoter by 40~50% level. DNA-affinity purification identified a protein which could bind to the promoter region of the purF gene. The protein showed high similarity to the CRP-family transcriptional regulator encoded by NCgl0120 in C. glutamicum. The size of the screened protein agreed with the expected protein size from the ORF NCgl0120. The corresponding gene in C. ammoniagenes encoded a 42 kDa polypeptide composed of 400 amino acids with expected pI of 4.9. The encoded protein showed 14.1% and 15.8% identity with E. coli and Bacillus subtilis PurR, respectively, suggesting that the isolated protein might be a novel type of regulatory protein involved in the regulation of purine metabolism.

Intramolecular DNA Triplexes in Escherichia. coli (Methylase를 사용한 Escherichia coli에서 Triplex 존재에 관한 연구)

  • Rhim, Hyangshuk;Kim, Sungjo;Kang, Seongman
    • Korean Journal of Microbiology
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    • v.34 no.4
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    • pp.231-235
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    • 1998
  • We have introduced a genetic assay to study the existence of intramolecular triplexes in Escherichia coli. A plasmid containing the gene that encodes a temperature-sensitive EcoRI methylase was cotransformed with different plasmids containing inserts, $(G)_9AATTC(G)_9$ and $(GAA)_9TTC(GAA)_8$, that are able to form intramolecular triplexes in vitro. Inhibition of methylation in vivo was found for $(G)_9AATTC(G)_9$ and $(GAA)_9TTC(GAA)_8$, suggesting that the pur pyr sequences adopt unusual strucures in E. coli. In addition, experiments using two dimensional gel electrophoresis confirmed that intramolecular triplexes are formed for the pur pyr sequences under negative supercoiling. These results demonstrate the existence of intramolecular triplexes in E. coli.

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Effect of Pile Driving on Three Layered Pipeline according to Soil Properties Variation (지반 물성값에 따른 항타 진동이 지중 삼중관에 미치는 거동 분석)

  • Yoo, Han-Kyu;Choi, Joung-Hyun;Won, Jong-Hwa;Kim, Moon-Kyum
    • Proceedings of the Korean Geotechical Society Conference
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    • 2010.03a
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    • pp.765-770
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    • 2010
  • In this study, the behavior of underground pipeline subjected to pile driving is examined using the verified finite element model based on the field experiment. Young's modules of surface soil is varied and elastic modulus of the other soil layer is fixed. The pile driving force model proposed by Mounir E. Mabsout in 1999 was used and it was functions of time and of force. The forcing function applied on this study considers the kinetic energy of ram located at 1.2m height with 7 tonf. The 3-layered pipeline is composed of steel(inner) pipe, PUR(Polyurethane Resin, filler) and HDPE(outer) pipe, and the length/diameter of main steel pipe is 20m/0.8m(O.D). It is used for district heating pipes in Korea. The results are expressed in terms of Von Mises stress, displacement, and vibration velocity for each soil condition. From the results of the analyses, PUR which is originally intended as a thermal insulation of inner pipe shows performance as a structural member which distributes external pressure.

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A Study on the Fire Characteristics of Sandwich Panels by ISO 9705 - Full-scale Room Test (ISO 9705 - Full-scale Room Test 시험방법을 이용한 샌드위치 패널의 화재 특성 연구)

  • Kweon, Oh-Sang;Yoo, Yong-Ho;Kim, Heung-Youl;Lee, Jeong-Il
    • Fire Science and Engineering
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    • v.23 no.6
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    • pp.39-45
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    • 2009
  • This paper studied on the fire characteristics of sandwich panels by ISO 9705 test (Full-scale room test). A comprehensive fire characteristics of sandwich panel was analyzed by applying the test result to the classification standard according to EN 13501-1 and Eurefic Research Program. Consequently, glass wool foam sandwich panel proved to be A class, incombustible - EPS Foam and incombustible - PUR Foam sandwich panels was class B. Also, EPS foam and PUR foam sandwich panels was class C because of flashover.

Infection status with helminthes in feral cats pur-chased from a market in Busan, Republic of Korea

  • SOHN Woon-Mok;CHAI Jong-Yil
    • Parasites, Hosts and Diseases
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    • v.43 no.3 s.135
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    • pp.93-100
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    • 2005
  • The present study was performed to investigate the infection status with helminth in a group of feral cats in Korea. More than 29 helminth species including adults or eggs were detected in visceral and fecal samples of the examined cats. Among these were a host of nematodes, including toxocarids, Ancylostoma sp. and the larva of Anisakis simplex; trematodes, including Clonorchis sinensis, Paragonimus westermani, Eurytrema pancreaticum, Pharyngostomum cordatum, Metagonimus spp., Heterophyes nocens, Pygidiopsis summa, Heterophyopsis continua, Stictodora fuscata, Stictodora lari, Acanthotrema felis, Stellantchasmus falcatus, Centrocestus armatus, Procerovum varium, Cryptocotyle sp., Echinostoma revolutum, Echinostoma hortense, Echinochasmus japonicus, Stephanoprora sp., Plagiorchis muris, Neodiplostomum sp. and diplostomulum. We also detected a variety of cestodes, including Spirometra erinacei, Taenia taeniaeformis and unidentified species of tapeworm. We also found examples of the acanthocephalan, Bolbosoma sp. In our assessment of the stools, we detected at least 12 species of helminth eggs. These findings confirmed that feral cats in Korea are infected with a variety of helminth parasite species. Furthermore, among the helminths detected, E. pancreaticum, S. fuscata, S. lari, A. felis, S. falcatus, C. armatus, P. varium, Cryptocotyle sp., E. revolutum, E. japonicus, Stephanoprora sp., P. muris, Neodiplostomum sp. and Bolbosoma sp. represent helminth fauna which have not been reported previously in feral cats in the Republic of Korea.

PLP-1 Binds Nematode Double-stranded Telomeric DNA

  • Im, Seol Hee;Lee, Junho
    • Molecules and Cells
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    • v.20 no.2
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    • pp.297-302
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    • 2005
  • The integrity and proper functioning of telomeres require association of telomeric DNA sequences with specific binding proteins. We have characterized PLP-1, a $PUR{\alpha}$ homolog encoded by F45E4.2, which we previously identified as a candidate double stranded telomere binding protein, by affinity chromatography followed by mass spectrometry. PLP-1 bound double-stranded telomeric DNA in vitro as shown by competition assays. Core binding was provided by the third and fourth nucleotides of the TTAGGC telomeric repeat. This is quite different from the binding sequence of CEH-37, another C. elegans telomere binding protein, suggesting that multiple proteins may bind nematode telomeric DNA simultaneously in vivo.

Transcriptional Analysis Responding to Propanol Stress in Escherichia coli (대장균에서 프로판올 스트레스에 관한 전사분석)

  • Park, Hye-Jin;Lee, Jin-Ho
    • Journal of Life Science
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    • v.22 no.3
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    • pp.417-427
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    • 2012
  • We compared the transcriptome in response to propanol stress in wild-type and propanol-resistant mutant Escherichia coli using the DNA microarray technique. The correlation value of RNA expression between the propanol-treated wild type and the untreated-one was about 0.949, and 50 genes were differentially expressed by more than twofold in both samples. The correlation value of RNA expression between the propanol-treated mutant and the untreated one was about 0.951, and 71 genes in two samples showed differential expression patterns. However, the values between the wild type and mutant, regardless of propanol addition, were 0.974-0.992 and only 1-2 genes were differentially expressed in the two strains. The representative characteristics among differentially expressed genes in W3110 or P19 treated with propanol compared to untreated samples were up-regulation of hest shock response genes and down-regulation of genes relating to ribosome biosynthesis. In addition, many genes were regulated by transcription regulation factors such as ArcA, CRP, FNR, H-NS, GatR, or PurR and overexpressed by sigma factor RpoH. We confirmed that RpoH mediated an important host defense function in propanol stress in E. coli W3110 and P19 by comparison of cell growth rate among the wild type, rpoH disruptant mutant, and rpoH-complemented strain.

Uropathogenic Escherichia coli ST131 in urinary tract infections in children

  • Yun, Ki Wook;Lee, Mi-Kyung;Kim, Wonyong;Lim, In Seok
    • Clinical and Experimental Pediatrics
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    • v.60 no.7
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    • pp.221-226
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    • 2017
  • Purpose: Escherichia coli sequence type (ST) 131, a multidrug-resistant clone causing extraintestinal infections, has rapidly become prevalent worldwide. However, the epidemiological and clinical features of pediatric infections are poorly understood. We aimed to explore the characteristics of ST131 Escherichia coli isolated from Korean children with urinary tract infections. Methods: We examined 114 uropathogenic E. coli (UPEC) isolates from children hospitalized at Chung-Ang University Hospital between 2011 and 2014. Bacterial strains were classified into STs by partial sequencing of seven housekeeping genes (adk, fumC, gyrB, icd, mdh, purA, and recA). Clinical characteristics and antimicrobial susceptibility were compared between ST131 and non-ST131 UPEC isolates. Results: Sixteen UPEC isolates (14.0%) were extended-spectrum ${\beta}-lactamase$ (ESBL)-producers; 50.0% of ESBL-producers were ST131 isolates. Of all the isolates tested, 13.2% (15 of 114) were classified as ST131. There were no statistically significant associations between ST131 and age, sex, or clinical characteristics, including fever, white blood cell counts in urine and serum, C-reactive protein, radiologic abnormalities, and clinical outcome. However, ST131 isolates showed significantly lower rates of susceptibility to cefazolin (26.7%), cefotaxime (40.0%), cefepime (40.0%), and ciprofloxacin (53.3%) than non-ST131 isolates (65.7%, 91.9%, 92.9%, and 87.9%, respectively; P<0.001 for all). ESBL was more frequently produced in ST131 (53.3%) than in non-ST131 (8.1%) isolates (P<0.01). Conclusion: ST131 E. coli isolates were prevalent uropathogens in children at a single medical center in Korea between 2011 and 2014. Although ST131 isolates showed higher rates of antimicrobial resistance, clinical presentation and outcomes of patients were similar to those of patients infected with non-ST131 isolates.

Characterization of the Starch Degradation Activity of recombinant glucoamylase from Extremophile Deinococcus geothermalis (극한성 미생물Deinococcus geothermalis 유래 재조합 글루코아밀레이즈의 전분 분해 활성 특징)

  • Jang, Seung-Won;Kwon, Deok-Ho;Park, Jae-Bum;Jung, Jong-Hyun;Ha, Suk-Jin
    • Journal of Industrial Technology
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    • v.39 no.1
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    • pp.15-19
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    • 2019
  • This work focused on characterization of the starch degradation activity from extremophile strain Deinococcus geothermalis. Glucoamylase gene from D. geothermalis was cloned and overexpressed by pET-21a vector using E. coli BL21 (DE3). In order to characterize starch degrading activity of recombinant glucoamylase, enzyme was purified using HisPur Ni-NTA column. The recombinant glucoamylase from D. geothermalis exhibited the optimum temperature as $45^{\circ}C$ for starch degradation activity. And highly acido-stable starch degrading activity was shown at pH 2. For further optimization of starch degrading activity with metal ion, various metal ions ($AgCl_2$, $HgCl_2$, $MnSO_4{\cdot}4H_2O$, $CoCl_2{\cdot}6H_2O$, $MgSO_4$, $ZnSO_4{\cdot}7H_2O$, $K_2SO_4$, $FeCl_2{\cdot}4H_2O$, NaCl, or $CuSO_4$) were added for enzyme reaction. As results, it was found that $FeCl_2{\cdot}4H_2O$ or $MnSO_4{\cdot}4H_2O$ addition resulted in 17% and 9% improved starch degrading activity, respectively. The recombinant glucoamylase from D. geothermalis might be used for simultaneous saccharification and fermentation (SSF) process at high acidic conditions.