• 제목/요약/키워드: lrp

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트립토판 돌연변이 루신-반응 조절 단백질의 형광 특성 (Fluorescence Characteristics of a Tryptophan Mutant of Leucine-responsive Regulatory Protein (Lrp))

  • 로버트 포쿠;이의호;이찬용
    • 미생물학회지
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    • 제50권4호
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    • pp.275-280
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    • 2014
  • 루신-반응 조절 단백질(Lrp)은 18.8 kDa의 분자량을 갖는 164개의 아미노산으로 이루어진 글로벌 조절 단백질으로서, 야생형의 단백질(Lrp Wt)에는 아미노산 중 가장 강한 자체 형광을 띠는 트립토판이 존재하지 않는다. Lrp 단백질의 구조변이에 대한 정보를 줄 수 있는 형광분석을 위하여 Lrp Wt과 트립토판이루신-반응 영역에 단지 하나 존재하는 돌연변이 단백질(Lrp R145W)을 분리 정제하였다. Lrp R145W 단백질은 이들 ilvIH 오페론에서 고안된 Lrp 결합 특정 DNA와 아미노산 루신과의 결합 후에 형광이 감소하였으며 acrylamide, urea 등에 의해서도 급격히 쇄광하는 양상을 보였다. 이들 형광 실험 결과는 Lrp의 3차원적 구조 및 배향을 연구에 중요한 정보를 제공하여 줄 수 있을 것이다.

대장균 Leucine-Responsive Regulatory Protein의 정제 및 형광분광학적 특성 분석 (Purification and Fluorometric Analysis of Leucine-Responsive Regulatory Protein from Escherichia coli)

  • 이찬용;김성철;서초희
    • 미생물학회지
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    • 제46권1호
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    • pp.104-108
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    • 2010
  • Leucine-responsive regulatory protein (Lrp)는 대장균 (Escherichia coli)에서 발견된 '글로벌 조절자(global regulator)'로서 Lrp-regulon이 leucine에 의하여 상이한 형태의 조절 양상을 나타낸다. 6XHis-tag 시스템으로 제조한 야생형 Lrp (Lrp Wt)와 돌연변이 Lrp (Lrp R145W) 단백질을 정제하여 그들의 생화학적 성질을 조사하였다. 이들은 gel retardation assay를 통하여 ilv 오페론의 프로모터 영역 consensus 염기서열인 21bp의 이중가닥 DNA와 결합하여 복합체를 형성하는 것을 확인 하였다. 형광성 아미노산인 tryptophan을 지닌 Lrp R145W은 단백질의 농도가 증가함에 따라 형광이 커졌으며, 아미노산 leucine에 의하여 형광성의 변화가 관찰되었다. 즉 1 ${\mu}M$의 Lrp R145W 단백질에 leucine을 첨가하여 결합시키면 약 20 ${\mu}M$까지는 형광이 증가하다가 그 이상의 농도에서는 감소하는 양상을 얻었다. 이들 실험 결과는 leucine과 Lrp의 결합 양상 및 구조변이에 관한 심층연구에 있어서 Lrp의 고유 형광성이 요긴하게 쓰일 수 있음을 시사한다.

저밀도 리포단백질 수용체 관련 단백질 5(LRP5) 유전자 적중 생쥐의 개발 (Development of Low Density Lipoprotein Receptor-Related Protein 5 (LRP5) Gene Targeted Mouse)

  • 박효영;김철민;이상미;정영희;문승주;강만종
    • Reproductive and Developmental Biology
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    • 제29권1호
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    • pp.19-24
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    • 2005
  • 저밀도 리포단백질 수용체 관련 단백질 5(LRP5)는 간과 췌장을 포함하여 많은 조직에서 발현하며 아포리포단백질 E와 결합한다. 이와 같은 LRP5 유전자의 체내 기능을 규명하기 위하여 LRP5 유전자가 결손된 생쥐를 개발하였다. 먼지 LRP5 genomic DNA는 TT2 ES 세포로부터 분리하였으며 LRP5 유전자의 엑손 18에 neo 유전자를 삽입한 vector를 구축하고 TT2 ES 세포에 도입하였다. 178개의 G418 내성을 보인 세포 중 상동유전자 재조합에 의하여 targeting vector가 LRP5 유전자 위치에 삽입된 clone은 3개였다. 키메라 생쥐는 상실배기 수정을 ES 세포와 응집시켜 생산하였으며 생산된 키메라 생쥐는 C57BL/6 생쥐와 교미를 유도하여 heterozygous를 얻었다. 또한 이들 heterozygous간의 교배에 의하여 LRP5 유전자 결손 생쥐를 생산하였다. 이러한 생쥐는 LRP5 유전자의 체내 기능연구에 있어서 모델로 이용될 것으로 생각된다.

The novel gene LRP15 is regulated by DNA methylation and confers increased efficiency of DNA repair of ultraviolet-induced DNA damage

  • Xu, Zhou-Min;Gao, Wei-Ran;Mei, Qi;Chen, Jian;Lu, Jing
    • BMB Reports
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    • 제41권3호
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    • pp.230-235
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    • 2008
  • LRP15 is a novel gene cloned from lymphocytic cells, and its function is still unknown. Bioinformatic data showed that LRP15 might be regulated by DNA methylation and had an important role in DNA repair. In this study, we investigate whether the expression of LRP15 is regulated by DNA methylation, and whether overexpression of LRP15 increases efficiency of DNA repair of UV-induced DNA damage in HeLa cells. The results showed (1) the promoter of LRP15 was hypermethylated in HeLa cells, resulting a silence of its expression. Gene expression was restored by a demethylating agent, 5-aza-2'-deoxycytidine, but not by a histone deacetylase inhibitor, trichostatin A; (2) overexpression of LRP15 inhibited HeLa cell proliferation, and the numbers of cells in the G2/M phase of the cell cycle in cells transfected with LRP15 increased about 10% compared with controls; (3) cyclin B1 level was much lower in cells overexpressing LRP15 than in control cells; and (4) after exposure to UV radiation, the LRP15-positive cells showed shorter comet tails compared with the LRP15-negative cells. From these results we conclude that the expression of LRP15 is controlled by methylation in its promoter in HeLa cells, and LRP15 confers resistance to UV damage and accelerates the DNA repair rate.

교차결합 실험을 통한 루신 대응 조절 단백질의 4차 구조 분석 (Analysis of quaternary structure of leucine-responsive regulatory protein (Lrp) by crosslink experiments)

  • 이의호;로버트 포쿠;로이 트엔 응구엔;이찬용
    • 미생물학회지
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    • 제53권4호
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    • pp.297-303
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    • 2017
  • 루신-대응 조절 단백질(Lrp)은 18.8 kDa의 분자량을 갖는 글로벌 조절 단백질로서 대장균과 같은 장내세균과에서 많은 대사작용 오페론의 기능적 활성도를 조절한다. 단백질의 4차 구조를 규명하기 위한 목적으로 Lrp단백질 코드하는 유전자가 삽입된 재조합 플라스미드 pQE vector를 발현시킨 6 ${\times}$ His-tag Lrp 야생형과 $^3H$로 표지된 Lrp를 분리 정제한 후 cross linker들인 glutaraldehyde, 1,2,3,4-diepoxy-butane (DEB), ethylene glycol bis (succinimidyl succinate) (EGS)으로 cross link 실험을 수행하여 Lrp가 $0.3{\mu}M$ 이하의 낮은 농도에서나 $5{\mu}M$의 높은 농도에서 이량체, 사량체, 육량체, 팔량체로 존재할 수 있음을 확인하였다.

대장균의 Global 조절 단백질인 Lrp (Leucine-responsive Regulatory Protein)의 생화학적 특성 (Biochemical Characteristics of Lrp (Leucine-responsive Regulatory Protein) as a Global Regulator in Escherichia coli)

  • 이찬용;김소영;김류련
    • 미생물학회지
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    • 제42권4호
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    • pp.239-245
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    • 2006
  • Leucine-responsive Regulatory Protein (Lrp)는 global 조절 단백질로서 아미노산의 합성 및 분해 작용과 pilin 생합성을 포함하는 다양한 대사기능을 조절하는데 관여한다. 또한 Lrp가 대장균의 성장 정지기의 유전자 발현에 매우 중요한 역할을 하는 것으로 최근의 여러 실험 결과에서 밝혀지고 있다. 따라서 본 총설에서는 세균들이 영양부족 등의 환경적인 스트레스상황을 어떻게 인지하고 유전자 발현 조절에 그 정보를 반영해 나가는지를 제시해 주는 종은 모텔 시스템으로서 Lrp의 생화학적 특성을 기술하였다.

Activation of the Vibrio vulnificus cadBA Operon by Leucine-Responsive Regulatory Protein is Mediated by CadC

  • Rhee, Jee-Eun;Kim, Kun-Soo;Choi, Sang Ho
    • Journal of Microbiology and Biotechnology
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    • 제18권11호
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    • pp.1755-1761
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    • 2008
  • The present study revealed that Lrp, a leucine-responsive regulatory protein, is involved in the regulation of cadBA transcription through activation of $P_{cadBA}$. The influence of Lrp on $P_{cadBA}$ was mediated by CadC, and thereby, CadC was able to compensate for the lack of Lrp in the activation of $P_{cadBA}$. Western blot analyses and EMSA demonstrated that the cellular level of CadC was not significantly affected by Lrp, and that Lrp exerted its effect by directly binding to $P_{cadBA}$. These combined results suggested that CadC and Lrp function cooperatively to activate the $P_{cadBA}$ rather than sequentially in a regulatory cascade.

Clinical Usefulness of LabChip Real-time PCR using Lab-On-a-Chip Technology for Diagnosing Malaria

  • Kim, Jeeyong;Lim, Da Hye;Mihn, Do-CiC;Nam, Jeonghun;Jang, Woong Sik;Lim, Chae Seung
    • Parasites, Hosts and Diseases
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    • 제59권1호
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    • pp.77-82
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    • 2021
  • As malaria remains a major health problem worldwide, various diagnostic tests have been developed, including microscopy-based and rapid diagnostic tests. LabChip real-time PCR (LRP) is a small and portable device used to diagnose malaria using lab-on-a-chip technology. This study aimed to evaluate the diagnostic performance of LRP for detecting malaria parasites. Two hundred thirteen patients and 150 healthy individuals were enrolled from May 2009 to October 2015. A diagnostic detectability of LRP for malaria parasites was compared to that of conventional RT-PCR. Sensitivity of LRP for Plasmodium vivax, P. falciparum, P. malariae, and P. ovale was 95.5%, 96.0%, 100%, and 100%, respectively. Specificity of LRP for P. vivax, P. falciparum, P. malariae, and P. ovale was 100%, 99.3%, 100%, and 100%, respectively. Cohen's Kappa coefficients between LRP and CFX96 for detecting P. vivax, P. falciparum, P. malariae, and P. ovale were 0.96, 0.98, 1.00, and 1.00, respectively. Significant difference was not observed between the results of LRP and conventional RT-PCR and microscopic examination. A time required to amplify DNAs using LRP and conventional RT-PCR was 27 min and 86 min, respectively. LRP amplified DNAs 2 times more fast than conventional RT-PCR due to the faster heat transfer. Therefore, LRP could be employed as a useful tool for detecting malaria parasites in clinical laboratories.

Gene Targeting of Low Density Lipoprotein(LDL) Receptor Related Protein 5(LRP5) Involved in the Wnt Signaling Pathway

  • Jeong, Young-Hee;Kim, Suck-Ho;Kim, Dong-Ho;Moon, Seung-Ju;Tokuo Yamamoto;Kang, Man-Jong
    • 한국동물번식학회:학술대회논문집
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    • 한국동물번식학회 2002년도 춘계학술발표대회 발표논문초록집
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    • pp.82-82
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    • 2002
  • The Wnt signaling pathway plays pivotal roles in embryonic development and oncogenesis through various signaling molecules inculding Frizzled receptor, recently characterized LRP5/6 and Dickkopf protein. Although Wnt signaling has been characterized in both developmental and oncogenic processes, little is known about its function in the normal adult. The ability of LRP5 to bind apolipoprotein E(apoE) and the abundant expression of LRP5 transcripts in hepatocytes, raise the possibility that LRP5 plays a role in the hepatic clearance of ApoE-containing chylomicron remonants, a major plasma lipoprotein carrying diet-derived cholesterol. (omitted)

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High Expression of Lung Resistance Protein mRNA at Diagnosis Predicts Poor Early Response to Induction Chemotherapy in Childhood Acute Lymphoblastic Leukemia

  • Bhatia, Prateek;Masih, Shet;Varma, Neelam;Bansal, Deepak;Trehan, Amita
    • Asian Pacific Journal of Cancer Prevention
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    • 제16권15호
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    • pp.6663-6668
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    • 2015
  • Background: Treatment failure in leukemia is due to either pharmacokinetic resistance or cell resistance to drugs. Materials and Methods: Gene expression of multiple drug resistance protein (MDR-1), multidrug resistance-related protein (MRP) and low resistance protein (LRP) was assessed in 45 pediatric ALL cases and 7 healthy controls by real time PCR. The expression was scored as negative, weak, moderate and strong. Results: The male female ratio of cases was 2.75:1 and the mean age was 5.2 years. Some 26/45 (58%) were in standard risk, 17/45(38%) intermediate and 2/45 (4%) in high risk categorie, 42/45 (93%) being B-ALL and recurrent translocations being noted in 5/45 (11.0%). Rapid early response (RER) at day 14 was seen in 37/45 (82.3%) and slow early response (SER) in 8/45 (17.7%) cases. Positive expression of MDR-1, LRP and MRP was noted in 14/45 (31%), 15/45 (33%) and 27/45 (60%) cases and strong expression in 3/14 (21%), 11/27 (40.7%) and 8/15 (53.3%) cases respectively. Dual or more gene positivity was noted in 17/45 (38%) cases. 46.5 % (7/15) of LRP positive cases at day 14 were in RER as compared to 100% (30/30) of LRP negative cases (p<0.05). All 8 (100%) LRP positive cases in SER had strong LRP expression (p=<0.05). Moreover, only 53.3% of LRP positive cases were in haematological remission at day 30 as compared to 100% of LRP negative cases (p=<0.05). Conclusions: Our study indicated that increased LRP expression at diagnosis in pediatric ALL predicts poor response to early treatment and hence can be used as a prognostic marker. However, larger prospective studies with longer follow up are needed, to understand the clinical relevance of drug resistance proteins.