• Title/Summary/Keyword: Dimensional Gel Electrophoresis

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The Fate of Strain-Specific Protein in xD Strain of Amoeba proteus (Amoeba proteus xD Strain의 변이주 특이성 단백질의 운영)

  • 안태인
    • The Korean Journal of Zoology
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    • v.26 no.3
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    • pp.181-192
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    • 1983
  • Cytosol protein patterns of two strains of A. proteus, tD and xD strain, were compared by two dimensional gel electrophoresis. Among the 200 major polypeptides that could be stained by silver stain method, tD strain contained a cell specific protein whose molecular weight was 45,000 dalton, pI 5.9. On the other hand, the cytosol and the symbiotic vesicles of xD strain contained a symbiosis specific protein (M.W. 29,000; pI 5.5). The fate of the symbiosis specific protein depended on the presence of symbiotic bacteria in the experiment of high temperature effect and of experimental infection. The significance of these results is discussed in relation to their function in organismic association on the basis of the previous findings.

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Tissue proteomics for cancer biomarker development - Laser microdissection and 2D-DIGE -

  • Kondo, Tadashi
    • BMB Reports
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    • v.41 no.9
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    • pp.626-634
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    • 2008
  • Novel cancer biomarkers are required to achieve early diagnosis and optimized therapy for individual patients. Cancer is a disease of the genome, and tumor tissues are a rich source of cancer biomarkers as they contain the functional translation of the genome, namely the proteome. Investigation of the tumor tissue proteome allows the identification of proteomic signatures corresponding to clinico-pathological parameters, and individual proteins in such signatures will be good biomarker candidates. Tumor tissues are also a rich source for plasma biomarkers, because proteins released from tumor tissues may be more cancer specific than those from non-tumor cells. Two-dimensional difference gel electrophoresis (2D-DIGE) with novel ultra high sensitive fluorescent dyes (CyDye DIGE Fluor satulation dye) enables the efficient protein expression profiling of laser-microdissected tissue samples. The combined use of laser microdissection allows accurate proteomic profiling of specific cells in tumor tissues. To develop clinical applications using the identified biomarkers, collaboration between research scientists, clinicians and diagnostic companies is essential, particularly in the early phases of the biomarker development projects. The proteomics modalities currently available have the potential to lead to the development of clinical applications, and channeling the wealth of produced information towards concrete and specific clinical purposes is urgent.

Metagenomic and Proteomic Analyses of a Mangrove Microbial Community Following Green Macroalgae Enteromorpha prolifera Degradation

  • Wu, Yijing;Zhao, Chao;Xiao, Zheng;Lin, Hetong;Ruan, Lingwei;Liu, Bin
    • Journal of Microbiology and Biotechnology
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    • v.26 no.12
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    • pp.2127-2137
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    • 2016
  • A mangrove microbial community was analyzed at the gene and protein levels using metagenomic and proteomic methods with the green macroalgae Enteromorpha prolifera as the substrate. Total DNA was sequenced on the Illumina HiSeq 2000 PE-100 platform. Two-dimensional gel electrophoresis in combination with liquid chromatography tandem mass spectrometry was used for proteomic analysis. The metagenomic data revealed that the orders Pseudomonadales, Rhizobiales, and Sphingomonadales were the most prevalent in the mangrove microbial community. By monitoring changes at the functional level, proteomic analyses detected ATP synthase and transporter proteins, which were expressed mainly by members of the phyla Proteobacteria and Bacteroidetes. Members of the phylum Proteobacteria expressed a high number of sugar transporters and demonstrated specialized and efficient digestion of various glycans. A few glycoside hydrolases were detected in members of the phylum Firmicutes, which appeared to be the main cellulose-degrading bacteria. This is the first report of multiple "omics" analysis of E. prolifera degradation. These results support the fact that key enzymes of glycoside hydrolase family were expressed in large quantities, indicating the high metabolic activity of the community.

Comprehensive Analysis of Proteomic Differences between Escherichia coli K-12 and B Strains Using Multiplexed Isobaric Tandem Mass Tag (TMT) Labeling

  • Han, Mee-Jung
    • Journal of Microbiology and Biotechnology
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    • v.27 no.11
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    • pp.2028-2036
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    • 2017
  • The Escherichia coli K-12 and B strains are among the most frequently used bacterial hosts for scientific research and biotechnological applications. However, omics analyses have revealed that E. coli K-12 and B exhibit notably different genotypic and phenotypic attributes, even though they were derived from the same ancestor. In a previous study, we identified a limited number of proteins from the two strains using two-dimensional gel electrophoresis and tandem mass spectrometry (MS/MS). In this study, an in-depth analysis of the physiological behavior of the E. coli K-12 and B strains at the proteomic level was performed using six-plex isobaric tandem mass tag-based quantitative MS. Additionally, the best lysis buffer for increasing the efficiency of protein extraction was selected from three tested buffers prior to the quantitative proteomic analysis. This study identifies the largest number of proteins in the two E. coli strains reported to date and is the first to show the dynamics of these proteins. Notable differences in proteins associated with key cellular properties, including some metabolic pathways, the biosynthesis and degradation of amino acids, membrane integrity, cellular tolerance, and motility, were found between the two representative strains. Compared with previous studies, these proteomic results provide a more holistic view of the overall state of E. coli cells based on a single proteomic study and reveal significant insights into why the two strains show distinct phenotypes. Additionally, the resulting data provide in-depth information that will help fine-tune processes in the future.

Effect of Low Doses of Genistein and Equol on Protein Expression Profile in MCF-7 Cells

  • Kim, Jang-Hoon;Lim, Hyun-Ae;Lee, Jeong-Soon;Sung, Mi-Kyung;Kim, Young-Kyoon;Yu, Ri-Na;Kim, Jong-Sang
    • Food Science and Biotechnology
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    • v.14 no.6
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    • pp.854-859
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    • 2005
  • Although action modes of equol and genistein have been extensively studied, their precise roles in tumor cells remain elusive. To address possible effects of these compounds on protein expression in mammary tumor cells, proteins modulated in MCF-7 mammary tumor cells when incubated in absence and presence of 10 uM equol or genistein were identified through 2-dimensional gel electrophoresis, MALDI-TOF MS/MS, and NCBInr database search using Mascot software. Most proteins differentially expressed in MCF-7 cells after treatment with 10 uM genistein or equol were identified as being the same. Exposure to both compounds caused decreased cellular expression of RNA-binding protein regulatory subunit and oncogene DJ1 tubulin beta-1 chain, and increased expression of heterogeneous ribonucleoproteins F and L, KH-type splicing regulatory protein, and translation elongation factor EF-Tu precursor. Genistein and equol at dose used in this study showed common action mechanism.

Induction of Drought Stress Resistance by Multi-Functional PGPR Bacillus licheniformis K11 in Pepper

  • Lim, Jong-Hui;Kim, Sang-Dal
    • The Plant Pathology Journal
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    • v.29 no.2
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    • pp.201-208
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    • 2013
  • Drought stress is one of the major yield affecting factor for pepper plant. The effects of PGPRs were analyzed in relation with drought resistance. The PGPRs inoculated pepper plants tolerate the drought stress and survived as compared to non-inoculated pepper plants that died after 15 days of drought stress. Variations in protein and RNA accumulation patterns of inoculated and non-inoculated pepper plants subjected to drought conditions for 10 days were confirmed by two dimensional polyacrylamide gel electrophoresis (2D-PAGE) and differential display PCR (DD-PCR), respectively. A total of six differentially expressed stress proteins were identified in the treated pepper plants by 2D-PAGE. Among the stress proteins, specific genes of Cadhn, VA, sHSP and CaPR-10 showed more than a 1.5-fold expressed in amount in B. licheniformis K11-treated drought pepper compared to untreated drought pepper. The changes in proteins and gene expression patterns were attributed to the B. licheniformis K11. Accordingly, auxin and ACC deaminase producing PGPR B. licheniformis K11 could reduce drought stress in drought affected regions without the need for overusing agrochemicals and chemical fertilizer. These results will contribute to the development of a microbial agent for organic farming by PGPR.

Proteomic Approach to the Cytotoxicity of 5-FU(Fluorouracil) in Colon Cancer Cells (대장암 세포에서 5-FU(Fluorouracil)의 세포독성과 관련된 단백체 분석)

  • Lee, Seo-Young;Song, Jin-Su;Roh, Si-Hun;Kim, Geun-Tae;Hong, Soon-Sun;Kim, Hie-Joon;Kwon, Sung-Won;Park, Jeong-Hill
    • YAKHAK HOEJI
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    • v.53 no.3
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    • pp.145-150
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    • 2009
  • We evaluated cytotoxic effect based on the MTT assay and identified altered proteins in 5-FU(fluorouracil) treated HT29 cells using two-dimensional gel electrophoresis and MALDI-TOF/TOF-MS. As proteins inducing apoptosis, siah binding protein 1 and p47 protein isoform a were up-regulated and tumor protein translationally-controlled 1 was down-regulated by 5-FU treatment. And mannose 6 phosphate receptor binding protein 1 controls DNA mismatch repair system was increased. We suggest 5-FU promotes a cytotoxicity under the action of these proteins in colon cancer cells.

Ginsenoside-Rp1-induced apolipoprotein A-1 expression in the LoVo human colon cancer cell line

  • Kim, Mi-Yeon;Yoo, Byong Chul;Cho, Jae Youl
    • Journal of Ginseng Research
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    • v.38 no.4
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    • pp.251-255
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    • 2014
  • Background: Ginsenoside Rp1 (G-Rp1) is a novel ginsenoside derived from ginsenoside Rk1. This compound was reported to have anticancer, anti-platelet, and anti-inflammatory activities. In this study, we examined the molecular target of the antiproliferative and proapoptotic activities of G-Rp1. Methods: To examine the effects of G-Rp1, cell proliferation assays, propidium iodine staining, proteomic analysis by two-dimensional gel electrophoresis, immunoblotting analysis, and a knockdown strategy were used. Results: G-Rp1 dose-dependently suppressed the proliferation of colorectal cancer LoVo cells and increased their apoptosis. G-Rp1 markedly upregulated the protein level of apolipoprotein (Apo)-A1 in LoVo, SNU-407, DLD-1, SNU-638, AGS, KPL-4, and SK-BR-3 cells. The knockdown of Apo-A1 by its small-interfering RNA increased the levels of cleaved poly(ADP-ribose) polymerase and p53 and diminished the proliferation of LoVo cells. Conclusion: These results suggest that G-Rp1 may act as an anticancer agent by strongly inhibiting cell proliferation and enhancing apoptosis through upregulation of Apo-A1.

Differential Effects of Two Period Genes on the Physiology and Proteomic Profiles of Mouse Anterior Tibialis Muscles

  • Bae, Kiho;Lee, Kisoo;Seo, Younguk;Lee, Haesang;Kim, Dongyong;Choi, Inho
    • Molecules and Cells
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    • v.22 no.3
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    • pp.275-284
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    • 2006
  • The molecular components that generate and maintain circadian rhythms of physiology and behavior in mammals are present both in the brain (suprachiasmatic nucleus; SCN) and in peripheral tissues. Examination of mice with targeted disruptions of either mPer1 or mPer2 has shown that these two genes have key roles in the SCN circadian clock. Here we show that loss of the clock gene mPer2 affects forced locomotor performance in mice without altering muscle contractility. A proteomic analysis revealed that the anterior tibialis muscles of the mPer2 knockout mice had higher levels of glycolytic enzymes such as triose phosphate isomerase and enolase than those of either the wild type or mPer1 knockout mice. In addition, the level of expression of HSP90 in the mPer2 mutant mice was also significantly higher than in wildtype mice. These results suggest that the reduced locomotor endurance of the mPer2 knockout mice reflects a greater dependence on anaerobic metabolism under stress conditions, and that the two canonical clock genes, mPer1 and mPer2, play distinct roles in the physiology of skeletal muscle.

The Physiological Role of CPR1 in Saccharomyces cerevisiae KNU5377 against Menadione Stress by Proteomics

  • Kim, Il-Sup;Yun, Hae-Sun;Kwak, Sun-Hye;Jin, Ing-Nyol
    • Journal of Microbiology
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    • v.45 no.4
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    • pp.326-332
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    • 2007
  • In order to understand the functional role of CPRl in Saccharomyces cerevisiae KNU5377 with regard to its multi-tolerance characteristics against high temperatures, inorganic acids, and oxidative stress conditions, whole cellular proteins were analyzed via liquid chromatography electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS). This procedure was followed by two-dimensional (2-D) gel electrophoresis. Under menadione stress conditions, the 23 upregulated proteins were clearly identified only in the wild- type strain of KNU5377. Among the proteins, Sodl1p Tsa1p, Ahp1, Cpr1p, Cpr3, Ssb2p, and Hsp12p were identified as components of antioxidant systems or protein-folding related systems. The CPR1 protein could not be completely detected in the $cpr1{\Delta}$ mutant of KNU5377 and the other upregulated proteins in the wild-type strain evidenced a clear correlation with the results of immunoblot analysis. Moreover, a reduction in growth patterns (about 50%) could be observed in the $cpr1{\Delta}$ mutant, as compared with that of the wild-type strain under mild MD stress conditions. These results indicate that the upregulation of CPR1 may contribute to tolerance against MD as an inducer of oxidative stress.