• Korean Journal of Microbiology
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• v.37 no.2
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• pp.109-113
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• 2001

As the complete genomic sequences accumulate, the use of global techniques became possible. DNA microarray is a powerful technology for measuring global mRNA levels. This method, however, does not provide information on post-translational modifications of proteins. In addition, mRNA levels do not strictly correlate with protein concentrations, especially for lower-abundance proteins. Therefore, studies at the level of transcription are not sufficient to understand cellular activity. Proteomic techniques to analyze protein expression and function at the large-scale have been developed and used. This review introduces a simple explanation for proteomic analysis and examples of how proteomics is applied in cell biology.

• Korean Journal of Environmental Agriculture
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• v.25 no.4
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• pp.371-381
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• 2006

New proteomic techniques have been pioneered extensively in recent years, enabling the high-throughput and systematic analyses of cellular proteins in combination with bioinformatic tools. Furthermore, the development of such novel proteomic techniques facilitates the elucidation of the functions of proteins under stress or disease conditions, resulting in the discovery of biomarkers for responses to environmental stimuli. The ultimate objective of proteomics is targeted toward the entire proteome of life, subcellular localization biochemical activities, and the regulation thereof. Comprehensive analysis strategies of proteomics can be classified into three categories: (i) protein separation via 2-dimensional gel electrophoresis (2-DE) or liquid chromatography (LC), (ii) protein identification via either Edman sequencing or mass spectrometry (MS), and (iii) proteome quantitation. Currently, MS-based proteomics techniques have shifted from qualitative proteome analysis via 2-DE or 2D-LC coupled with off-line matrix assisted laser desorption ionization (MALDI) and on-line electrospray ionization (ESI) MS, respectively, toward quantitative proteome analysis. In vitro quantitative proteomic techniques include differential gel electrophoresis with fluorescence dyes. protein-labeling tagging with isotope-coded affinity tags, and peptide-labeling tagging with isobaric tags for relative and absolute quantitation. In addition, stable isotope-labeled amino acids can be in vivo labeled into live culture cells via metabolic incorporation. MS-based proteomics techniques extend to the detection of the phosphopeptide mapping of biologically crucial proteins, which ale associated with post-translational modification. These complementary proteomic techniques contribute to our current understanding of the manner in which life responds to differing environment.

• Journal of Plant Biotechnology
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• v.37 no.2
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• pp.196-204
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• 2010

With the completion of genome sequencing of several organisms, attention has been focused to determine the function and functional network of proteins by proteome analysis. The recent techniques of proteomics have been advanced quickly so that the high-throughput and systematic analyses of cellular proteins are enabled in combination with bioinformatics tools. Furthermore, the development of proteomic techniques helps to elucidate the functions of proteins under stress or diseased condition, resulting in the discovery of biomarkers responsible for the biological stimuli. Ultimate goal of proteomics orients toward the entire proteome of life, subcellular localization, biochemical activities, and their regulation. Comprehensive analysis strategies of proteomics can be classified as three categories: (i) protein separation by 2-dimensional gel electrophoresis (2-DE) or liquid chromatography (LC), (ii) protein identification by either Edman sequencing or mass spectrometry (MS), and (iii) quanitation of proteome. Currently MS-based proteomics turns shiftly from qualitative proteome analysis by 2-DE or 2D-LC coupled with off-line matrix assisted laser desorption ionization (MALDI) and on-line electrospray ionization (ESI) MS, respectively, to quantitative proteome analysis. Some new techniques which include top-down mass spectrometry and tandem affinity purification have emerged. The in vitro quantitative proteomic techniques include differential gel electrophoresis with fluorescence dyes, protein-labeling tagging with isotope-coded affinity tag, and peptide-labeling tagging with isobaric tags for relative and absolute quantitation. In addition, stable isotope labeled amino acid can be in vivo labeled into live culture cells through metabolic incorporation. MS-based proteomics extends to detect the phosphopeptide mapping of biologically crucial protein known as one of post-translational modification. These complementary proteomic techniques contribute to not only the understanding of basic biological function but also the application to the applied sciences for industry.

• Analytical Science and Technology
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• v.24 no.6
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• pp.407-413
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• 2011

Over the last decade sophisticated and powerful microcapillary HPLC for proteomic analysis have been developed increasingly and interfaced with high resolution tandem mass spectrometers. Separation prior to mass spectrometric (MS) analysis removes impurities, and concentrates analytes in the narrow elution peaks, resulting in increased sensitivity of MS analysis. This review will focus on the recent advances of on-line highperformance separation techniques based on microfluidic chips for complex proteomic analysis.

• Molecular & Cellular Toxicology
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• v.1 no.1
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• pp.7-12
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• 2005

Proteomics has recently received intense scientific interest after the completion of the Human Genome Project, because this genome-based high technology allows to search new drug targets or diagnostic markers. Many proteome projects including Human plasma proteome projects (HPPP), Human liver proteome projects (HLPP), Human brain proteome projects (HBPP), and Mouse and Rat Proteome Project (MRPP) have been carried out and proteomic analytical techniques have been developed in second dimensional electrophoresis (2-DE) and LC/MS system. This powerful method has been applied in toxicology producing a new term "Toxicoproteomics". In this review, recent proteome projects, proteomic technologies, and toxicoproteomics will be discussed.

• BMB Reports
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• v.47 no.3
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• pp.149-157
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• 2014

The recent dramatic improvements in high-resolution mass spectrometry (MS) have revolutionized the speed and scope of proteomic studies. Conventional MS-based proteomics methodologies allow global protein profiling based on expression levels. Although these techniques are promising, there are numerous biological activities yet to be unveiled, such as the dynamic regulation of enzyme activity. Chemical proteomics is an emerging field that extends these types proteomic profiling. In particular, activity-based protein profiling (ABPP) utilizes small-molecule probes to monitor enzyme activity directly in living intact subjects. In this mini-review, we summarize the unique roles of smallmolecule probes in proteomics studies and highlight some recent examples in which this principle has been applied.

• Mass Spectrometry Letters
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• v.4 no.1
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• pp.1-9
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• 2013

Fungal biotechnology has been well established in food and healthcare sector, and now being explored for lignocellulosic biorefinery due to their great potential to produce a wide array of extracellular enzymes for nutrient recycling. Due to global warming, environmental pollution, green house gases emission and depleting fossil fuel, fungal enzymes for lignocellulosic biomass refinery become a major focus for utilizing renewal bioresources. Proteomic technologies tender better biological understanding and exposition of cellular mechanism of cell or microbes under particular physiological condition and are very useful in characterizing fungal secretome. Hence, in addition to traditional colorimetric enzyme assay, mass-spectrometry-based quantification methods for profiling lignocellulolytic enzymes have gained increasing popularity over the past five years. Majority of these methods include two dimensional gel electrophoresis coupled to mass spectrometry, differential stable isotope labeling and label free quantitation. Therefore, in this review, we reviewed more commonly used different proteomic techniques for profiling fungal secretome with a major focus on two dimensional gel electrophoresis, liquid chromatography-based quantitative mass spectrometry for global protein identification and quantification. We also discussed weaknesses and strengths of these methodologies for comprehensive identification and quantification of extracellular proteome.

• Journal of Microbiology and Biotechnology
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• v.17 no.2
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• pp.207-217
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• 2007

The Saccharomyces0 cerevisiae KNU5377 strain, which was isolated from spoilage in nature, has the ability to convert biomass to alcohol at high temperatures and it can resist against various stresses [18, 19]. In order to understand the defense mechanisms of the KNU5377 strain under menadione (MD) as oxidative stress, we used several techniques for study: peptide mass fingerprinting (PMF) by matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry (MS) followed by two-dimensional (2D) gel electrophoresis, liquid chromatography electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS), and surface-enhanced laser desorption ionization-time of flight (SELDI-TOF) technology. Among the 35 proteins identified by MALDI-TOF MS, 19 proteins including Sod1p, Sod2p, Tsa1p, and Ahp1p were induced under stress condition, while 16 proteins were augmented under normal condition. In particular, five proteins, Sod1p, Sod2p, Ahp1p, Rib3p, Yaf9p, and Mnt1p, were induced in only stressed cells. By LC-ESI-MS/MS analysis, 37 proteins were identified in normal cells and 49 proteins were confirmed in the stressed cells. Among the identified proteins, 32 proteins were found in both cells. Five proteins including Yel047cp and Met6p were only upregulated in the normal cells, whereas 17 proteins including Abp1P and Sam1p were elevated in the stressed cells. It was interesting that highly hypothetical proteins such as Ynl281wp, Ygr279cp, Ypl273wp, Ykl133cp, and Ykr074wp were only expressed in the stressed cells. SELDI-TOF analysis using the SAX2 and WCX2 chips showed that highly multiple-specific protein patterns were reproducibly detected in ranges from 2.9 to 27.0 kDa both under normal and stress conditions. Therefore, induction of antioxidant proteins, hypothetical proteins, and low molecular weight proteins were revealed by different proteomic techniques. These results suggest that comparative analyses using proteomics might contribute to elucidate the defense mechanisms of KNU5377 under MD stress.

• Food Science of Animal Resources
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• v.33 no.2
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• pp.281-286
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• 2013

Salmonella Gallinarum (SG) is known as an important pathogen that causes fowl typhoid in chickens. To investigate SG outer-membrane proteins (OMPs) as a vaccine candidate, we used proteomic mapping and database analysis techniques with extracted OMPs. Also, extracted OMPs were evaluated in several aspects to their safety, immune response in their host and protective effects. Our research has established a proteomic map and database of immunogenic SG-OMPs used as inactive vaccine against salmonellosis in chickens. A total of 22 spots were detected by 2-dimensional gel electrophoresis and immunogenic protein analysis. Eight spots were identified by Matrix-Assisted Laser Desorption/Ionization-Time of Flight-Mass spectrometry (MALDI-TOF-MS) and peptide mass fingerprinting (PMF) and categorized into four different types of proteins. Among these proteins, OmpA is considered to be an immunogenic protein and involved in the hosts' immune system. To estimate the minimum safety dose in chickens, 35 brown layers were immunized with various concentrations of OMPs, respectively. Consequently, all chickens immunized with more than a $50{\mu}g$ dose were protected against challenges. Moreover, intramuscular administration of OMPs to chickens was more effective compared to subcutaneous administration. These results suggest that the adjuvanted SG-OMP vaccine not only induces both the humoral and cellular immune response in the host but also highly protects the hosts' exposed to virulent SG with $50{\mu}g$ OMPs extracted by our method.

• Journal of Microbiology and Biotechnology
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• v.11 no.6
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• pp.1111-1114
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• 2001

An extracellular fibrinolytic-enzyme-producing bacterium was isolated from Doen-Jang, a Korean traditional fermented flood, and identified as Bacillus sp. DJ based on its morphology and cellular fatty acid composition. The total extracellular fibrinase (EF) from Bacillus sp. DJ was analyzed using three fibrin zymographic techniques, SDS-fibrin zymography (SDS-FZ), isoelectrofocucing-fibrin zymographs(IEF-FZ), and a two-dimensional SDS-fibrin zymographic analysis (2D SDS-FZ). As a result, the EP map of Bacillus sp. DJ was established. The results suggest that the 2D SDS-FZ method will be a useful tool for the proteomic approach for many other bacterial pretenses.