• Proceedings of the Korean Society of Crop Science Conference
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• 2017.06a
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• pp.59-59
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• 2017

Rice (Oryza sativa L.) is the most important crop and its yield must be improved to feed the increasing global population. Recently developed high-yielding varieties with extra-large sink capacity often have a problem in unstable grain-filling. Therefore, understanding limiting factors for improving grain-filling and controlling them are essential for further improvement of rice grain yield. However, since grain-filling rate was determined by complex sink-source balance, the ability of grain-filling was very difficult to evaluate. Source ability for 'grain' was not only determined by the ability of carbon assimilation in leaves, but also that of carbon translocation from leaves to panicles. Sink strength was determined by the complex carbon metabolism from sucrose degradation to starch synthesis. Hence, to evaluate the grain-filling ability and determine its regulatory steps, the whole picture of carbon flow from photosynthesis at leaves to starch synthesis at grains must be revealed in a metabolite level. In this study, the yield and grain growth rate of three high-yielding varieties, which show high sink capacity commonly, were compared. Momiroman showed lower grain filling rate and slower grain growth rate than the other varieties, Hokuriku 193 and Tequing. To clarify the limiting point in the carbon flow of Momiroman, $CO_2$ labeled by stable isotope ($^{13}C$) was fed to three varieties during ripening period. The ratio of $^{13}C$ left in the stem was higher in Momiroman 24 hours after feeding, suggesting inefficient carbon translocation of Momiroman. More interestingly, $^{13}C$ translocation from soluble fraction to insoluble one in the grain seemed to be slower in Momiroman. To get the further insight in a metabolite level, we are now trying the $^{13}C$ labeling metabolome analysis in the developing grains.

• BMB Reports
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• v.45 no.7
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• pp.396-401
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• 2012

Overnutrition is one of the major causes of non-alcoholic fatty liver disease (NAFLD). NAFLD is characterized by an accumulation of lipids (triglycerides) in hepatocytes and is often accompanied by high plasma levels of free fatty acids (FFA). In this study, we compared the energy metabolism in acute steatotic and non-steatotic primary mouse hepatocytes. Acute steatosis was induced by pre-incubation with high concentrations of oleate and palmitate. Labeling experiments were conducted using [$U-^{13}C_5$,$U-^{15}N_2$] glutamine. Metabolite concentrations and mass isotopomer distributions of intracellular metabolites were measured and applied for metabolic flux estimation using transient $^{13}C$ metabolic flux analysis. FFAs were efficiently taken up and almost completely incorporated into triglycerides (TAGs). In spite of high FFA uptake rates and the high synthesis rate of TAGs, central energy metabolism was not significantly changed in acute steatotic cells. Fatty acid ${\beta}$-oxidation does not significantly contribute to the detoxification of FFAs under the applied conditions.

• 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.

• Bulletin of the Korean Chemical Society
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• v.35 no.8
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• pp.2367-2370
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• 2014

"Cat's eye"-shaped $[^{57}Co]CoO@SiO_2$ core-shell nanostructure was prepared by the reverse microemulsion method combined with radioisotope technique to investigate a potential imaging agent for a single photon emission computed tomography (SPECT) in nuclear medicine. The core cobalt oxide nanorods were obtained by thermal decomposition of $Co-(oleate)_2$ precursor from radio isotope Co-57 containing cobalt chloride and sodium oleate. The $SiO_2$ coating on the surface of the core cobalt oxide nanorods was produced by hydrolysis and a condensation reaction of tetraethylorthosilicate (TEOS) in the water phase of the reverse microemulsion system. In vivo test, micro SPECT image was acquired with nude mice after 30 min of intravenous injection of $[^{57}Co]CoO@SiO_2$ core-shell nanostructure.

• Proceedings of the Korean Society of Applied Pharmacology
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• 1995.04a
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• pp.80-80
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• 1995

T4 Endonuclease V (Mw 16,000) acts as a repair enzyme for UV induced pyrimidine dimers in DNA. Many researchers have studied the biochemical characteristics of the enzyme. However the precise action mechanism of T4 endo V has not fully elucidated yet. In our laboratory NMR spectroscopy technique is being used for the structural study of T4 endo V. Because of its low temperature stability and high content of ${\alpha}$-helix, the conventional $^1$H NMR technique was inapplicable. Therefore we utilized stable isotope labeling technique and so far prepared about 10 amino acid specific labeled proteins. The HSQC spectra of amino acid specific labeled proteins will help us to interpret the triple resonance 3D, 4D data which are under processing, We also studied the behaviors of specific amino acid residues whose roles might be critical. When the enzyme labeled by $\^$15/N-Thr was mixed with the substrate oligonucleotide (semispecific -TT- sequence), one crosspeak in its HSQC spectrum was completely desappeared, which means that one of seven Thr residues is in the binding site of the enzyme with DNA, This result is well consistent with previous report that implicated the Thr 2 residue in the activity of the enzyme. Similar studies were carried on the behaviors of Arg and Tyr residues.

• Journal of Ginseng Research
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• v.41 no.4
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• pp.566-571
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• 2017

Background: A number of reports have described the protective effects of ginsenoside Rg1 (Rg1) in Alzheimer's disease (AD). However, the protective mechanisms of Rg1 in AD remain elusive. Methods: To investigate the potential mechanisms of Rg1 in ${\beta}$-amyloid peptide-treated SH-SY5Y cells, a comparative proteomic analysis was performed using stable isotope labeling with amino acids in cell culture combined with nano-LC-MS/MS. Results: We identified a total of 1,149 proteins in three independent experiments. Forty-nine proteins were significantly altered by Rg1 after exposure of the cells to ${\beta}$-amyloid peptides. The protein interaction network analysis showed that these altered proteins were clustered in ribosomal proteins, mitochondria, the actin cytoskeleton, and splicing proteins. Among these proteins, mitochondrial proteins containing HSD17B10, AARS2, TOMM40, VDAC1, COX5A, and NDUFA4 were associated with mitochondrial dysfunction in the pathogenesis of AD. Conclusion: Our results suggest that mitochondrial proteins may be related to the protective mechanisms of Rg1 in AD.

• Bulletin of the Korean Chemical Society
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• v.30 no.11
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• pp.2647-2650
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• 2009

Cold shock proteins (Csps) are a subgroup of the cold-induced proteins on reduction of the growth temperature below the physiological temperature. They preferentially bind to single-stranded nucleic acids to translational regulation via RNA chaperoning. Csp plays important role in cold adaptations for the psychrophilic microorganism. Recently, Cold shock protein from psychrophilic bacteria, Colwellia psychrerythraea (CpCsp) has been identified. Three dimensional structures of a number of Csps from various microorganisms have been solved by NMR spectroscopy or X-ray crystallography, but structures of psychrophilic Csps were not studied yet. Therefore, cloning and purification protocols for further structural study of psychrophilic Csp have been optimized in this study. CpCsp was expressed in E. coli with pET-11a vector system and purified by ion exchange, size exclusion, and reverse phase chromatography. Expression and purification of CpCsp in M9 minimal media was carried out and $^{15}N$-labeled proteins with high purity over 90% was obtained. Further study will be carried out to investigate the tertiary structure and dynamics of CpCsp.

• Journal of Radiopharmaceuticals and Molecular Probes
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• v.2 no.1
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• pp.9-16
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• 2016

Carbon-11 is one of the most sensitive and desirable positron emission tomography radio-isotope, which offers the capacity to be incorporated, through a covalent bond, into biologically active molecules without altering their biological properties. Carbon-11 can be obtained from the cyclotron with two different chemical forms: $[^{11}C]CO_2$ and $[^{11}C]CH_4$. [$^{11}C$]Methyl iodide has been widely used as a highly reactive labelling precursor that can be applied to label carbon-11 with biologically active molecules via alkylation of N-, O-, or S-nucleophiles. A more recent and still challenging labeling method is transition metal mediated $^{11}C$-carbonylation. Advances in organic chemistry, radiochemistry and improved automated techniques greatly encourage researchers to develop more carbon-11 labelled radiotracers for molecular imaging studies. This mini-review will introduce a historical track of carbon-11 chemistry combining with examples and its role in near future.

• Molecules and Cells
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• v.45 no.3
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• pp.158-167
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• 2022

Ubiquitin (Ub) is post-translationally modified by Ub itself or Ub-like proteins, phosphorylation, and acetylation, among others, which elicits a variety of Ub topologies and cellular functions. However, N-terminal (Nt) modifications of Ub remain unknown, except the linear head-to-tail ubiquitylation via Nt-Met. Here, using the yeast Saccharomyces cerevisiae and an Nt-arginylated Ub-specific antibody, we found that the detectable level of Ub undergoes Nt-Met excision, Nt-deamination, and Nt-arginylation. The resulting Nt-arginylated Ub and its conjugated proteins are upregulated in the stationary-growth phase or by oxidative stress. We further proved the existence of Nt-arginylated Ub in vivo and identified Nt-arginylated Ub-protein conjugates using stable isotope labeling by amino acids in cell culture (SILAC)-based tandem mass spectrometry. In silico structural modeling of Nt-arginylated Ub predicted that Nt-Arg flexibly protrudes from the surface of the Ub, thereby most likely providing a docking site for the factors that recognize it. Collectively, these results reveal unprecedented Nt-arginylated Ub and the pathway by which it is produced, which greatly expands the known complexity of the Ub code.

• Parasites, Hosts and Diseases
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• v.55 no.2
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• pp.121-128
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• 2017

Protozoan viruses may influence the function and pathogenicity of the protozoa. Trichomonas vaginalis is a parasitic protozoan that could contain a double stranded RNA (dsRNA) virus, T. vaginalis virus (TVV). However, there are few reports on the properties of the virus. To further determine variations in protein expression of T. vaginalis, we detected 2 strains of T. vaginalis; the virus-infected ($V^+$) and uninfected ($V^-$) isolates to examine differentially expressed proteins upon TVV infection. Using a stable isotope N-terminal labeling strategy (iTRAQ) on soluble fractions to analyze proteomes, we identified 293 proteins, of which 50 were altered in $V^+$ compared with $V^-$ isolates. The results showed that the expression of 29 proteins was increased, and 21 proteins decreased in $V^+$ isolates. These differentially expressed proteins can be classified into 4 categories: ribosomal proteins, metabolic enzymes, heat shock proteins, and putative uncharacterized proteins. Quantitative PCR was used to detect 4 metabolic processes proteins: glycogen phosphorylase, malate dehydrogenase, triosephosphate isomerase, and glucose-6-phosphate isomerase, which were differentially expressed in $V^+$ and $V^-$ isolates. Our findings suggest that mRNA levels of these genes were consistent with protein expression levels. This study was the first which analyzed protein expression variations upon TVV infection. These observations will provide a basis for future studies concerning the possible roles of these proteins in host-parasite interactions.