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채영기,Woo-Young Kang,Seong Hwan Kim,Jong Eun Joo,Joon Kil Han,Boo Whan Hong 대한화학회 2010 Bulletin of the Korean Chemical Society Vol.31 No.2
Metabolites of colorectal cancer tissues from 12 patients were analyzed and compared with those of the normal tissues by two-dimensional NMR spectroscopy. NMR data were analyzed with the help of the metabolome database and the statistics software. Cancerous tissues showed significantly altered metabolic profiles as compared to the normal tissues. Among such metabolites, the concentrations of taurine, glutamate, choline were notably increased in the cancerous tissues of most patients, and those of glucose, malate, and glycerol were decreased. Changes in individual metabolites varied significantly from patient to patient, but the combination of such changes could be used to distinguish cancerous tissues from normal ones, which could be done by PCA analysis. The traditional chemometric analysis was also performed using AMIX software. By comparing those two results, the analysis via 1H-13C HSQC spectra proved to be more robust and effective in assessing and classifying global metabolic profiles of the colorectal tissues.
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채영기,Seol Hyun Kim,James E. Ellinger,John L. Markley 대한화학회 2013 Bulletin of the Korean Chemical Society Vol.34 No.12
Saccharomyces cerevisiae, which is a common species of yeast, is by far the most extensively studied model of a eukaryote because although it is one of the simplest eukaryotes, its basic cellular processes resemble those of higher organisms. In addition, yeast is a commercially valuable organism for ethanol production. Since the yeast data can be extrapolated to the important aspects of higher organisms, many researchers have studied yeast metabolism under various conditions. In this report, we analyzed and compared metabolites of Saccharomyces cerevisiae under salt and pH stresses of various strengths by using two-dimensional NMR spectroscopy. A total of 31 metabolites were identified for most of the samples. The levels of many identified metabolites showed gradual or drastic increases or decreases depending on the severity of the stresses involved. The statistical analysis produced a holistic outline: pH stresses were clustered together, but salt stresses were spread out depending on the severity. This work could provide a link between the metabolite profiles and mRNA or protein profiles under representative and well studied stress conditions.
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채영기,Seol Hyun Kim,James J. Ellinger,John L. Markley 대한화학회 2012 Bulletin of the Korean Chemical Society Vol.33 No.12
The recombinant expression of proteins has been the method of choice to meet the demands from proteomics and structural genomics studies. Despite its successful production of many heterologous proteins, Escherichia coli failed to produce many other proteins in their native forms. This may be related to the fact that the stresses resulting from the overproduction interfere with cellular processes. To better understand the physiological change during the overproduction phase, we profiled the metabolites along the time course of the recombinant protein expression. We identified 32 metabolites collected from different time points in the protein production phase. The stress induced by protein production can be characterized by (A) the increased usage of aspartic acid, choline, glycerol, and N-acetyllysine; and (B) the accumulation of adenosine, alanine, oxidized glutathione, glycine, N-acetylputrescine, and uracil. We envision that this work can be used to create a strategy for the production of usable proteins in large quantities.
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채영기,김설현 대한화학회 2015 Bulletin of the Korean Chemical Society Vol.36 No.1
Escherichia coli has been widely used as a host to produce recombinant proteins. However, because of its prokaryotic nature, it often fails to produce eukaryotic proteins in their native or correctly folded forms. To devise a systematic way to find such a condition that produces a large amount of correctly folded proteins, we chose to perturb the composition of intracellular metabolites by imposing different stresses such as the addition of sorbitol, NaCl, pH, and ethanol, and to monitor which metabolites showed the largest differences in the case of successful protein production. We profiled the metabolites under stresses by using two-dimensional NMR spectroscopy, and identified 39 metabolites after the protein production phase. We found that mild NaCl stress increased the yield of the target protein. Mildly high pH stress also helped in producing more of the target protein, and there is likely a connection between the isoelectric point of the target protein and the pH of medium for a higher yield. Exogenous sorbitol appeared to have been used to produce an osmoprotectant (trehalose) or direct energy source (glucose); thus it is advised that its use should not include isotopic labeling. Mild ethanol stress did not cause any significant change either in the metabolite profile or in the target protein production. We hope this work might shed light on how to improve production of the target protein by adjusting the external environment.
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채영기,김설현 대한화학회 2016 Bulletin of the Korean Chemical Society Vol.37 No.10
Rice is one of the most important crops that feed almost half of the world's population. With the increasing concern of consumers on the integrity of the product, efforts have been made to develop analytic techniques to discriminate rice products according to their origins or cultivars, but those efforts were mostly based on elemental analysis. We postulated that such discrimination would be possible with the global metabolite profiles. Nineteen metabolites of three different rice cultivars from four different geographical origins were identified from the extracts and compared with one another by the two-dimensional nuclear magnetic resonance (NMR) spectroscopy. NMR data were analyzed with the help of the metabolome database and the statistics software. The different rice samples were successfully separated in the principal component space, showing that the global metabolite profiles can be used to discriminate geographic origins. Our results show that the metabolite analysis via1H–13C heteronuclear single quantum coherence spectra combined with the statistical method can be applied to discriminate the geographic origins or cultivars of rice samples, thus can provide a means to inspect and pick up fraudulent labeling or adulteration.
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채영기,강창모 대한화학회 2020 Bulletin of the Korean Chemical Society Vol.41 No.9
Humans are continuously exposed to radiation, originating from natural backgrounds or artificial sources. We focused on the X-ray irradiations to which people are exposed during routine medical checkups. We probed the cellular metabolic responses by feeding human umbilical vein endothelial cells with three different 13C-labeled molecules, which was followed by irradiation with three different doses of X-rays, which was followed by four different incubation periods. We monitored the downstream metabolites via two-dimensional NMR spectroscopy to figure out what input was best suited for our purpose. We found that even though the glucose provided the highest number of metabolized products, the glutamine, which was the lowest, provided as much information as the glucose. Second, the glucose and the glutamine were suitable for the purpose of probing the recovery process, but pyruvate was not suitable. Lastly, 48?h seemed to be needed for the cells to recover from the irradiation shock.
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채영기,김설현,엄윤진 대한화학회 2019 Bulletin of the Korean Chemical Society Vol.40 No.7
In producing large amounts of heterologous proteins, researchers most often use Escherichia coli as a host thanks to its extensively studied genetics, simple growth procedure, and low cost. However, the desired protein is often produced only in a form of inclusion bodies. Researchers have tried to devise a way to circumvent such a problem, and the ones using fusion partners seem to be the most successful. Based on our previous observation that the host metabolome was related to the outcomes of protein expression patterns, we proceeded to perturb the metabolome by applying a salt stress to see if we could shake up metabolite compositions to make them better suited for soluble expression of the target protein. We examined a subset of the metabolites which had been partially labeled with input 13C-glucose. We tested 11 genes under 4 different NaCl concentrations, and identified 18 metabolites using the heteronuclear single quantum coherence NMR experiment. Most of the proteins kept their expression profiles unchanged, but two proteins were converted from inclusion bodies to a soluble form with increased NaCl concentration. Through the statistical analysis, we could identify a region where the soluble protein production was favored in the metabolite space. We hope that this work would provide an alternative strategy to produce the recombinant proteins in their soluble or native forms, not only in E. coli but also in other hosts.
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