• Molecules and Cells
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• v.38 no.5
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• pp.381-389
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• 2015

Autophagy is a lysosome-dependent intracellular degradation process that allows recycling of cytoplasmic constituents into bioenergetic and biosynthetic materials for maintenance of homeostasis. Since the function of autophagy is particularly important in various stress conditions, perturbation of autophagy can lead to cellular dysfunction and diseases. Accumulation of abnormal protein aggregates, a common cause of neurodegenerative diseases, can be reduced through autophagic degradation. Recent studies have revealed defects in autophagy in most cases of neurodegenerative disorders. Moreover, deregulated excessive autophagy can also cause neurodegeneration. Thus, healthy activation of autophagy is essential for therapeutic approaches in neurodegenerative diseases and many autophagy-regulating compounds are under development for therapeutic purposes. This review describes the overall role of autophagy in neurodegeneration, focusing on various therapeutic strategies for modulating specific stages of autophagy and on the current status of drug development.

• Korean Journal of Food Science and Technology
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• v.21 no.5
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• pp.714-720
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• 1989

Effects of acetylation on conformational changes of glycinin was studied using solvent perturbation, second derivative spectroscopy, near uv circular dichroism spectra and viscosity. Glycinin with purity of more than 93% was used for the experiment. Modification was carried out with acetic anhydride and glycinin with lysine residue modification of 0%, 28%, 65%, 85%, and 95% were used for the experiment. The result of solvent perturbation using some selected perturbants, such as glycerol, ethylene glycol, and dimethyl sulfoxide revealed that acetylation has caused increase In solvent accessibility of tyrosine residues from less than 40% in native protein to more than 70% for 95% acetylated glycinin. This was confirmed by second derivative spectroscopy. Near ultraviolet circular dichroism revealed that the spectra of native and acetylated glycinin were almost identical differing only in intensity and no other useful information could be derived from it. However, in the case of 95% acetylated glycinin the influence of tryptophan on the spectrum was more pronounced Specific viscosity of glycinin also increased by modification, the extent of which depended upon the degree of acetylation. These results supported that acetylation had caused globular conformation of glycinin to be expanded and denatured.

• Toxicological Research
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• v.21 no.4
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• pp.347-353
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• 2005

Eukaryotic initiation factor 4E (elF4E) is a key element for cap-dependent protein translation controlled by affinity between elF4E and 4E-binding protein 1 (4E-BP1). Rapamycin can also affect protein translation by regulating 4E-BP1 phosphorylation. Tobacco-specific nitrosamine, 4(N-methyl-N-nitrosamino )-1-(3-pyridyl)-1-butanone (NNK) is a strong lung carcinogen, but its precise lung cancer induction mechanism remains unknown. Relative roles of cap-dependent and -independent protein translation in terms of NNK-induced lung carcinogenesis were elucidated using normal human bronchial epithelial cells. NNK concentrations applied in this study did not decrease cell viability. Addition of NNK restored rapamycin-induced decrease of protein synthesis and rapamycin-induced phosphorylation of 4E-BP1, and increased expression levels of mTOR, ERK1/2, p70S6K, and Raf-1 in a concentration-dependent manner. NNK also caused perturbation of normal cell cycle progression. Taken together, NNK might cause toxicity through the combination of restoration of 4E-BP1 phosphorylation and increase of elF4E as well as mTOR protein expression, interruption of Raf1/ERK as well as the cyclin G-associated p53 network. Our data could be applied towards elucidation of the molecular basis for lung cancer treatment.

• KSBB Journal
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• v.26 no.3
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• pp.199-205
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• 2011

Using tetrameric ${\beta}$-galactosidase as a model protein, anchoring motives were screened in Bacillus subtilis spore display system. Eleven spore coat proteins were selected considering their expression levels and the location in the spore coat layer. After chromosomal single-copy homologous integration in the amyE site of Bacillus subtilis chromosome, cotE and cotG were chosen as possible spore surface anchoring motives with their higher whole cell ${\beta}$-galactosidase activity. PAGE and Wester blot of extracted fraction of outer layer of purified spore, which express CotE-LacZ or CotG-LacZ fusion verified the existence of exact size of fusion protein and its location in outer coat layer of purified spore. ${\beta}$-galactosidase activity of spore with CotE-LacZ or CotG-LacZ fusion reached its highest value around 16~20 h of culture time in terms of whole cell and purified spore. After intensive spore purification with lysozyme treatment and renografin treatment, spore of BJH135, which expresses CotE-LacZ, retained only 1~2% of its whole cell ${\beta}$-galactosidase activity. Whereas spore of BJH136, which has cotG-lacZ cassette in the chromosome, retained 10~15% of its whole cell ${\beta}$-galactosidase activity, proving minor perturbation of CotG-LacZ, when incorporated in the spore coat layer of Bacillus subtilis compared to CotE-LacZ. Usage of Bacillus subtilis WB700, of which 7 proteases are knocked-out and thereby resulting in 99.7% decrease in protease activity of the host, did not prevent the proteolytic degradation of spore surface expressed CotG-LacZ fusion protein.

• Nutrition Research and Practice
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• v.1 no.1
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• pp.19-28
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• 2007

To identify regulatory molecules which play key roles in the development of obesity, we investigated the transcriptional profiles in 3T3-L1 cells at early stage of differentiation and analyzed the promoter sequences of differentially regulated genes. One hundred and sixty-one (161) genes were found to have significant changes in expression at the 2nd day following treatment with differentiation cocktail. Among them, 86 transcripts were up-regulated and 75 transcripts were down-regulated. The 161 transcripts were classified into 10 categories according to their functional roles; cytoskeleton, cell adhesion, immune, defense response, metabolism, protein modification, protein metabolism, regulation of transcription, signal transduction and transporter. To identify transcription factors likely involved in regulating these differentially expressed genes, we analyzed the promoter sequences of up- or - down regulated genes for the presence of transcription factor binding sites (TFBSs). Based on coincidence of regulatory sites, we have identified candidate transcription factors (TFs), which include those previously known to be involved in adipogenesis (CREB, OCT-1 and c-Myc). Among them, c-Myc was also identified by our microarray data. Our approach to take advantage of the resource of the human genome sequences and the results from our microarray experiments should be validated by further studies of promoter occupancy and TF perturbation.

• Animal cells and systems
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• v.11 no.1
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• pp.9-15
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• 2007

The naphthoquinone analog (2,3-dichloro-6,9-dihydroxy-1,4-naphtoquinone, NA) has an inhibitory effect on cdc25A protein phosphatase in vitro, which is responsible for G1/S transition during cell cycle. However, the exact mechanism inducing the growth inhibition is not understood. In this study, we investigated the regulatory mechanisms of growth arrest induced by NA, as a new potent inhibitor of cdc25A phosphatase, in human hepatocarcinoma SK-hep-1 cells. We found that NA induced the G1 arrest by perturbation of protein tyrosine dephosphorylation of Cdk2, which may be resulting from inhibition of cdc25A phosphatase. In addition, p21 was expressed in a p53-independent manner and participated in the NA-induced G1 arrest by inhibiting Cdk2 activity. Although the exact mechanism is not known, the p21 expression might be related to MAPK activation. From these results, we suggest that NA induces G1 arrest via inhibition of cdc25A and induction of p53-independent p21 expression in SK-Hep-1 cells.

• Journal of the Korean Magnetic Resonance Society
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• v.21 no.3
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• pp.102-108
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• 2017

Escherichia coli responds to ever-changing external and internal stresses by rapidly adjusting its physiology for better survival. This adjustment occurs at all levels including metabolites as well as mRNAs and proteins. Although there has been many reports describing E. coli's adaptation to various stresses regarding transcriptomics or proteomics, only a few investigations have been reported regarding this adaptation viewed from metabolites' perspective. We applied four different types of stresses at four different doses as imposed by NaCl, sorbitol, ethanol, and pH to investigate the similarities or differences among the stresses, and which stress causes the largest perturbation of the metabolite composition. We profiled the metabolites under such external stresses by using two-dimensional NMR spectroscopy and identified 39 metabolites including amino acids, sugars, organic acids, and nucleic acids. According to our statistical analysis, the osmotic stress caused by sorbitol differentiated itself from others, while NaCl showed the largest dose dependent metabolic perturbations. We hope this work will form a foundation on which an approach to a successful protein production is systematically provided by a favorable metabolic environment by imposing proper external stresses.

• Animal cells and systems
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• v.15 no.1
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• pp.1-8
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• 2011

Perturbation of metabolism with increased expression of lipogenic enzymes is a common characteristic of human cancers, including prostate cancer. In the present work the overexpression of stearoyl-CoA desaturase (SCD) in LNCaP cells led to increased mRNA levels of fatty acid synthase (FAS) and acetyl-CoA-carboxylase-a, whereas micro RNA-mediated silencing of SCD inhibited the expression of these lipogenic genes in LNCaP cells. Treatment with the FAS-specific inhibitor cerulenin inhibited SCD induction of LNCaP cell proliferation. In addition, a transient transfection assay revealed the capability of cerulenin to suppress SCD and dihydrotestosterone induction of androgen receptor transcriptional activity. Furthermore, overexpression of SCD in LNCaP cells produced marked resistance to ceramide-induced cell death with reduced poly(ADP-ribose) polymerase (PARP) cleavage. In contrast, silencing of SCD expression increased Bax protein in LNCaP cells. Furthermore, addition of ceramide to SCD knockdown LNCaP cells increased cell death and caspase-3 activity with drastic increase of PARP cleavage. Together, the data indicate that SCD may provide resistance of prostate cancer cells to ceramide-induced cell death.

• Journal of the Korean Magnetic Resonance Society
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• v.19 no.2
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• pp.67-73
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• 2015

The microcystin is a cyclic heptapeptide from metabolites of cyanobacteria in the genera mycrocystis, anabaeba as a result of eutrophication. It has been known that microcystin-LR is a potent inhibitor of the catalytic subunits of protein phosphatase-1 (PP-1) as well as powerful tumor promoter. The active site of microcystin actually has two metal ions $Fe^{2+}/Zn^{2+}$ close to the nucleophilic portion of PP-1-microcystin complex. We report the isolation and purification of this microcystin-LR from cyanobacteria (blue-green algae) obtained from Daechung Dam in Chung-cheong Do, Korea. Microcystin-LR was extracted from solid-phase extraction (SPE) sample preparation using a CN cartridge. The cyanobacteria extract was purified to obtain microcystin-LR by HPLC method and identified by LC/MS. The detail structural studies that can elucidate the possible role of monovalent and divalent metal ions in PP-1-microcystin complexation were carried out by utilizing molecular dynamics. Conformational changes in metal binding for ligands were monitored by molecular dynamic computation and potential of mean force (PMF) using the method of the free energy perturbation. The microcystin-metal binding PMF simulation results exhibit that microcystin can have very stable binding free energy of -10.95 kcal/mol by adopting the $Mg^{2+}$ ion at broad geometrical distribution of $0.5{\sim}4.5{\AA}$, and show that the $K^+$ ion can form a stable metal complex rather than other monovalent alkali metal ions.

• Journal of Physiology & Pathology in Korean Medicine
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• v.20 no.6
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• pp.1516-1523
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• 2006

The water extract of Danchunhwan(DCH) has been traditionally used for treatment of dementia damage in oriental medicine. However, little is known about the mechanism by which the water extract of DCH rescues cells from neurodegenerative disease such as Alzheimer's disease. This study was designed to investigate the protective mechanisms of DCH on ${\beta}$-amyloid or $H_2O_2$-induced cytotoxicity in SH-SY5Y neuronblastoma cells. ${\beta}$-amyloid and $H_2O_2$ markedly decreased the viability of SH-SY5Y cells, which was characterized with apparent apoptotic features such as membrane blebbing as well as fragmentation of genomic DNA and nuclei. However, the water extract of DCH significantly reduced both ${\beta}$-amyloid or $H_2O_2$-induced cell death and apoptotic characteristics through reduction of intracellular peroxide generation. Also, the water extract of DCH prevented prevented the mitochondrial dysfunction including the disruption of mitochondria membrane permeability transition (MPT) and the perturbation in Bcl-2 family protein expressions in $H_2O_2$-treated SH-SY5Y cells.