• IMMUNE NETWORK
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• 제4권3호
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• pp.184-189
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• 2004

Background: Hu syndrome, a neurological disorder, is characterized by the remote effect of small cell lung cancer on the neural degeneration. The suspicious effectors for this disease are anti-Hu autoantibodies or Hu-related CD8+ T lymphocytes. Interestingly, the same effectors have been suggested to act against tumor growth and this phenomenon may represent natural tumor immunity. For these diagnostic and therapeutic reasons, the demand for antibodies against Hu protein is rapidly growing. Methods: Polyclonal and monoclonal antibodies were generated using recombinant HuR protein. Western blot analyses were performed to check the specificity of generated antibodies using various recombinant proteins and cell lysates. Extracellular stimuli for HuR expression had been searched and HuR-associated proteins were isolated from polysome lysates and then separated in a 2-dimensional gel. Results: Polyclonal and monoclonal antibodies against HuR protein were generated and these antibodies showed HuR specificity. Antibodies were also useful to detect and immunoprecipitate endogenous HuR protein in Jurkat and BJAB. This report also revealed that TNF-${\alpha}$ treatment in BJAB up-regulated HuR expression. Lastly, protein profile in HuR-associated mRNAprotein complexes was mapped by 2-dimensional gel electrophoresis. Conclusion: This study reported that new antibodies against HuR protein were successfully generated. Currently, project to develop a diagnostic kit is in process. Also, this report showed that TNF-${\alpha}$ up-regulated HuR expression in BJAB and protein profile associated with HuR protein was mapped.

• 한국미생물·생명공학회지
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• 제18권5호
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• pp.490-495
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• 1990

토양 분리균인 Bacillus spp.가 생산하는 inulinase를 ammonium sulfate 분획, 열처리, DEAE Sephadex Cl-6B ion exchange chromatography, Sephadex G-100 및 Sephadex G-150 gel 여과 등의 과정을 거쳐 단일 단백질로 분리 정제하였다. 정제 inulinase는 분자량이 약 56,000인 효소로서 pH6.0,$50^{\circ}C$에서 최대 활성을 나타내었으며 $Co^{2+}$와 $Mn^{2+}$에 의해서 현저한 활성화 효과를 보였다. 또한 본 효소는 sucrose와 raffinose에 대해서도 높은 활성을 나타내므로 $\beta$-D-fructofuranosidase(EC 3.2.1.26)로 분류되는 exo-inulinase인 것으로 확인되었다. 한편 효소활성에 필수적인 아미노산 잔기는 tryptophan과 histidine인 것으로 분석되었으며 inulin과 sucrose에 대한 $K_m$값은 각각 $2.0 \times 1.0^[-3}M, 1.0 \times 10^[-2}M$로 산출되었다.

• Molecules and Cells
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• 제23권1호
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• pp.1-10
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• 2007

Invading pathogens are recognized by diverse germline-encoded pattern-recognition receptors (PRRs) which are distributed in three different cellular compartments: extracellular, membrane, and cytoplasmic. In mammals, the major extracellular PRRs such as complements may first encounter the invading pathogens and opsonize them for clearance by phagocytosis which is mediated by membrane-associated phagocytic receptors including complement receptors. The major membrane-associated PRRs, Toll-like receptors, recognize diverse pathogens and generate inflammatory signals to coordinate innate immune responses and shape adaptive immune responses. Furthemore, certain membrane-associated PRRs such as Dectin-1 can mediate phagocytosis and also induce inflammatory response. When these more forefront detection systems are avoided by the pathogens, cytoplasmic PRRs may play major roles. Cytoplasmic caspase-recruiting domain (CARD) helicases such as retinoic acid-inducible protein I (RIG-I)/melanoma differentiation-associated gene 5 (MDA5), mediate antiviral immunity by inducing the production of type I interferons. Certain members of nucleotide-binding oligomerization domain (NOD)-like receptors such as NALP3 present in the cytosol form inflammasomes to induce inflammatory responses upon ligand recognition. Thus, diverse families of PRRs coordinately mediate immune responses against diverse types of pathogens.

• 동의생리병리학회지
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• 제27권5호
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• pp.563-569
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• 2013

This study was undertaken to apply the yin-yang theory in action potential. In order to apply the yin-yang theory in action potential, nature of yin and yang, interrelation of yin and yang and action potential in cell were reviewed. According to the yin-yang theory, inner cellular space corresponds to yin, but outer cellular space corresponds to yang. If we classify ions in intracellular fluid or extracellular fluid by nature of yin and yang, potassium(K+) corresponds to yang within yin(陰中之陽), protein(Pr-) corresponds to yin within yin(陰中之陰) in intracellular fluid, and sodium(Na+) corresponds to yang within yang(陽中之陽), chloride(Cl-) corresponds to yin within yang(陽中之陰) in extracellular fluid. Double donnan equilibrium and equilibrium potential were caused by intracellular anion(Pr-) and extracellular cation(Na+) are related with mutual rooting of yin and yang(陰陽互根) and opposition of yin and yang(陰陽對立). The influx and efflux of ion through cell membrane means waxing and waning of yin and yang(陰陽消長), the change of membrane potential means yin-yang conversion(陰陽轉化) during action potential.

• Journal of Microbiology and Biotechnology
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• 제14권5호
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• pp.1004-1008
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• 2004

An extracellular acid phytase from Pseudomonas fragi Y9451 was purified to homogeneity from the culture supernatant by salting-out, DEAE-Sepharose column chromatography, CM-Sepharose column chromatography, and Sephacryl S-300 gel filtration. The molecular weight of the purified enzyme was estimated to be 74 kDa on gel filtration and 54 kDa and 25 kDa on SDS-PAGE, suggesting that the native enzyme was a heterodimeric protein. The purified enzyme was most active at pH 4.5 and $70^{\circ}C$ and fairly stable from pH 4.0- 6.0. It was specific for phytate and exhibited a $K_{m}$ value of 27 mM (sodium phytate, pH 4.5, $50^{\circ}C$). The phytase activity was strongly inhibited (at maximum by 87%) by $Fe^{3+},\;Cu^{2+},\;Fe^{2+}$, and $Zn^{2+}$ at 5 mM concentration, and greatly inhibited by $Ca^{2+}$ at 10 mM concentration. However, EDTA notably stimulated the phytase activity at 10 mM concentration. With optimum pH and stability, Pseudomonas fragi phytase could be a potential candidate for animal feed applications.

• Journal of Microbiology
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• 제37권1호
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• pp.14-20
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• 1999

Two extracellular proteases, EP I and EP II, from cells of Oligotropha carboxydovorans (formerly Pseudomonas carboxydovorans) DSM 1227 grown in nutrient broth were purified to greater than 95% homogeneity in five steps using azocasein as a substrate. The final specific activities of EPs I and II were 214.9 and 667.4 units per mg of protein. The molecular weights of native EPs I and II were determined to be 23,000. Sodium dodecyl sulfate-gel electrophoresis revealed the two enzymes to be monomers. The enzymes were found to be serine-type proteases. The activity of EP I was stimulated by Ca2+, Mg2+, and Ba2+, but that of EP II was not. The enzymes were completely inhibited by Fe2+, Hg2+, Co2+, Zn2+, and Cd2+. EDTA and EGTA exhibited a strong inhibitory effect on EP I. The optimal pH for the two enzymes was pH 9.0. The optimal temperatures for EP I and II were 60 and 50$^{\circ}C$, respectively. The enzymes were stable under alkaline conditions. The thermal stability of EP I was higher than that of EP II. Cell-free extracts did not inhibit the purified enzymes. The enzymes were active on casein, azocasein, azocoll, and carbon monoxide dehydrogenase, but weakly active with bovine serum albumin.

• Journal of Microbiology and Biotechnology
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• 제16권10호
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• pp.1485-1490
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• 2006

In this study, comparative replicational properties of Hyphantria cunea nucleopolyhedrovirus (HycuNPV) in Lymantria dispar (IPLB-LdElta) and Spodoptera frugiperda (IPLB-Sf21) cell lines were investigated. Our microscopic observations showed that cytopathic effects (CPEs) in LdElta cells appeared 12 h later than those in Sf21 cells. Whereas polyhedral inclusion bodies (PIBs) formed at 48 h postinfection (p.i.) in LdElta cells, it formed at 36 h p.i. in Sf21 cells. Extracellular virus production determined according to the 50% tissue culture infective dose ($TCID_{50}$) method in LdElta cells started about 12 h later when compared with Sf21 cells. Titers of extracellular virus in LdElta and Sf21 cells were calculated as $1.77{\times}10^9$ plaque forming units (PFU)/ml and $5.6{\times}10^9PFU/ml$, respectively, at 72 h p.i. We also showed that viral DNA replication began at 12 h p.i. in both cell lines. Viral protein synthesis was determined by SDS-polyacrylamide gel electrophoresis (PAGE) and polyhedrin synthesis was observed at 12 h p.i. in both cell lines. The results indicate that while the synthesis of macromolecules is 12 h later and production of extracellular virus is almost 3-fold lower in LdElta cells compared with those in Sf21 cells, the LdElta cell line is still a productive cell line for infection of HycuNPV.

• 생약학회지
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• 제50권1호
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• pp.18-24
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• 2019

Andrographolide, the main compound of Andrographis paniculata (A. paniculata), shows various biological properties including anti-viral, anti-inflammatory, anti-diabetic, and hepatoprotective effects. Our previous study has shown that A. paniculata extract exerts antiaging effects by activation of stemness in epidermal stem cells (EpSCs). In this study, we investigated the effect of andrographolide as a main compound of A. paniculata on EpSCs and its mechnism of action using several in vitro assays. Andrographolide increased the proliferation of EpSCs and induced cell cycle progression. Additionally, andrographolide increased VEGF production and the expression of stem cell markers integrin ${\beta}1$ and p63. Furthermore, phosphorylation levels of extracellular signal-regulated kinase 1/2 (ERK1/2), S6 ribosomal protein (S6RP) and Akt were increased by andrographolide. Taken together, these results indicate that andrographolide-induced proliferation of EpSCs is mediated by the ERK1/2, Akt-dependent pathway with increased production of VEGF and upregulated stemness through integrin ${\beta}1$ and p63.

• Journal of Microbiology and Biotechnology
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• 제33권5호
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• pp.582-590
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• 2023

Stress granules (SGs) are cytoplasmic aggregates of RNA-protein complexes that form in response to various cellular stresses and are known to restrict viral access to host translational machinery. However, the underlying molecular mechanisms of SGs during viral infections require further exploration. In this study, we evaluated the effect of SG formation on cellular responses to coxsackievirus B3 (CVB3) infection. Sodium arsenite (AS)-mediated SG formation suppressed cell death induced by tumor necrosis factor-alpha (TNF-a)/cycloheximide (CHX) treatment in HeLa cells, during which G3BP1, an essential SG component, contributed to the modulation of apoptosis pathways. SG formation in response to AS treatment blocked CVB3-mediated cell death, possibly via the reduction of mitochondrial reactive oxygen species. Furthermore, we examined whether AS treatment would affect small extracellular vesicle (sEV) formation and secretion during CVB3 infection and modulate human monocytic cell (THP-1) response. CVB3-enriched sEVs isolated from HeLa cells were able to infect and replicate THP-1 cells without causing cytotoxicity. Interestingly, sEVs from AS-treated HeLa cells inhibited CVB3 replication in THP-1 cells. These findings suggest that SG formation during CVB3 infection modulates cellular response by inhibiting the release of CVB3-enriched sEVs.

• Journal of Korean Neurosurgical Society
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• 제46권4호
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• pp.389-396
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• 2009

Objective : Triptolide (TP) has been reported to suppress the expression of mitogen-activated protein kinase (MAPK) phosphatase-1 (MKP-1), of which main function is to inactivate the extracellular signal-regulated kinase-1/2 (ERK-1/2), the p38 MAPK and the c-Jun N-terminal kinase-1/2 (JNK-1/2), and to exert antiproliferative and pro-apoptotic activities. However, the mechanisms underlying antiproliferative and pro-apoptotic activities of TP are not fully understood. The purpose of this study was to examine whether the down-regulation of MKP-1 expression by TP would account for antiproliferative activity of TP in immortalized HT22 hippocampal cells. Methods : MKP-1 expression and MAPK phosphorylation were analyzed by Western blot. Cell proliferation was assessed by $^3H$-thymidine incorporation. Small interfering RNA (siRNA) against MKP-1, vanadate (a phosphatase inhibitor), U0126 (a specific inhibitor for ERK-1/2), SB203580 (a specific inhibitor for p38 MAPK), and SP600125 (a specific inhibitor for JNK-1/2) were employed to evaluate a possible mechanism of antiproliferative action of TP. Results : At its non-cytotoxic dose, TP suppressed MKP-1 expression, reduced cell growth, and induced persistent ERK-1/2 activation. Similar growth inhibition and ERK-1/2 activation were observed when MKP-1 expression was blocked by MKP-1 siRNA and its activity was inhibited by vanadate. The antiproliferative effects of TP, MKP-1 siRNA, and vanadate were significantly abolished by U0126, but not by SB203580 or SP600125. Conclusion : Our findings suggest that TP inhibits the growth of immortalized HT22 hippocampal cells via persistent ERK-1/2 activation by suppressing MKP-1 expression. Additionally, this study provides evidence supporting that MKP-1 may play an important role in regulation of neuronal cell growth.