• Bulletin of the Korean Chemical Society
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• v.34 no.4
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• pp.1089-1095
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• 2013

The tumor necrosis factor-receptor 1 (TNFR1)-associated death domain protein (TRADD) contains an N-terminal TRAF binding domain and a C-terminal death domain. TRADD is known to interact directly with TNF receptor 2 (TNFR2) and the Fas-associated death domain protein (FADD), which are signal transducers that activate NF-${\kappa}B$ and induce apoptosis, respectively. To date, there has been no structural information on the TRADD and FADD death domain (DDs) complex. In this study, the death domains of TRADD and FADD were co-expressed and purified from Escherichia coli for structural characterization. We found that human TRADD (hTRADD) interacted strongly with mouse FADD (mFADD) via their DDs and interacted weakly with human FADD (hFADD)-DD. Moreover, the structures of the TRADD-DD:FADD-DD complexes were separately modeled from predicted structures in the protein data bank (PDB). The results of this study will have important applications in human diseases such as cancer, AIDS, degenerative and autoimmune diseases, and infectious diseases.

• BMB Reports
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• v.35 no.1
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• pp.24-27
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• 2002

Apoptotic cell death is an active process mediated by various signaling pathways, which include the caspase cascade and the stress-activated protein kinase pathways. The caspase cascade is activated by two distinct routes: one from cell surface and the other from mitochondria. Activation of the route from cell surface requires the cellular components that include membrane receptors, adaptor proteins such as TRADD and FADD, and caspase-8, while activation of the other from mitochondria requires Apaf-1, caspase-9, and cytosolic cytochrome c. On the other hand, persistent stimulation of the stress-activated protein kinase pathway is also shown to mediate apoptosis in many cell types. Gene-targeting studies with jnk- or jip-null mice, in particular, strongly suggest that this signaling pathway plays a pivotal role in the cellular machinery for apoptosis.

• The Journal of Internal Korean Medicine
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• v.25 no.1
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• pp.28-45
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• 2004

Objectives : The main purpose of this study is to evaluate the effect of Injinchunggan-tang on $TNF-{\alpha}$ signal transmission system. Materials and Methods : We analyzed the following with quantitative RT-PCR method; the effect of Injinchunggan-tang on secretion of $TNF-\alpha$ mRNA/protein and stability, the effect on gene revelation that consists of signal transmission system (TRAIL, NIK, A20, TRADD, RAIDD, RIP TNFR-I, TNFR-II, TRAF1, TRAF2, FADD), the one on activation of p38, Erk1/2 MAPK and the rate of nuclear $NF-{\kappa}B/cytosolic\;NF-{\kappa}B$ in HepG2 cell. We also analyzed the inhibitory effect of Injinchunggan-tang on the apoptosis of HepG2 cell that $TNF-{\alpha}$ induces and the $NF-{\kappa}B$ restraint effected by transfection of $I{\kappa}B{\Delta}N$ through tryphan blue exclusion assay. Results : Injinchunggan-tang prohibits revelation of $TNF-{\alpha}$ mRNA in HepG2 cell and the creation of protein. However, it has no effect on the stability of $TNF-{\alpha}$ mRNA. While it did not have any effect on the generation of TRAIL, NIK, A20, TRADD, RAIDD and RIP genes, Injinchunggan-tang reduces the revelation of TNFR-I, TNFR-II, TRAF1, TRAF2 and FADD genes. It has been confirmed that Injinchunggan-tang restraints the revelation of $TNF-{\alpha}$ mRNA that is promoted by ethanol, acetaldehyde, lipopolysaccharide, in proportion to the treatment density and time. It activated $NF-{\kappa}B$ of HepG2 cell and promoted activation of $NF-{\kappa}B$ that is occurred by $TNF-{\alpha}$. It has been observed that the restraint effect against the $TNF-{\alpha}$ inducing apoptosis is lost when it is intercepted the function of $NF-{\kappa}B$ in HepG2 cell. Conclusion: It has been confirmed that Injinchunggan-tang has restraining effect against the revelation of $TNF-{\alpha}$ and mRNA that is constituent element of TNF-a signal transmission system. It also has been revealed that it restraints the activation of p38, Erk1/2 by $TNF-{\alpha}$. Through this prohibiting effect, it is inferred that it restraints signal transmission among various cells that are related to inflammation reaction. Meanwhile, Injinchunggan-tang protects liver cell from apoptosis that is caused by $TNF-{\alpha}$, by maintaining the activating function for $NF-{\kappa}B$.

• Journal of Microbiology and Biotechnology
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• v.22 no.12
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• pp.1665-1672
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• 2012

An 80% ethanol extract of Hizikia fusiforme was obtained and followed by successive fractionation using the organic solvents n-hexane, ethyl acetate, and n-butanol to identify the antioxidative substance. The aqueous part of the nbutanol fractionation step, showing high antioxidative activity, was subjected to reverse-phase liquid chromatography. As a result, a substance purified from a BB-2 fraction showed high antioxidative activity. The m/z 419 [M+H] molecular ion peak in the fraction was observed by the analysis of the ESI-LC/MS spectrum. By the analysis of 1H NMR (500 MHz, DMSO-$d_6$) and $^{13}C$ NMR (125 MHz, DMSO-$d_6$) spectra, a unique compound of the fraction was biochemically identified as a 5-hydroxy-3,6,7,8,3',4'-hexamethoxyflavone (5HHMF). We also investigated the effect of 5HHMF on human gastric AGS carcinoma cells. Western blot analysis suggested that the flavone substantially increased the levels of the death receptor-associated apoptosis mediators Fas, Fas L, FADD, TRADD, and DR4 in a concentration-dependent manner. The levels of Fas, Fas L, TRADD, and DR4 in the cells treated with 5HHMF ($5{\mu}g/ml$) were approximately 26.4-, 12.8-, 6.7-, and 9.8-times higher than those of non-treated cells, respectively. Of note, the level of FADD protein in the cells exposed to 5HHMF ($1{\mu}g/ml$) increased approximately 9.6-times. In addition, the cleavage of caspase-3, -8, and -9 in cultured AGS cells treated with 5HHMF was significantly confirmed. Therefore, our results suggest that 5HHMF from H. fusiforme is involved in the induction of death receptor-associated apoptosis mediators in human gastric AGS carcinoma cells.