• Korean Journal of Microbiology
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• v.52 no.2
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• pp.236-241
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• 2016

The Schizosaccharomyces pombe $sdu1^+$ gene, belonging to the PPPDE superfamily of deubiquitinating enzyme (DUB) genes, was previously shown to encode a protein with ubiquitin C-terminal hydrolase (UCH) activity and to participate in the response against oxidative and nitrosative stresses. This work focused on the reactive oxygen species (ROS)-dependent regulation of the S. pombe $sdu1^+$ gene. UCH activities, encoded by the $sdu1^+$ gene, were attenuated in the S. pombe cells exposed to $H_2O_2$, superoxide radical-generating menadione (MD), and nitric oxide (NO)-generating sodium nitroprusside (SNP). Reduced glutathione (GSH) and its precursor N-acetylcysteine (NAC) were able to significantly enhance the UCH activities in the absence or presence of $H_2O_2$. However, the influences of both GSH and NAC on the ROS levels in the absence or presence of $H_2O_2$ were opposite to their effects on the UCH activities under the same conditions. The UCH activities in the Sdu1-overexpressing S. pombe cells were also diminished under exposure to $H_2O_2$, MD and SNP, but still remained to be higher than those in the vector control cells. In brief, it is proposed that the S. pombe $sdu1^+$ gene is regulated by ROS in a negative manner, the meaning of which largely remains elusive.

• Applied Biological Chemistry
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• v.46 no.4
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• pp.305-310
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• 2003

Hydrophobic core variant of ubiquitin appeared to have partially folded structure at pH around 2. The intrinsic tryptophan fluorescence emission maximum of this ubiquitin variant at pH 2 showed slight blue shift compare to that of unfolded state, suggesting that some residual tertiary structures remain in this solvent condition. At the same solvent condition, this ubiquitin variant binds with hydrophobic dye, 8-anilinonaphthalene-1-sulfonic acid(AMS), which is known to bind to exposed hydrophobic surface. Furthermore, far-UV circular dichroic spectrum of this ubiquitin variant in the diminished pH was remarkably different from the far-UV CD spectrum of the native state or unfolded state. Based on the molar ellipticity at 220 nm, this ubiquitin variant at pH 2 appeared to have significant amount of secondary structures. All these observations suggest that this ubiquitin variant in the diminished solvent pH has loosely folded hydrophobic core with some secondary structures, which are key features of molten globule conformation. Since molten globule has long been considered as a protein folding intermediate, it is considered that this hydrophobic core variant ubiquitin will serve as a valuable model to study protein folding process.

• KSBB Journal
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• v.24 no.3
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• pp.217-226
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• 2009

p53 undergoes various post-translational modifications, including phosphorylation, ubiquitination, sumoylation, acetylation, methylation, and poly(ADP-ribosyl)ation. Modification of p53 widely affects to various functions of p53. Acetylation and phosphorylation of p53 have been studied for regulating its transcriptional activity which is observed in various stress condition. Otherwise, ubiquitination of p53 by Mdm2 has been well-studied as a canonical ubiquitin-mediated proteasomal degradation pathway. Moreover several investigators have recently reported that ubiquitination of p53 modulates not only its proteasome-dependent degradation by poly-ubiquitination but also its localization and transcriptional activity by mono-ubiquitination which usually does not serve the proteasome dependent degradation. Here we review recent studies on the cellular functions of p53 regulated by post-translational modifications, particularly focusing on mechanisms of ubiquitination.

• Journal of the Korean Chemical Society
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• v.49 no.5
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• pp.479-487
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• 2005

• Clinical and Experimental Reproductive Medicine
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• v.32 no.3
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• pp.231-242
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• 2005

연구목적: 본 연구는 아직 그 기능이 파악되지 않은 탈유비퀴틴효소 중 하나인 HIDE에 대한 기본적인 생화학적 특징과 고환에서의 발현 양상을 파악하고 있다. 연구재료 및 방법: 인간의 HIDE 유전자를 클로닝하여 효소활성이 있는지 세포 외 실험을 통해 확인하였고, 아미노산 서열을 분석하여 진화상 보존된 부분을 찾아 그 기능을 파악한 다음 HSP90과의 상호작용을 공동면역침전반응으로 확인하였다. HIDE의 조직별 발현양상을 파악하기 위해서 인간과 쥐의 RNA 블롯과 쥐의 단백질 블롯을 이용하여 각각 노던 블롯팅과 웨스턴 블롯팅을 수행하여 고환에서 많이 발현된다는 것을 알았고 이 사실을 바탕으로 쥐의 고환을 절개하여 면역조직화학반응으로써 고환 내의 HIDE 단백질의 발현양상을 파악하였다. 결　과: HIDE는 세포 외에서 유비퀴틴 잔기를 제거하는 탈유비퀴틴 활성이 있으나 세포 내에서 전체적인 유비퀴틴 복합체를 줄여주는 효과는 없었다. HIDE는 HSP90이라는 분자 샤페론과 상호작용한다. HIDE의 전사체는 고환에서 가장 많이 발현되며 다른 조직에서도 소량 발현된다. HIDE의 단백질은 웨스턴 블롯상에서 고환에서만 확인되었다. 고환 내에서의 HIDE의 발현양상은 왕성한 감수분열을 하는 정모세포에서 높았으며 지지세포나 정조세포에는 발현되지 않았다. 결　론: HIDE는 분자 샤페론 HSP90과 상호작용하며 고환 내의 감수분열 중인 세포에서 많이 발현되는 것으로 보아 감수분열이나 정자형성에 관여하는 것으로 보인다.

• Proceedings of the Korean Society of Crop Science Conference
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• 2021.10a
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• pp.193-193
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• 2021

• Journal of Life Science
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• v.28 no.9
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• pp.1007-1015
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• 2018

The E6-associated protein (E6AP) is known to induce the ubiquitination and proteasomal degradation of HCV core protein and thereby directly impair capsid assembly, resulting in a decline in HCV replication. To counteract this anti-viral host defense system, HCV core protein has evolved a strategy to inhibit E6AP expression via DNA methylation. In the present study, we further explored the mechanism by which HCV core protein inhibits E6AP expression. HCV core protein upregulated both the protein levels and enzyme activities of DNA methyltransferase 1 (DNMT1), DNMT3a, and DNMT3b to inhibit E6AP expression via promoter hypermethylation in HepG2 cells but not in Hep3B cells, which do not express p53. Interestingly, p53 overexpression alone in Hep3B cells was sufficient to activate DNMTs in the absence of HCV core protein and thereby inhibit E6AP expression via promoter hypermethylation. In addition, upregulation of p53 was absolutely required for the HCV core protein to inhibit E6AP expression via promoter hypermethylation, as evidenced by both p53 knockdown and ectopic expression experiments. Accordingly, levels of the ubiquitinated forms of HCV core protein were lower in HepG2 cells than in Hep3B cells. Based on these observations, we conclude that HCV core protein evades ubiquitin-dependent proteasomal degradation in a p53-dependent manner.

• Journal of the Korean Chemical Society
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• v.58 no.6
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• pp.560-568
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• 2014

The contribution of electrostatic interactions to protein folding reaction was studied by using mutant ubiquitin with lysine to alanine mutation at residue position 29. Based on the three dimensional structure of ubiquitin, lysine 29 is located close to negatively charged glutamate 16 and aspartate 21 and considered to stabilize the native state of ubiquitin by electrostatic interactions between these residues. The equilibrium unfolding experiment showed that the native stability was decreased by about ~20% upon mutation. This observation indicates lysine 29 indeed forms electrostatic interactions with nearby residues. Folding kinetics measurements using stopped-flow device and quantitative analysis of kinetics data indicate that ubiquitin folds from unfolded state to native state via intermediate state as observed previously. This intermediate state was observed to form immediately after the initiation of folding reaction. The folding intermediate was shown to be destabilized considerably upon lysine to alanine mutation. These observations indicate that electrostatic interactions can form early stage of protein folding and hence lead the folding reaction.

• Journal of Life Science
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• v.28 no.10
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• pp.1132-1139
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• 2018

The stability of diverse cellular proteins in eukaryotes is regulated via ubiquitination. Moreover, E3 ligase plays a crucial role in determining substrate specificity and transfers ubiquitins into the substrates during the ubiquitination process. As a type of multi-subunit E3 ligase, cullin4 (CUL4)-based E3 ligase (CRL4) complex is involved in a variety of cellular processes, such as hormonal and stress responses in plants. In spite of several reports on the versatile roles of CRL4 in various signalings in Arabidopsis, CRL4's function in rice has been poorly known. To learn about CRL4-mediated cellular processes in rice in more detail, OsCUL4 that exhibits the highest homology with Arabidopsis CUL4 was isolated, and its expression patterns in various tissues and in response to plant hormones and abiotic stresses were monitored. Exogenous application of ABA or cytokinin increased the transcript levels of the OsCUL4 gene. Moreover, OsCUL4 was significantly upregulated in response to drought and salt stresses. These findings imply that OsCUL4 may be functionally related to ABA- and/or cytokinin-mediated cellular responses. OsCUL4 directly interacted with OsDDB1, an adaptor protein of CRL4, indicating that OsCUL4 can act as a scaffold protein of CRL4. An expression study on the OsCUL4 gene from this report could be used as a starting point to elucidate cellular responses in which a CRL4-mediated ubiquitination process is involved in rice.

• Journal of Life Science
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• v.32 no.6
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• pp.476-481
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• 2022

Ubiquitin signaling regulates virtually all aspects of eukaryotic biology and dynamic processes in which protein substrates are modified by ubiquitin. To regulate these processes, deubiquitinating enzymes (DUBs) cleave ubiquitin or ubiquitin-like proteins from these substrates. DUBs have been implicated in the pathogenesis of cancer, leading to the development of increasing numbers of small-molecule DUB inhibitors. On the other hand, recent studies have focused on the function of DUBs in metabolic diseases such as obesity, diabetes, and fatty liver diseases. DUBs play a positive or negative role in the progression and development of metabolic diseases. Their involvement in cell pathology and regulation of major transcription factors in metabolic syndrome has been examined in vitro and in animal and human biopsies. UCH, USP7, and USP19 were linked to adipocyte differentiation, body weight gain, and insulin resistance in genetic or diet-induced obesity. CYLD, USP4, and USP18 were found to be closely associated with fatty liver diseases. In addition, these liver diseases were accompanied by body weight change in certain cases. Collectively, in this review, we discuss the current understanding of DUBs in metabolic diseases with a particular focus on obesity. We also provide basic knowledge and regulatory mechanisms of DUBs and suggest these enzymes as therapeutic targets for metabolic diseases.