• Bulletin of the Korean Chemical Society
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• 제35권12호
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• pp.3642-3644
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• 2014

• BMB Reports
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• 제44권7호
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• pp.458-461
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• 2011

Enteropeptidase is a serine protease secreted by the pancreas and converts inactive trypsinogen to active trypsin. Enteropeptidase cleaves the C-terminal end of the substrate recognition sequence Asp-Asp-Asp-Asp-Lys ($D_4K$). The assay for enteropeptidase has utilized $GD_4K$-conjugated 2-naphthylamine ($GD_4K$-NA) as a fluorogenic probe over the last 30 years. However, no other $D_4K$-conjugated fluorogenic substrates of enteropeptidase have been reported. Furthermore, naphthalene is known as carcinogenic to humans. In this study, we used shift in the emission spectrum of $GD_4K$-conjugated 7-amino-4-methylcoumarin ($GD_4K$-AMC) as a fluorogenic method to measure enteropeptidase activity. The kinetic analysis revealed that enteropeptidase has a $K_M$ of 0.025 mM and a $k_{cat}$ of 65 $sec^{-1}$ for $GD_4K$-AMC, whereas it has a $K_M$ of 0.5 to 0.6 mM and a $k_{cat}$ of 25 $sec^{-1}$ for $GD_4K$-NA. The optimum pH of $GD_4K$-AMC hydrolysis was pH 8.0. Our data indicate that $GD_4K$-AMC is more suitable as a substrate for enteropeptidase than $GD_4K$-NA.

• IMMUNE NETWORK
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• 제22권3호
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• pp.28.1-28.11
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• 2022

The 26S proteasome irreversibly hydrolyzes polyubiquitylated substrates to maintain protein homeostasis; it also regulates immune responses by generating antigenic peptides. An alternative form of the 26S proteasome is the immunoproteasome, which contains substituted catalytic subunits (β1i/PSMB9, β2i/PSMB10, and β5i/PSMB8) instead of constitutively expressed counterparts (β1/PSMB6, β2/PSMB7, and β5/PSMB5). The immunoproteasome expands the peptide repertoire presented on MHC class I molecules. However, how its activity changes in this context is largely elusive, possibly due to the lack of a standardized methodology to evaluate its specific activity. Here, we describe an assay protocol that measures the immunoproteasome activity of whole-cell lysates using commercially available fluorogenic peptide substrates. Our results showed that the most accurate assessment of immunoproteasome activity could be achieved by combining β5i-targeting substrate Ac-ANW-AMC and immunoproteasome inhibitor ONX-0914. This simple and reliable protocol may contribute to future studies of immunoproteasomes and their pathophysiological roles during viral infection, inflammation, and tumorigenesis.

• Rapid Communication in Photoscience
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• 제4권4호
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• pp.88-90
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• 2015

The design and development of fluorescent chemosensors have recently been intensively explored for sensitive and specific detection of environmentally and biologically relevant metal ions in aqueous solution and living cells. Herein, we report the photophysical results of rhodamine B based fluorogenic and chromogenic receptor for selective copper detection in the complete organic or mixed aqueous-organic media at neutral pH under ambient condition. The ligand exhibited the remarkable increment in the fluorescence emission and UV-visible absorption signal intensities at 587 and 547 nm, respectively, on induction of copper ion while the ligand solution remain completely silent on addition of varieties of other metal ions.

• Macromolecular Research
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• 제16권1호
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• pp.73-75
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• 2008

• 대한약학회:학술대회논문집
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• 대한약학회 2003년도 Proceedings of the Convention of the Pharmaceutical Society of Korea Vol.1
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• pp.125.2-126
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• 2003

Human tumor necrosis factor-alpha (TNFa) is a key pro-inflammatory cytokine produced by activated monocytes and macrophage as a part of the self-defence machinery. TNF-a converting enzyme (TACE) is the metalloproteinase that processes the membrane bound precursor of TNFa to the soluble component. (omitted)

• 대한의생명과학회지
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• 제18권4호
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• pp.438-444
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• 2012

N-acylethanolamines (NAEs) including endocannabinoids, anadamide, are long chain fatty acid ethanolamines and express ubiquitously in animal and plant tissues. NAEs have several pharmacological effects including anti-inflammatory, analgesic and anorexic effects. The levels of NAEs in tissues are strictly regulated by synthesizing and hydrolyzing enzymes because NAEs are not stored in the cell but rather made on demand. NAEs are hydrolyzed to free fatty acids and ethanolamines by fatty acid amide hydrolase and N-acylethanolamine-hydrolyzing acid amidase (NAAA). Here, we suggest the fluorescence-based assay system for NAAA. We developed N-(4-methy-2-oxo-2H-chromen-7-yl)palmitamide (PAAC) as a fluorogenic substrate for NAAA and we also generated NAAA stably expressing COSM6 cell line. When extracts of cells expressing NAAA were incubated with PAAC, NAAA specifically hydrolyzed PAAC to palmitic acids and fluorogenic dye, coumarin. Release of coumarin was monitored by using fluorometer. NAAA hydrolyzed PAAC with an apparent Km of $20.05{\mu}M$ and Vmax of 32.18 pmol/mg protein/min. This assay system can be used to develop inhibitors or activators of NAAA.

• 미생물학회지
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• 제28권3호
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• pp.229-235
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• 1990

평판배지상 세균 colony의 체외 세포 효소활성을 직접 측정할 수 있는 신속하고 정확한 방법에 대하여 기술하였다. 일반적으로 세균의 효소 특성을 살피기 위해서는 단백질, 전분, chitin, tween-80 등과 같은 고분자 물질을 첨가한 선택배지를 사용하고 있으나 그 방법상 여러 가지 문제점이 있다 그러므로 본 연구에서는 형광물질의 일종인 Methvlumbell liferyl(MUF) 기질이 일반적으보 사용되고 있는 천연 고분자 물질고 유사한가를 순수분리세균 균주를 이용하여 실험으로 검증하였다. MUF 기질 분해원리에 기초를 둔 기술한 새로운 방법은 순수 분리 균주는 물론 colony 계수에 사용되는 평판배지상에서도 세균의 체외세포 효소 특성을 정량적으로 측정 가능하게 한다. 본 새로운 방법을 이용하여 담수 생태계와 해양 퇴적토내 종속영양세균의 체외 효소 활성을 측정하여 고찰하였다.

• Bulletin of the Korean Chemical Society
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• 제31권3호
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• pp.615-619
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• 2010

A new coumarin-thiazolobenzo-crown ether based fluorogenic chemosensor BTC (1) was reported. The ion-selective binding properties of 1 with different alkali, alkaline earth metals and transitional metals were investigated in an ethanol-DMSO system. BTC (1) showed the highest binding constant toward $Hg^{2+}$ over $Ag^+$, $Pb^{2+}$ and $Cu^{2+}$.

• 한국염색가공학회:학술대회논문집
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• 한국염색가공학회 2010년도 제3회 국제학회
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• pp.132-132
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• 2010