Application of a cell-based protease assay for testing inhibitors of picornavirus 3C proteases

doi:10.1016/j.antiviral.2013.12.012

	Application of a cell-based protease assay for testing inhibitors of picornavirus 3C proteases
	van der Linden, Lonneke 2,3,4; Ulferts, Rachel 2,3,5; Nabuurs, Sander B.6; Kusov, Yuri 1,7; Liu, Hong8 ; George, Shyla 1,7; Lacroix, Celine 4; Goris, Nesya 9; Lefebvre, David 10; Lanke, Kjerstin H. W.2,3
刊名	ANTIVIRAL RESEARCH
	2014-03
卷号	103 页码:17-24
关键词	Cell-based assay Protease 3C(pro) Picornavirus Inhibitor
ISSN号	0166-3542
DOI	10.1016/j.antiviral.2013.12.012
文献子类	Article
英文摘要	Proteolytical cleavage of the picornaviral polyprotein is essential for viral replication. Therefore, viral proteases are attractive targets for anti-viral therapy. Most assays available for testing proteolytical activity of proteases are performed in vitro, using heterologously expressed proteases and peptide substrates. To deal with the disadvantages associated with in vitro assays, we modified a cell-based protease assay for picornavirus proteases. The assay is based on the induction of expression of a firefly luciferase reporter by a chimeric transcription factor in which the viral protease and cleavage sites are inserted between the GAL4 binding domain and the VP16 activation domain. Firefly luciferase expression is dependent on cleavage of the transcription factor by the viral protease. This biosafe assay enables testing the effect of compounds on protease activity in cells while circumventing the need for infection. We designed the assay for 3C proteases (3C(pro)) of various enteroviruses as well as of viruses of several other picoma-virus genera, and show that the assay is amenable for use in a high-throughput setting. Furthermore, we show that the spectrum of activity of 3C(pro) inhibitor AG7088 (rupintrivir) not only encompasses enterovirus 3C(pro) but also 3C(pro) of foot-and-mouth disease virus (FMDV), an aphthovirus. In contrary, AG7404 (compound 1), an analogue of AG7088, had no effect on FMDV 3C(pro) activity, for which we provide a structural explanation. (C) 2013 Elsevier B.V. All rights reserved.
资助项目	"Convenant KU Leuven-Radboud University Nijmegen" framework (FJMvK, JN)[00000000] ; European Union[260644] ; NWO-VICI Grant[91812628] ; Chinese Academy of Sciences for a Visiting Professorship for Senior International Scientists[2010T1S6] ; KU Leuven geconcerteerde onderzoeksactie (JN)[GOA/10/014]
WOS关键词	MOUTH-DISEASE VIRUS ; STRUCTURE-BASED DESIGN ; VITRO ANTIVIRAL ACTIVITY ; IN-VITRO ; BIOLOGICAL EVALUATION ; 3C-LIKE PROTEASES ; LEADER PROTEIN ; CLEAVAGE ; REPLICATION ; TRANSLATION
WOS研究方向	Pharmacology & Pharmacy ; Virology
语种	英语
出版者	ELSEVIER SCIENCE BV
WOS记录号	WOS:000332821000003
内容类型	期刊论文
源URL	[http://119.78.100.183/handle/2S10ELR8/277167]
专题	药物化学研究室中科院受体结构与功能重点实验室新药研究国家重点实验室
通讯作者	van Kuppeveld, Frank J. M.
作者单位	1.Med Univ Lubeck, Inst Biochem, Ctr Struct & Cell Biol Med, D-23538 Lubeck, Germany; 2.Radboud Univ Nijmegen, Med Ctr, Nijmegen Ctr Mol Life Sci, Dept Med Microbiol, NL-6525 ED Nijmegen, Netherlands; 3.Radboud Univ Nijmegen, Med Ctr, Nijmegen Inst Infect Inflammat & Immun, NL-6525 ED Nijmegen, Netherlands; 4.Univ Louvain, Lab Virol & Chemotherapy, Rega Inst Med Res, Louvain, Belgium; 5.Univ Utrecht, Fac Vet Med, Dept Infect Dis & Immunol, Div Virol, NL-3508 TD Utrecht, Netherlands; 6.Radboud Univ Nijmegen, Med Ctr, Nijmegen Ctr Mol Life Sci, Ctr Mol & Biomol Informat, NL-6525 ED Nijmegen, Netherlands; 7.Med Univ Lubeck, German Ctr Infect Res DZIF, D-23538 Lubeck, Germany; 8.Chinese Acad Sci, Shanghai Inst Biol Sci, Shanghai Inst Mat Med, State Key Lab Drug Res,Grad Sch, Shanghai, Peoples R China; 9.Okapi Sci NV, Heverlee, Belgium; 10.Vet & Agrochem Res Ctr, Brussels, Belgium;
推荐引用方式 GB/T 7714	van der Linden, Lonneke,Ulferts, Rachel,Nabuurs, Sander B.,et al. Application of a cell-based protease assay for testing inhibitors of picornavirus 3C proteases[J]. ANTIVIRAL RESEARCH,2014,103:17-24.
APA	van der Linden, Lonneke.,Ulferts, Rachel.,Nabuurs, Sander B..,Kusov, Yuri.,Liu, Hong.,...&van Kuppeveld, Frank J. M..(2014).Application of a cell-based protease assay for testing inhibitors of picornavirus 3C proteases.ANTIVIRAL RESEARCH,103,17-24.
MLA	van der Linden, Lonneke,et al."Application of a cell-based protease assay for testing inhibitors of picornavirus 3C proteases".ANTIVIRAL RESEARCH 103(2014):17-24.