首页|TMPRSS2 and SARS-CoV-2 SPIKE interaction assay for uHTS

TMPRSS2 and SARS-CoV-2 SPIKE interaction assay for uHTS

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SARS-CoV-2,the coronavirus that causes the disease COVID-19,has claimed millions of lives over the past 2 years.This demands rapid development of effective therapeutic agents that target various phases of the viral replication cycle.The interaction between host transmembrane serine protease 2(TMPRSS2)and viral SPIKE protein is an important initial step in SARS-CoV-2 infection,offering an opportunity for therapeutic development of viral entry inhibitors.Here,we report the development of a time-resolved fluorescence/Förster resonance energy transfer(TR-FRET)assay for monitoring the TMPRSS2-SPIKE interaction in lysate from cells co-expressing these proteins.The assay was configured in a 384-well-plate format for high-throughput screening with robust assay performance.To enable large-scale compound screening,we further miniaturized the assay into 1536-well ultrahigh-throughput screening(uHTS)format.A pilot screen demonstrated the utilization of the assay for uHTS.Our optimized TR-FRET uHTS assay provides an enabling platform for expanded screening campaigns to discover new classes of small-molecule inhibitors that target the SPIKE and TMPRSS2 protein-protein interaction.

SARS-CoV-2COVID-19protein-protein interactionTMPRSS2SPIKEhigh-throughput screeningTR-FRET

Danielle Cicka、Qiankun Niu、Min Qui、Kun Qian、Eric Miller、Dacheng Fan、Xiulei Mo、Andrey A.Ivanov、Stefan G.Sarafianos、Yuhong Du、Haian Fu

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Department of Pharmacology and Chemical Biology,Emory University School of Medicine,Atlanta,GA 30322,USA

Graduate Program in Molecular and Systems Pharmacology,Laney Graduate School of Emory University,Atlanta,GA 30322,USA

Emory Chemical Biology Discovery Center,Emory University School of Medicine,Atlanta,GA 30322,USA

Winship Cancer Institute of Emory University,Atlanta,GA 30322,USA

Laboratory of Biochemical Pharmacology,Department of Pediatrics,Emory University School of Medicine,Atlanta,GA 30322,USA

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Emory School of Medicine COVID Catalyst-I3 awardNCI Emory Lung Cancer SPORE Career Enhancement Program(A.A.I.)Emory Initiative on Biological Discovery through Chemical Innovation(A.A.I.)R01AI167356(S.G.S.)

P50CA217691

2023

10.1093/jmcb/mjad017

分子细胞生物学报(英文版)
中国科学院上海生命科学研究院,生物化学与细胞生物学研究所,中国细胞生物学学会

分子细胞生物学报(英文版)

CSTPCDCSCD
影响因子：0.595
ISSN：1673-520X
年,卷(期)：2023.15(3)
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