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SARS-CoV-2 proteins

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    High-throughput fluorescent assay for inhibitor screening of proteases from RNA viruses

    Authors: Bara Cihlova; Andrea Huskova; Jiri Böserle; Radim Nencka; Evzen Boura; Jan Silhan; Tsuguhisa Nakayama; Jayakar V. Nayak; Peter K. Jackson; Raul Andino; Xiaokun Shu; Nicholas A. Crossland; Christopher S. Chen; Darrell N. Kotton; Susan C. Baker; John H. Connor; Florian Douam; Andrew Emili; Mohsan Saeed; Ilayda Sahin; Cavit Kerem Kayhan; Fatma Tokat; Gurler Akpinar; Murat Kasap; Ayse Sesin Kocagoz; Ugur Ozbek; Dilek Telci; Fikrettin Sahin; Koray Yalcin; Siret Ratip; Umit Ince; Guldal Suyen; Ercument Ovali; Liam Fergusson; Marta Conti; Marius Rameil; Vanessa Nakonecnij; Jakob Vanhoefer; Leonard Schmiester; Muying Wang; Emily E Ackerman; Jason E Shoemaker; Jeremy Zucker; Kristie L Oxford; Jeremy Teuton; Ebru Kocakaya; Gokce Yagmur Summak; Kristina Hanspers; Martina Kutmon; Susan Coort; Lars Eijssen; Friederike Ehrhart; Rex D. A. B.; Denise Slenter; Marvin Martens; Robin Haw; Bijay Jassal; Lisa Matthews; Marija Orlic-Milacic; Andrea Senff-Ribeiro; Karen Rothfels; Veronica Shamovsky; Ralf Stephan; Cristoffer Sevilla; Thawfeek Mohamed Varusai; Jean-Marie Ravel; Vera Ortseifen; Silvia Marchesi; Piotr Gawron; Ewa Smula; Laurent Heirendt; Venkata Satagopam; Guanming Wu; Anders Riutta; Martin Golebiewski; Stuart Owen; Carole Goble; Xiaoming Hu; Rupert Overall; Dieter Maier; Angela Bauch; John A Bachman; Benjamin M Gyori; Carlos Vega; Valentin Groues; Miguel Vazquez; Pablo Porras; Luana Licata; Marta Iannuccelli; Francesca Sacco; Denes Turei; Augustin Luna; Ozgun Babur; Sylvain Soliman; Alberto Valdeolivas; Marina Esteban-Medina; Maria Pena-Chilet; Tomas Helikar; Bhanwar Lal Puniya; Anastasia Nesterova; Anton Yuryev; Anita de Waard; Dezso Modos; Agatha Treveil; Marton Laszlo Olbei; Bertrand De Meulder; Aurelien Naldi; Aurelien Dugourd; Laurence Calzone; Chris Sander; Emek Demir; Tamas Korcsmaros; Tom C Freeman; Franck Auge; Jacques S Beckmann; Jan Hasenauer; Olaf Wolkenhauer; Egon Willighagen; Alexander R Pico; Chris Evelo; Lincoln D Stein; Henning Hermjakob; Julio Saez-Rodriguez; Joaquin Dopazo; Alfonso Valencia; Hiroaki Kitano; Emmanuel Barillot; Charles Auffray; Rudi Balling; Reinhard Schneider; - the COVID-19 Disease Map Community

    doi:10.1101/2020.10.27.357418 Date: 2020-10-28 Source: bioRxiv

    Spanish flu and other influenza outbreaks, the recent Zika epidemics, and the ongoing COVID-19 MESHD COVID-19 MESHD pandemic are the most profound examples of severe widespread diseases that are caused by RNA viruses. Perhaps less well-known yet dangerous RNA viruses cause deadly diseases such as polio, Ebola, measles, rubella MESHD, yellow fever MESHD, dengue fever MESHD, etc. To combat a particular viral disease by diminishing its spread and number of fatal cases, effective vaccines and antivirals are indispensable. Therefore, quick access to the means of discovery of new treatments for any epidemic outbreak is of great interest and in vitro, biochemical assays are the basis of drug discovery. The recent outbreak of the coronavirus pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) demands an affordable and reliable assay for testing antivirals. Here, we developed a quick and inexpensive high-throughput fluorescent assay to test inhibitors of viral proteases. Accordingly, we employed this assay to sample inhibitors for papain-like protease PROTEIN from SARS-CoV-2. In addition, we validated this assay for screening inhibitors of flaviviral protease from the tick-borne encephalitis MESHD virus to emphasize a broad range of applications of our approach. This fluorescent high-throughput assay is based on fluorescent energy transfer (FRET) between two distinct fluorescent proteins (eGFP and mCherry) connected via a substrate polypeptide. When the substrate is cleaved, FRET is abolished and the change in fluorescence corresponds to reaction progress. Our data show that this assay can be used for testing the inhibitors in the 96 or 384 well plates format with robust and reproducible outcomes.

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