A Covalent Chemical Probe for NsP2 Cysteine Protease with Antialphaviral Activity and Proteome-wide Selectivity

Overview

Journal Res Sq

Date 2024 Nov 28

PMID 39606462

Authors

Anirban Ghoshal

Edwin G Tse

Mohammad Anwar Hossain

Kesatebrhan Haile Asressu

Eric M Merten

John D Sears

Stefanie Howell

Sumera Perveen

Jane Burdick

Noah L Morales

Sabian A Martinez

Isabella Law

Bennett J Davenport

Thomas E Morrison

Zachary J Streblow

Daniel N Streblow

Angie L Mordant

Thomas S Webb

Aurora Cabrera

Laura E Herring

Cheryl H Arrowsmith

Kenneth H Pearce

Nathaniel J Moorman

Mark T Heise

Rafael Miguez Counago

Peter J Brown

Timothy M Willson

Affiliations

Soon will be listed here.

Abstract

RA-0003022 () was identified as a high-quality covalent chemical probe for nsP2 cysteine protease (nsP2pro). Isoxazole covalently captured the active site C478 and inactivated the enzyme with a / ratio of 6000 Ms. A negative control analog RA-0025453 () retained the covalent warhead but demonstrated >100-fold decrease in enzyme inhibition. Isoxazoles and were stable across a wide range of pH in solution and upon prolonged storage as solids. The covalent chemical probe was inactive across a panel of 23 human and viral cysteine proteases and demonstrated remarkable proteome-wide selectivity by two chemoproteomic methods. Isoxazole reduced viral titer against infectious isolates of , , and Venezuelan Equine Encephalitis viruses demonstrating its activity on both New and Old World alphaviruses. Isoxazole and its negative control will find utility as covalent chemical probes to study the role of the nsP2pro in alphaviral replication and virulence.

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