The intrinsically disordered SARS-CoV-2 nucleoprotein in dynamic complex with its viral partner nsp3a.

Luiza Mamigonian Bessa,Anas Malki,Maiia Botova,Laura Mariño Perez,Nicola Salvi,Malene Ringkjøbing Jensen,Max Nanao,Rob W H Ruigrok,Aldo R Camacho-Zarco,Serafima Guseva,Martin Blackledge,Damien Maurin

doi:10.1126/sciadv.abm4034

Abstract

The processes of genome replication and transcription of SARS-CoV-2 represent important targets for viral inhibition. Betacoronaviral nucleoprotein (N) is a highly dynamic cofactor of the replication-transcription complex (RTC), whose function depends on an essential interaction with the amino-terminal ubiquitin-like domain of nsp3 (Ubl1). Here, we describe this complex (dissociation constant - 30 to 200 nM) at atomic resolution. The interaction implicates two linear motifs in the intrinsically disordered linker domain (N3), a hydrophobic helix (219LALLLLDRLNQL230) and a disordered polar strand (243GQTVTKKSAAEAS255), that mutually engage to form a bipartite interaction, folding N3 around Ubl1. This results in substantial collapse in the dimensions of dimeric N, forming a highly compact molecular chaperone, that regulates binding to RNA, suggesting a key role of nsp3 in the association of N to the RTC. The identification of distinct linear motifs that mediate an important interaction between essential viral factors provides future targets for development of innovative strategies against COVID-19.

Highlights

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the origin of the respiratory coronavirus disease 2019 (COVID-19) pandemic, is a member of the betacoronavirus genus, including Middle East respiratory syndrome (MERS)–CoV and SARS-CoV
Betacoronaviral replication- transcription complexes (RTCs) are associated with membrane networks in the form of viral replication organelles called double- membrane vesicles (DMVs) constituted from membranes participating in the host secretory pathway [4]
Very similar entropic and enthalpic contributions are measured for the complexes comprising constructs containing sN3 and Ubl1, suggesting that the interaction site present in sN3 dominates the thermodynamics of complex formation

Summary

Introduction

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the origin of the respiratory coronavirus disease 2019 (COVID-19) pandemic, is a member of the betacoronavirus genus, including Middle East respiratory syndrome (MERS)–CoV and SARS-CoV. The processes of replication and transcription of viral RNA represent important targets for viral inhibition, and the development of rational strategies to achieve this end requires a molecular understanding of the viral replication cycle. Coronaviridae are enveloped positive-sense single-strand RNA viruses that express their own replication machinery. Replication of the SARS-CoV-2 genome is carried out by the RNA-dependent RNA polymerase complex, whose functional modes have been investigated by cryo–electron microscopy [1,2,3]. Betacoronaviral replication- transcription complexes (RTCs) are associated with membrane networks in the form of viral replication organelles called double- membrane vesicles (DMVs) constituted from membranes participating in the host secretory pathway [4]. DMVs have been shown to constitute active sites of viral RNA synthesis [5, 6]

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