WHAT IS COVID-19

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WHAT IS COVID-19

A novel coronavirus (COVID-19 virus) outbreak has caused a global pandemic resulting in tens of thousands of infections and thousands of deaths worldwide. The RNA-dependent RNA polymerase (RdRp, also named nsp12) is the central component of coronaviral replication/transcription machinery and appears to be a primary target for the antiviral drug, remdesivir. We report the cryo-EM structure of COVID-19 virus full-length nsp12 in complex with cofactors nsp7 and nsp8 at 2.9-Å resolution. In addition to the conserved architecture of the polymerase core of the viral polymerase family, nsp12 possesses a newly identified β-hairpin domain at its N terminus. A comparative analysis model shows how remdesivir binds to this polymerase. The structure provides a basis for the design of new antiviral therapeutics targeting viral RdRp.

Corona Virus Disease 2019 (COVID-19) caused by a novel coronavirus emerged in December 2019 and has since become a global pandemic. COVID-19 virus is reported to be a new member of the betacoronavirus genus and is closely related to severe acute respiratory syndrome coronavirus (SARS-CoV) and to several bat coronaviruses . Compared to SARS-CoV and MERS-CoV, COVID-19 virus exhibits faster human-to-human transmission, thus leading to the WHO declaration of a world-wide public health emergency  

CoVs employ a multi-subunit replication/transcription machinery. A set of non-structural proteins (nsp) produced as cleavage products of the ORF1a and ORF1ab viral polyproteins (assemble to facilitate viral replication and transcription. A key component, the RNA-dependent RNA polymerase (RdRp, also known as nsp12), catalyzes the synthesis of viral RNA and thus plays a central role in the replication and transcription cycle of COVID-19 virus, possibly with the assistance of nsp7 and nsp8 as co-factors Nsp12 is therefore considered a primary target for nucleotide analog antiviral inhibitors such as remdesivir, which shows potential for the treatment of COVID-19 viral infections To inform drug design we have determined the structure of nsp12, in complex with its cofactors nsp7 and nsp8 by cryo-Electron Microscopy (Cryo-EM) using two different protocols, one in the absence of DTT (Dataset-1) and the other in the presence of DTT (Dataset-2).

The bacterially expressed full-length COVID-19 virus nsp12 (residues S1-Q932) was incubated with nsp7 (residues S1-Q83) and nsp8 (residues A1-Q198), and the complex was then purified . Cryo-EM grids were prepared using this complex and preliminary screening revealed excellent particle density with good dispersion. After the collection and processing of 7,994 micrograph movies, we obtained a 2.9-Å resolution 3D reconstruction of an nsp12 monomer in complex with one nsp7-nsp8 pair and an nsp8 monomer, as was previously observed for SARS-CoV . In addition to the nsp12-nsp7-nsp8 complex, we also observed single particle classes corresponding to the nsp12-nsp8 dimer, as well as individual nsp12 monomers, but these do not give atomic resolution reconstructions. However, the nsp12-nsp7-nsp8 complex reconstruction provides the structural information for complete structural analysis