NSP3 polyprotein of SARS-CoV2: structural studies and its exploration in drug screening campaign – COVNSP3

Severe Acute Respiratory syndrome (SARS) is an alarming emerging infectious disease caused by coronavirus (CoV). The current COVID-19 pandemic, which originated in the Wuhan region of China, is caused by a novel coronavirus, SARS-CoV2 which is more infectious than the 2003 SARS-CoV and which has led to the partial or total confinement of the populations of several countries, including France. In order to address the current, and potential future outbreaks of COVID-19, it is extremely important to both improve our understanding of the novel virus and to develop potential therapeutic agents. Here, we propose the largest polyprotein of SARS-CoV2, non-structural protein 3 (Nsp3) as an important therapeutic target. Nsp3 is a polyprotein precursor on which 15 individual proteins crucial to the function and life cycle of the virus are linked together before autocleavage of the chain to produce individual proteins. Although some current potential drugs likely target some of these individual proteins, a more efficient strategy may be to stop the virus in its tracks by targeting the pre-cleaved precursor. Elucidation of the atomic structure of Nsp3 will be crucial to such a strategy, revealing new drug binding sites thus enhancing the possibility to inhibit the ability of the virus to produce the individual proteins that are crucial to its life cycle and infectivity. The aims of this proposal are to reveal, at close to atomic resolution, the three-dimensional structures of full length Nsp3 and of its cytoplasmic region, and to pursue the first steps towards identifying potential therapeutic molecules. The project will exploit an integrated, interdisciplinary and multi-level approach, involving the recombinant production and purification of the Nsp3 targets coupled with cryo-electron microscopy, X-ray crystallography and ligand/fragment screening campaigns. Polyprotein production will be carried out using a single chain technology in which the individual domains of Nsp3 will be tethered covalently with modified protease sites. This methodology will also be used to identify the stable core of Nsp3, which will also be used in structural studies and fragment screening protocols. The project proposed here thus not only has a direct impact on tackling the present COVID-19 pandemic by identifying new therapeutic targets, it will also open up new avenues for the study any viral polyprotein and/or multi-protein complexes.