David Dulin
Assistant Professor, Vrije Universiteit Amsterdam (The Netherlands) and Friedrich-Alexander University Erlangen-Nuremberg (Germany)
https://daviddulinlab.com/
“Mechanochemistry and drug targeting of the SARS-CoV-2 replication-transcription complex from a single molecule perspective”
Subhas C. Bera, Mona Seifert, Pim A. A. America, Pauline van Nies, Flávia S. Papini, Jay Marecky, Robert N. Kirchdoerfer, Irina Artsimovitch, Bruno Canard, Jamie J. Arnold, Yan Xiang, Martin Depken, Kevin Raney, Craig E. Cameron, and David Dulin
The SARS-CoV-2 pandemic has been and still is a terrible burden for our societies. While the vaccine response has been incredibly fast, drugs development is still in its infancy, and lack to cure patients infected by SARS-CoV-2, or by future coronavirus outbreaks. The main antiviral drug target is the replication-transcription complex (RTC), a conserved large multiprotein machinery in charge of replicating and transcribing the ~30 kb long positive single stranded (ss) RNA coronavirus genome. The RTC is made of a core RTC formed of the RNA dependent RNA polymerase (RdRp) nsp12, and the associated factors nsp7 and nsp8, in a stoichiometry 1:1:2, to which associates many factors, such as the helicase nsp13 and the exonuclease nsp14. While the cryoEM revolution has revealed the structure of many partial RTCs, functional studies have lagged behind, shadowing our knowledge on the structure-function relationship of these RTCs, and delaying the finding drug targets and novel antiviral drugs. To fill this gap, we have pioneered a high throughput magnetic tweezers assay to monitor the elongation dynamics of the SARS-CoV-2 RTC over 1 kilobase long template at near single base resolution in the presence of all NTPs. Here, I will present how we applied this assay to reveal the nucleotide addition cycle of SARS-CoV-2 core RTC (Bera, Seifert et al., Cell Reports 2021), and the mechanism of action of several nucleotide analogs, such as Remdesivir and ddhC (Seifert, Bera et al., eLife 2021). I will also present our latest investigations of the core RTC mechanochemistry when including other viral co-factors.