TY  - JOUR
A1  - Kokic, Goran
A1  - Hillen, Hauke S.
A1  - Tegunov, Dimitry
A1  - Dienermann, Christian
A1  - Seitz, Florian
A1  - Schmitzova, Jana
A1  - Farnung, Lucas
A1  - Siewert, Aaron
A1  - Höbartner, Claudia
A1  - Cramer, Patrick
T1  - Mechanism of SARS-CoV-2 polymerase stalling by remdesivir
JF  - Nature Communications
N2  - Remdesivir is the only FDA-approved drug for the treatment of COVID-19 patients. The active form of remdesivir acts as a nucleoside analog and inhibits the RNA-dependent RNA polymerase (RdRp) of coronaviruses including SARS-CoV-2. Remdesivir is incorporated by the RdRp into the growing RNA product and allows for addition of three more nucleotides before RNA synthesis stalls. Here we use synthetic RNA chemistry, biochemistry and cryoelectron microscopy to establish the molecular mechanism of remdesivir-induced RdRp stalling. We show that addition of the fourth nucleotide following remdesivir incorporation into the RNA product is impaired by a barrier to further RNA translocation. This translocation barrier causes retention of the RNA 3ʹ-nucleotide in the substrate-binding site of the RdRp and interferes with entry of the next nucleoside triphosphate, thereby stalling RdRp. In the structure of the remdesivir-stalled state, the 3ʹ-nucleotide of the RNA product is matched and located with the template base in the active center, and this may impair proofreading by the viral 3ʹ-exonuclease. These mechanistic insights should facilitate the quest for improved antivirals that target coronavirus replication.
KW  - SARS-CoV-2 polymerase
KW  - Remdesivir
KW  - RNA-dependent RNA polymerase
KW  - Molecular mechanism
KW  - Biochemistry
KW  - Cryoelectron microscopy
KW  - RNA
Y1  - 2021
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-220979
VL  - 12
ER  - 
TY  - JOUR
A1  - Kabinger, Florian
A1  - Stiller, Carina
A1  - Schmitzová, Jana
A1  - Dienemann, Christian
A1  - Kokic, Goran
A1  - Hillen, Hauke S.
A1  - Höbartner, Claudia
A1  - Cramer, Patrick
T1  - Mechanism of molnupiravir-induced SARS-CoV-2 mutagenesis
JF  - Nature Structural & Molecular Biology
N2  - Molnupiravir is an orally available antiviral drug candidate currently in phase III trials for the treatment of patients with COVID-19. Molnupiravir increases the frequency of viral RNA mutations and impairs SARS-CoV-2 replication in animal models and in humans. Here, we establish the molecular mechanisms underlying molnupiravir-induced RNA mutagenesis by the viral RNA-dependent RNA polymerase (RdRp). Biochemical assays show that the RdRp uses the active form of molnupiravir, β-d-\(N^4\)-hydroxycytidine (NHC) triphosphate, as a substrate instead of cytidine triphosphate or uridine triphosphate. When the RdRp uses the resulting RNA as a template, NHC directs incorporation of either G or A, leading to mutated RNA products. Structural analysis of RdRp–RNA complexes that contain mutagenesis products shows that NHC can form stable base pairs with either G or A in the RdRp active center, explaining how the polymerase escapes proofreading and synthesizes mutated RNA. This two-step mutagenesis mechanism probably applies to various viral polymerases and can explain the broad-spectrum antiviral activity of molnupiravir.
KW  - Molnupiravir
KW  - RNA-Dependent RNA Polymerase
KW  - SARS-CoV2 Replication Impairment
KW  - Molnupiravir-Induced RNA Mutagenesis Mechanism
KW  - Cryoelectron Microscopy
Y1  - 2021
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-254603
VL  - 28
ER  -