Small-molecule-mediated rescue of mutant viral polymerases

Sammanfattning

Live attenuated vaccines are currently our greatest weapon in the fight against infectious disease, but they come with the downsides of severe infections in sick individuals and potential reversion of the vaccine strain to a virulent state. In order to further improve the safety of these vaccines and the trust that the community has in them, the downsides need to be minimized. We propose that the concept of synthetic auxotrophy can be applied to viral polymerases, thereby making the virus dependent on a small molecule for replication. Towards a proof-of-concept, we mutated tryptophan residues within a viral polymerase with the aims of inactivating the enzymatic activity, and then rescuing activity using indole. Here I establish transcription assays in vitro and in vivo to study the activities of SARS-COV2 RNA-dependent RNA polymerase and bacteriophage T7 RNA polymerase expressed in Escherichia coli. Although all preparations of RNA-dependent RNA polymerase appeared to be inactive, preliminary results showed active T7 RNA polymerase that was inactivated by our mutagenesis. This provides a starting point for potential rescue by small molecules.