Mutations in the monkeypox virus replication complex: Potential contributing factors to the 2022 outbreak

Attributes contributing to the current monkeypox virus (MPXV) outbreak remain unknown. It has been estab-lished that mutations in viral proteins may alter phenotype and pathogenicity. To assess if mutations in the MPXV DNA replication complex (RC) contribute to the outbreak, we conducted a temporal analysis of available MPXV sequences to identify mutations, generated a DNA replication complex (RC) using structures of related viral and eukaryotic proteins, and structure prediction method AlphaFold. Ten mutations within the RC were identified and mapped onto the RC to infer role of mutations. Two mutations in F8L (RC catalytic subunit), and two in G9R (a processivity factor) were-100% prevalent in the 2022 sequences. F8L mutation L108F emerged in 2022, whereas W411L emerged in 2018, and persisted in 2022. L108 is topologically located to enhance DNA binding affinity of F8L. Therefore, mutation L108F can change the fidelity, sensitivity to nucleoside inhibitors, and processivity of F8L. Surface exposed W411L likely affects the binding of regulatory factor(s). G9R mutations S30L and D88 N in G9R emerged in 2022, and may impact the interaction of G9R with E4R (uracil DNA gly-cosylase). The remaining six mutations that appeared in 2001, reverted to the first (1965 Rotterdam) isolate. Two nucleoside inhibitors brincidofovir and cidofovir have been approved for MPXV treatment. Cidofovir resistance in vaccinia virus is achieved by A314T and A684V mutations. Both A314 and A684 are conserved in MPXV. Therefore, resistance to these drugs in MPXV may arise through similar mechanisms.