No abstract

During the RNA replication, coronaviruses require proofreading to maintain the integrity of their large genomes. Nsp14 associates with viral polymerase complex to excise the mismatched nucleotides. Aside from the exonuclease activity, nsp14 methyltransferase domain mediates cap methylation, facilitating translation initiation and protecting viral RNA from recognition by the innate immune sensors. The nsp14 exonuclease activity is modulated by a protein co-factor nsp10. While the nsp10/nsp14 complex structure is available, the mechanistic basis for nsp10 mediated modulation remains unclear in the absence of nsp14 structure. Here we provide a crystal structure of nsp14 in an apo-form. Comparative analysis of the apo- and nsp10 bound structures explain the modulatory role of the co-factor protein. Further, the structure presented in this study rationalizes the recently proposed idea of nsp14/nsp10/nsp16 ternary complex.

Coronaviruses protect their single-stranded RNA genome with the methylated cap added during the replication. This capping process is carried out by several nonstructural proteins (nsp) encoded in the viral genome. The methylation itself is performed consecutively by two methyltransferases, nsp14 and nsp16, which interact with nsp10 protein acting as a co-factor. The nsp14 protein also carries the exonuclease domain, which also serves as a part of the proofreading system during the replication of the large RNA genome. The available crystal structures suggest that the concomitant interaction between these three proteins is impossible due to the structural clash, and it is generally accepted that the nsp16 and nsp14 bind with the nsp10 separately. Here, we show that nsp14, nsp10, and nsp16 form a methylation complex despite the odds. Due to spatial proximity, this interaction is beneficial for forming mature capped viral mRNA. Further, it modulates the exonuclease activity of nsp14, protecting the viral RNA at the replication site. Our findings show that nsp14 is more amenable to allosteric regulation and may serve as a molecular target for the therapy.