Structural and Phylogenetic Analysis on the Proofreading Activity of SARS-CoV-2

Maintenance of genomic integrity is a fundamental characteristic of viruses, however, RNA-dependent RNA Polymerase (RdRp) lacks exonuclease activity for proofreading. To facilitate genomic proofreading in viruses, an independent exonuclease domain assists RdRp to maintain fidelity during replication. In contrast to high fidelity in DNA viruses, RNA viruses have to evolve into new variants through comparatively delicate mutagenesis activity for genetic diversity. Coronavirideae, a family of single positive-stranded RNA (+ ssRNA) viruses, meticulously sustain a balance between genetic diversity and large-size RNA genome. In coronaviruses, the proofreading activity is accomplished by an exonuclease (ExoN) domain located at the N-terminal of non-structural protein 14 (nsp14). ExoN is responsible for the new variants and antiviral resistance towards nucleotide analogs. Here, we provide an evolutionary characterization of ExoN by using a well-defined phylogenetic pipeline and structural analysis based on host and habitat. We carried out a phylogenetic analysis on ExoN, methyltransferase domain, nsp14, and whole genomes of ExoN-containing viruses. Furthermore, a three-dimensional structural comparison of the ExoN domain from various sources is also carried out to understand structural preservation. Our study has unveiled the evolutionary trajectories and structural conservation of the ExoN domain within the Coronaviridae family, highlighting its distinct evolutionary path independent of other domains. Structural analyses revealed minimal variance in RMSD values, underscoring the conserved nature of ExoN despite diverse ecological settings.