Avoiding Regions Symptomatic of Conformational and Functional Flexibility to Identify Antiviral Targets in Current and Future Coronaviruses

Abstract

Within the last 15 years, two related coronaviruses (Severe Acute Respiratory Syndrome [SARS]-CoV and Middle East Respiratory Syndrome [MERS]-CoV) expanded their host range to include humans, with increased virulence in their new host. Coronaviruses were recently found to have little intrinsic disorder compared with many other virus families. Because intrinsically disordered regions have been proposed to be important for rewiring interactions between virus and host, we investigated the conservation of intrinsic disorder and secondary structure in coronaviruses in an evolutionary context. We found that regions of intrinsic disorder are rarely conserved among different coronavirus protein families, with the primary exception of the nucleocapsid. Also, secondary structure predictions are only conserved across 50–80% of sites for most protein families, with the implication that 20–50% of sites do not have conserved secondary structure prediction. Furthermore, nonconserved structure sites are significantly less constrained in sequence divergence than either sites conserved in the secondary structure or sites conserved in loop. Avoiding regions symptomatic of conformational flexibility such as disordered sites and sites with nonconserved secondary structure to identify potential broad-specificity antiviral targets, only one sequence motif (five residues or longer) remains from the >10,000 starting sites across all coronaviruses in this study. The identified sequence motif is found within the nonstructural protein (NSP) 12 and constitutes an antiviral target potentially effective against the present day and future coronaviruses. On shorter evolutionary timescales, the SARS and MERS clades have more sequence motifs fulfilling the criteria applied. Interestingly, many motifs map to NSP12 making this a prime target for coronavirus antivirals.

Potential RNA-dependent RNA polymerase inhibitors as prospective therapeutics against SARS-CoV-2

“…These targets represent potential drug-binding sites located in protein regions crucial for viral fitness and, therefore, conserved on long evolutionary time scales. Our study identified two to four such targets in the protease (NSP5), replicase (NSP7), helicase (NSP13) and exonuclease (NSP14), with nine targets identified in the RdRP (NSP12) [7].…”

“…A multiple sequence alignment of RdRP from SARS-CoV-2 (accession number: YP_009724389.1, range 4398-5324) and the RdRP sequences previously used to determine the target motifs [7] was constructed using mafft with L+INS+ i in Jalview [12]. The multiple sequence alignment was used to build a phylogenetic tree using PhyML with 100 bootstraps [13].…”

“…The positive-strand RNA genome of a coronavirus such as SARS-CoV and MERS-CoV encodes ~25 different protein products [7]. Four of these proteins have a structural function and assemble to form the capsid that encapsulates the viral genome.…”

“…If the key proteins in the replicase-transcriptase complex can be prevented from replicating the RNA genome, new virions will not form. Recently we identified regions in the key proteins in MERS-CoV and SARS-CoV that could serve as targets for antiviral drugs [7]. We took an evolutionary approach to identify regions of at least five consecutive sites not just conserved in sequence but also in structural properties with low conformational flexibility to reduce the risk of a target being displayed for only a fraction of the time [7].…”

“…Recently we identified regions in the key proteins in MERS-CoV and SARS-CoV that could serve as targets for antiviral drugs [7]. We took an evolutionary approach to identify regions of at least five consecutive sites not just conserved in sequence but also in structural properties with low conformational flexibility to reduce the risk of a target being displayed for only a fraction of the time [7]. These targets represent potential drug-binding sites located in protein regions crucial for viral fitness and, therefore, conserved on long evolutionary time scales.…”