Highly similar structural frames link the template tunnel and NTP entry tunnel to the exterior surface in RNA-dependent RNA polymerases

Abstract

RNA-dependent RNA polymerase (RdRp) is essential to viral replication and is therefore one of the primary targets of countermeasures against these dangerous infectious agents. Development of broad-spectrum therapeutics targeting polymerases has been hampered by the extreme sequence variability of these sequences. RdRps range in length from 400–800 residues, yet contain only ∼20 residues that are conserved in most species. In this study, we made structure-based comparisons that are independent of sequence composition using a recently developed algorithm. We identified residue-to-residue correspondences of multiple protein structures and created (two-dimensional) structure-based alignment maps of 37 polymerase structures that provide both sequence and structure details. Using these maps, we determined that ∼75% of each polymerase species consists of seven protein segments, each of which has high structural similarity to segments in other species, though they are widely divergent in sequence composition and order. We define each of these segments as a ‘homomorph’, and each includes (though most are much larger than) the well-known conserved polymerase motifs. All homomorphs contact the template tunnel or nucleoside triphosphate (NTP) entry tunnel and the exterior of the protein, suggesting they constitute a structural and functional skeleton common among the polymerases.

Evolution of Tertiary Structure of Viral RNA Dependent Polymerases

“…Features were selected and quantified manually according to criteria previously used for describing vRdPs [20], [24], [42] and are included in the Text S1.…”

“…DALI created structure based sequence alignment was validated and improved using the default settings in T-Coffee Expresso [35]. The resulting alignment was verified by comparison with previously published vRdP alignments [17], [24], [31], [36], [37].…”

Homology-Based Identification of a Mutation in the Coronavirus RNA-Dependent RNA Polymerase That Confers Resistance to Multiple Mutagens

“…In most cases, altered RdRp fidelity decreases fitness relative to wild-type (WT) viruses; this has been demonstrated for changes as small as a 1.2-fold difference in the accumulation of mutations (12,(14)(15)(16). Despite having as little as no amino acid identity outside conserved motifs (11)(12)(13)(14)(17)(18)(19), all described polymerase structures (including RdRps) resemble a "cupped right hand," with finger, palm, and thumb domains (20). The fingers form a channel that allows entry of the template RNA and ribonucleotide triphosphates (rNTPs) and assist in proper positioning of incoming nucleotides in the active site (21).…”

Vaccine-derived Mutation in Motif D of Poliovirus RNA-dependent RNA Polymerase Lowers Nucleotide Incorporation Fidelity

“…Most of the catalytic machinery resides in the palm region, which can be further divided into five major structural motifs (A-E). Motifs A and C contain absolutely conserved Asp residues critical for binding Mg 2ϩ ions required for RdRp function, and conserved residues in motif B are important for interacting with the sugar of the incoming nucleoside triphosphate (NTP) (15)(16)(17)(18)(19)(20)(21)(22). Structures of RdRps bound with RNA and/or nucleotide have highlighted structural rearrangements within these palm motifs that are necessary before nucleotide incorporation (23)(24)(25)(26)(27).…”