Motif D of Viral RNA-Dependent RNA Polymerases Determines Efficiency and Fidelity of Nucleotide Addition

• DOI: 10.1016/j.str.2012.06.012
• Publication Date: 2012-07-19T17:17:54Z
• Authors: Xiaorong Yang; Eric D. Smidansky; Kenneth R. Maksim...; David Lum; Jesse L. Welch; Jamie J. Arnold; Craig E. Cameron; David D. Boehr
• Keywords: Models, Molecular; 0303 health sciences; Magnetic Resonance Spectroscopy; Base Sequence; Nucleotides; Amino Acid Motifs; Hydrogen Bonding; RNA-Dependent RNA Polymerase; Poliovirus; Viral Proteins; 03 medical and health sciences; Structural Biology; Catalytic Domain; Biocatalysis; Molecular Biology; Base Pairing; Protein Binding

Fast, accurate nucleotide incorporation by polymerases facilitates expression and maintenance of genomes. Many polymerases use conformational dynamics of a conserved α helix to permit efficient nucleotide addition only when the correct nucleotide substrate is bound. This α helix is missing in structures of RNA-dependent RNA polymerases (RdRps) and RTs. Here, we use solution-state nuclear magnetic resonance to demonstrate that the conformation of conserved structural motif D of an RdRp is linked to the nature (correct versus incorrect) of the bound nucleotide and the protonation state of a conserved, motif-D lysine. Structural data also reveal the inability of motif D to achieve its optimal conformation after incorporation of an incorrect nucleotide. Functional data are consistent with the conformational change of motif D becoming rate limiting during and after nucleotide misincorporation. We conclude that motif D of RdRps and, by inference, RTs is the functional equivalent to the fidelity helix of other polymerases.