Motif D of viral RNA-dependent RNA polymerases determines efficiency and fidelity of nucleotide addition

Structure. 2012 Sep 5;20(9):1519-27. doi: 10.1016/j.str.2012.06.012. Epub 2012 Jul 19.

Abstract

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.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Motifs
  • Base Pairing
  • Base Sequence
  • Biocatalysis
  • Catalytic Domain
  • Hydrogen Bonding
  • Magnetic Resonance Spectroscopy
  • Models, Molecular
  • Nucleotides / chemistry*
  • Poliovirus / enzymology*
  • Protein Binding
  • RNA Replicase / chemistry*
  • Viral Proteins

Substances

  • Nucleotides
  • Viral Proteins
  • RNA Replicase