A mechanism for initiating RNA-dependent RNA polymerization

Nature. 2001 Mar 8;410(6825):235-40. doi: 10.1038/35065653.


In most RNA viruses, genome replication and transcription are catalysed by a viral RNA-dependent RNA polymerase. Double-stranded RNA viruses perform these operations in a capsid (the polymerase complex), using an enzyme that can read both single- and double-stranded RNA. Structures have been solved for such viral capsids, but they do not resolve the polymerase subunits in any detail. Here we show that the 2 A resolution X-ray structure of the active polymerase subunit from the double-stranded RNA bacteriophage straight phi6 is highly similar to that of the polymerase of hepatitis C virus, providing an evolutionary link between double-stranded RNA viruses and flaviviruses. By crystal soaking and co-crystallization, we determined a number of other structures, including complexes with oligonucleotide and/or nucleoside triphosphates (NTPs), that suggest a mechanism by which the incoming double-stranded RNA is opened up to feed the template through to the active site, while the substrates enter by another route. The template strand initially overshoots, locking into a specificity pocket, and then, in the presence of cognate NTPs, reverses to form the initiation complex; this process engages two NTPs, one of which acts with the carboxy-terminal domain of the protein to prime the reaction. Our results provide a working model for the initiation of replication and transcription.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Bacteriophage phi 6 / enzymology*
  • Bacteriophage phi 6 / genetics
  • Crystallography, X-Ray
  • Escherichia coli
  • Hepacivirus / enzymology*
  • Hepacivirus / genetics
  • Magnesium / metabolism
  • Manganese / metabolism
  • Models, Molecular
  • Protein Conformation
  • RNA, Double-Stranded / metabolism
  • RNA, Viral / metabolism*
  • RNA-Directed DNA Polymerase / chemistry
  • RNA-Directed DNA Polymerase / metabolism*
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / metabolism
  • Templates, Genetic
  • Transcription, Genetic


  • RNA, Double-Stranded
  • RNA, Viral
  • Recombinant Proteins
  • Manganese
  • RNA-Directed DNA Polymerase
  • Magnesium

Associated data

  • PDB/1HHS
  • PDB/1HHT
  • PDB/1HI0
  • PDB/1HI1
  • PDB/1HI8