Crystallographic and modeling studies of RNase III suggest a mechanism for double-stranded RNA cleavage

Structure. 2001 Dec;9(12):1225-36. doi: 10.1016/s0969-2126(01)00685-2.


Background: Aquifex aeolicus Ribonuclease III (Aa-RNase III) belongs to the family of Mg(2+)-dependent endonucleases that show specificity for double-stranded RNA (dsRNA). RNase III is conserved in all known bacteria and eukaryotes and has 1-2 copies of a 9-residue consensus sequence, known as the RNase III signature motif. The bacterial RNase III proteins are the simplest, consisting of two domains: an N-terminal endonuclease domain, followed by a double-stranded RNA binding domain (dsRBD). The three-dimensional structure of the dsRBD in Escherichia coli RNase III has been elucidated; no structural information is available for the endonuclease domain of any RNase III.

Results: We present the crystal structures of the Aa-RNase III endonuclease domain in its ligand-free form and in complex with Mn(2+). The structures reveal a novel protein fold and suggest a mechanism for dsRNA cleavage. On the basis of structural, genetic, and biological data, we have constructed a hypothetical model of Aa-RNase III in complex with dsRNA and Mg(2+) ion, which provides the first glimpse of RNase III in action.

Conclusions: The functional Aa-RNase III dimer is formed via mainly hydrophobic interactions, including a "ball-and-socket" junction that ensures accurate alignment of the two monomers. The fold of the polypeptide chain and its dimerization create a valley with two compound active centers at each end of the valley. The valley can accommodate a dsRNA substrate. Mn(2+) binding has significant impact on crystal packing, intermolecular interactions, thermal stability, and the formation of two RNA-cutting sites within each compound active center.

MeSH terms

  • Amino Acid Sequence
  • Binding Sites
  • Crystallography, X-Ray
  • Dimerization
  • Endoribonucleases / chemistry*
  • Escherichia coli Proteins*
  • Ligands
  • Manganese / metabolism
  • Models, Molecular
  • Molecular Sequence Data
  • Mutation
  • Nucleic Acid Conformation
  • Protein Binding
  • Protein Conformation
  • Protein Folding
  • Protein Structure, Secondary
  • Protein Structure, Tertiary
  • RNA, Double-Stranded / metabolism*
  • Ribonuclease III
  • Sequence Homology, Amino Acid


  • Escherichia coli Proteins
  • Ligands
  • RNA, Double-Stranded
  • Manganese
  • Endoribonucleases
  • Ribonuclease III
  • ribonuclease III, E coli

Associated data

  • PDB/1I4S
  • PDB/1JFZ