title: Crystallographic and modeling studies of RNase III suggest a mechanism for double-stranded RNA cleavage.
abstract: 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.
authors.affiliation: Macromolecular Crystallography Laboratory, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Frederick, MD 21702, USA.
authors.forename: J
authors.initials: J
authors.lastname: Blaszczyk
authors.forename: J E
authors.initials: JE
authors.lastname: Tropea
authors.forename: M
authors.initials: M
authors.lastname: Bubunenko
authors.forename: K M
authors.initials: KM
authors.lastname: Routzahn
authors.forename: D S
authors.initials: DS
authors.lastname: Waugh
authors.forename: D L
authors.initials: DL
authors.lastname: Court
authors.forename: X
authors.initials: X
authors.lastname: Ji
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D018360:Crystallography, X-Ray; D019281:Dimerization; D004722:Endoribonucleases / Q000737:chemistry*; D029968:Escherichia coli Proteins; D008024:Ligands; D008345:Manganese / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D011487:Protein Conformation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D012330:RNA, Double-Stranded / Q000378:metabolism*; D043244:Ribonuclease III; D017386:Sequence Homology, Amino Acid
chemical_list: D029968:Escherichia coli Proteins; D008024:Ligands; D012330:RNA, Double-Stranded; D008345:Manganese; D004722:Endoribonucleases; D043244:Ribonuclease III; C468366:ribonuclease III, E coli