2luq
Solution structure of double-stranded RNA binding domain of S.cerevisiae RNase III (rnt1p)
Structural highlights
FunctionRNT1_YEAST DsRNA-specific nuclease that cleaves eukaryotic pre-ribosomal RNA at the U3 snoRNP-dependent A0 site in the 5'-external transcribed spacer (ETS) and in the 3'-ETS. In vitro, cleaves synthetic 5'-ETS RNA A0 site in the absence of snoRNA or other factors. Has an essential growth function in addition to pre-rRNA processing. Publication Abstract from PubMedThe Saccharomyces cerevisiae RNase III enzyme Rnt1p preferentially binds to double-stranded RNA hairpin substrates with a conserved (A/u)GNN tetraloop fold, via shape-specific interactions by its double-stranded RNA-binding domain (dsRBD) helix alpha1 to the tetraloop minor groove. To investigate whether conformational flexibility in the dsRBD regulates the binding specificity, we determined the backbone dynamics of the Rnt1p dsRBD in the free and AGAA hairpin-bound states using NMR spin-relaxation experiments. The intrinsic microsecond-to-millisecond timescale dynamics of the dsRBD suggests that helix alpha1 undergoes conformational sampling in the free state, with large dynamics at some residues in the alpha1-beta1 loop (alpha1-beta1 hinge). To correlate free dsRBD dynamics with structural changes upon binding, we determined the solution structure of the free dsRBD used in the previously determined RNA-bound structures. The Rnt1p dsRBD has an extended hydrophobic core comprising helix alpha1, the alpha1-beta1 loop, and helix alpha3. Analysis of the backbone dynamics and structures of the free and bound dsRBD reveals that slow-timescale dynamics in the alpha1-beta1 hinge are associated with concerted structural changes in the extended hydrophobic core that govern binding of helix alpha1 to AGAA tetraloops. The dynamic behavior of the dsRBD bound to a longer AGAA hairpin reveals that dynamics within the hydrophobic core differentiate between specific and nonspecific sites. Mutations of residues in the alpha1-beta1 hinge result in changes to the dsRBD stability and RNA-binding affinity and cause defects in small nucleolar RNA processing invivo. These results reveal that dynamics in the extended hydrophobic core are important for binding site selection by the Rnt1p dsRBD. Intrinsic Dynamics of an Extended Hydrophobic Core in the S. cerevisiae RNase III dsRBD Contributes to Recognition of Specific RNA Binding Sites.,Hartman E, Wang Z, Zhang Q, Roy K, Chanfreau G, Feigon J J Mol Biol. 2012 Nov 28. pii: S0022-2836(12)00897-2. doi:, 10.1016/j.jmb.2012.11.025. PMID:23201338[1] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See AlsoReferences
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