PDBsum entry 3fs0

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dna_rna metals links
_MG ×5
Waters ×12
PDB id:
Name: RNA
Title: Class ii ligase ribozyme product-template duplex, structure 2
Structure: 5'-r( Gp Gp Up Gp Ap Gp Gp Cp Up G)-3'. Chain: a. Engineered: yes. Other_details: template strand of class ii ligase a4-11. 5'-r( Cp Cp Ap Gp Up Cp Gp Gp Ap Ap C)-3'. Chain: b. Engineered: yes. Other_details: product strand of class ii ligase a4-11, single stranded RNA with one 2'-5' linkage
Source: Synthetic: yes. Synthetic: yes
2.30Å     R-factor:   0.208     R-free:   0.234
Authors: J.N.Pitt,A.R.Ferre-D'Amare
Key ref: J.N.Pitt and A.R.Ferré-D'Amaré (2009). Structure-guided engineering of the regioselectivity of RNA ligase ribozymes. J Am Chem Soc, 131, 3532-3540. PubMed id: 19220054
08-Jan-09     Release date:   24-Feb-09    


J Am Chem Soc 131:3532-3540 (2009)
PubMed id: 19220054  
Structure-guided engineering of the regioselectivity of RNA ligase ribozymes.
J.N.Pitt, A.R.Ferré-D'Amaré.
Ribozyme-catalyzed RNA synthesis is central to the RNA world hypothesis. No natural RNA polymerase ribozymes have been discovered. However, ribozymes that catalyze the requisite chemistry, generating a new phosphodiester through attack of a terminal hydroxyl of an RNA on the alpha-phosphate of a triphosphate-activated oligonucleotide, have been isolated by in vitro selection. These experiments often yield ribozymes that generate 2'-5' phosphodiesters rather than conventional 3'-5' linkages. We have determined crystal structures of the duplex formed by the template segment of a representative 2'-5' RNA ligase ribozyme, the class II ligase, and its ligation product. The structures reveal a product-template duplex with a G x A pair at the ligation junction. This sheared pair is flanked on one side by a minor groove-broadening wedge comprised of two unpaired nucleotides. The reported structure of an independently isolated 3'-5' ligase ribozyme, the L1 ligase, shows a product-template duplex that shares the G x A pair with the class II ligase. However, this G x A pair is flanked by G x U wobbles, rather than an unpaired wedge. We demonstrate that these structural differences of the substrate-template duplexes are largely responsible for the divergent regioselectivity of the two ribozymes, independent of their catalytic moieties, by constructing chimeras. The L1 ligase with a class II substrate-template duplex shows a 30-fold increase in 2'-5' bond synthesis, while the class II ligase with an L1 substrate-template duplex produces 3'-5' bonds exclusively. These results demonstrate how local geometry inherent to the substrate-template duplexes controls the regioselectivity of ribozyme-catalyzed RNA ligation reactions.

Literature references that cite this PDB file's key reference

  PubMed id Reference
21857665 D.M.Shechner, and D.P.Bartel (2011).
The structural basis of RNA-catalyzed RNA polymerization.
  Nat Struct Mol Biol, 18, 1036-1042.
PDB codes: 3r1h 3r1l
20554048 A.R.Ferré-D'Amaré (2010).
Use of the spliceosomal protein U1A to facilitate crystallization and structure determination of complex RNAs.
  Methods, 52, 159-167.  
20947767 J.N.Pitt, and A.R.Ferré-D'Amaré (2010).
Rapid construction of empirical RNA fitness landscapes.
  Science, 330, 376-379.  
20693400 J.N.Pitt, I.Rajapakse, and A.R.Ferré-D'Amaré (2010).
SEWAL: an open-source platform for next-generation sequence analysis and visualization.
  Nucleic Acids Res, 38, 7908-7915.  
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