3FS0

Class II ligase ribozyme product-template duplex, structure 2

Summary for 3FS0

• Entry DOI: 10.2210/pdb3fs0/pdb
• Related: 3FTM
• Descriptor: 5'-R(*GP*GP*UP*GP*AP*GP*GP*CP*UP*G)-3', 5'-R(*CP*CP*AP*GP*UP*CP*GP*GP*AP*AP*C)-3', MAGNESIUM ION, ... (4 entities in total)
• Functional Keywords: ribozyme, ligase, 2'-5', 2-5, 2p5, rna
• Total number of polymer chains: 2
• Total formula weight: 6899.69

Authors

Pitt, J.N.,Ferre-D'Amare, A.R. (deposition date: 2009-01-08, release date: 2009-02-24, Last modification date: 2023-09-06)

Primary citation

Pitt, J.N.,Ferre-D'Amare, A.R.
Structure-guided engineering of the regioselectivity of RNA ligase ribozymes.
J.Am.Chem.Soc., 131:3532-3540, 2009

PubMed Abstract: 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.

PubMed: 19220054
DOI: 10.1021/ja8067325

Experimental method

X-RAY DIFFRACTION (2.3 Å)