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Updated: May 13, 2026

Nanomanipulation of Single RNA Molecules by Optical Tweezers
Published on: August 20, 2014
Capturing hammerhead ribozyme structures in action by modulating general base catalysis
Young-In Chi1, Monika Martick, Monica Lares
1Department of Molecular and Cellular Biochemistry, Center for Structural Biology, University of Kentucky, Lexington, Kentucky, United States of America. ychi@uky.edu
Abstract:
We have obtained precatalytic (enzyme-substrate complex) and postcatalytic (enzyme-product complex) crystal structures of an active full-length hammerhead RNA that cleaves in the crystal. Using the natural satellite tobacco ringspot virus hammerhead RNA sequence, the self-cleavage reaction was modulated by substituting the general base of the ribozyme, G12, with A12, a purine variant with a much lower pKa that does not significantly perturb the ribozyme's atomic structure. The active, but slowly cleaving, ribozyme thus permitted isolation of enzyme-substrate and enzyme-product complexes without modifying the nucleophile or leaving group of the cleavage reaction, nor any other aspect of the substrate. The predissociation enzyme-product complex structure reveals RNA and metal ion interactions potentially relevant to transition-state stabilization that are absent in precatalytic structures.
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