An active site rearrangement within the Tetrahymena group I ribozyme releases nonproductive interactions and allows formation of catalytic interactions

Raghuvir N. Sengupta, Sabine N.S. Van Schie, George Giambaşu, Qing Dai, Joseph D. Yesselman, Darrin York, Joseph A. Piccirilli, Daniel Herschlag

Research output: Contribution to journalArticle

4 Scopus citations

Abstract

Biological catalysis hinges on the precise structural integrity of an active site that binds and transforms its substrates and meeting this requirement presents a unique challenge for RNA enzymes. Functional RNAs, including ribozymes, fold into their active conformations within rugged energy landscapes that often contain misfolded conformers. Here we uncover and characterize one such "off-pathway" species within an active site after overall folding of the ribozyme is complete. The Tetrahymena group I ribozyme (E) catalyzes cleavage of an oligonucleotide substrate (S) by an exogenous guanosine (G) cofactor. We tested whether specific catalytic interactions with G are present in the preceding E•S•G and E•G ground-state complexes. We monitored interactions with G via the effects of 2'- and 3'-deoxy (-H) and -amino (-NH2) substitutions on G binding. These and prior results reveal that G is bound in an inactive configuration within E•G, with the nucleophilic 3'-OH making a nonproductive interaction with an active site metal ion termed MA and with the adjacent 2'-OH making no interaction. Upon S binding, a rearrangement occurs that allows both -OH groups to contact a different active site metal ion, termed MC, to make what are likely to be their catalytic interactions. The reactive phosphoryl group on S promotes this change, presumably by repositioning the metal ions with respect to G. This conformational transition demonstrates local rearrangements within an otherwise folded RNA, underscoring RNA's difficulty in specifying a unique conformation and highlighting Nature's potential to use local transitions of RNA in complex function.

Original languageEnglish (US)
Pages (from-to)32-48
Number of pages17
JournalRNA
Volume22
Issue number1
DOIs
StatePublished - Jan 2016

All Science Journal Classification (ASJC) codes

  • Molecular Biology

Keywords

  • Active site
  • Conformational change
  • Metal ion
  • Noncoding RNA
  • RNA catalysis

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    Sengupta, R. N., Van Schie, S. N. S., Giambaşu, G., Dai, Q., Yesselman, J. D., York, D., Piccirilli, J. A., & Herschlag, D. (2016). An active site rearrangement within the Tetrahymena group I ribozyme releases nonproductive interactions and allows formation of catalytic interactions. RNA, 22(1), 32-48. https://doi.org/10.1261/rna.053710.115