The 2′-hydroxyl group of the guanosine nucleophile donates a functionally important hydrogen bond in the Tetrahymena ribozyme reaction

James L. Hougland, Raghuvir N. Sengupta, Qing Dai, Shirshendu K. Deb, Joseph A. Piccirilli

Research output: Contribution to journalArticlepeer-review

16 Scopus citations

Abstract

In the first step of self-splicing, group I introns utilize an exogenous guanosine nucleophile to attack the 5′-splice site. Removal of the 2′-hydroxyl of this guanosine results in a 106-fold loss in activity, indicating that this functional group plays a critical role in catalysis. Biochemical and structural data have shown that this hydroxyl group provides a ligand for one of the catalytic metal ions at the active site. However, whether this hydroxyl group also engages in hydrogen-bonding interactions remains unclear, as attempts to elaborate its function further usually disrupt the interactions with the catalytic metal ion. To address the possibility that this 2′-hydroxyl contributes to catalysis by donating a hydrogen bond, we have used an atomic mutation cycle to probe the functional importance of the guanosine 2′-hydroxyl hydrogen atom. This analysis indicates that, beyond its role as a ligand for a catalytic metal ion, the guanosine 2′-hydroxyl group donates a hydrogen bond in both the ground state and the transition state, thereby contributing to cofactor recognition and catalysis by the intron. Our findings continue an emerging theme in group I intron catalysis: the oxygen atoms at the reaction center form multidentate interactions that function as a cooperative network. The ability to delineate such networks represents a key step in dissecting the complex relationship between RNA structure and catalysis.

Original languageEnglish (US)
Pages (from-to)7684-7694
Number of pages11
JournalBiochemistry
Volume47
Issue number29
DOIs
StatePublished - Jul 22 2008
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry

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