Productive folding to the native state by a group II intron ribozyme

Jennifer F. Swisher, Linhui J. Su, Michael Brenowitz, Vernon E. Anderson, Anna Marie Pyle

Research output: Contribution to journalArticle

75 Scopus citations

Abstract

Group II introns are large catalytic RNA molecules that fold into compact structures essential for the catalysis of splicing and intron mobility reactions. Despite a growing body of information on the folded state of group II introns at equilibrium, there is currently no information on the folding pathway and little information on the ionic requirements for folding. Folding isotherms were determined by hydroxyl radical footprinting for the 32 individual protections that are distributed throughout a group II intron ribozyme derived from intron ai5γ. The isotherms span a similar range of Mg2+ concentrations and share a similar index of cooperativity. Time-resolved hydroxyl radical footprinting studies show that all regions of the ribozyme fold slowly and with remarkable synchrony into a single catalytically active structure at a rate comparable to those of other ribozymes studied thus far. The rate constants for the formation of tertiary contacts and recovery of catalytic activity are identical within experimental error. Catalytic activity analyses in the presence of urea provide no evidence that the slow folding of the ai5γ intron is attributable to the presence of unproductive kinetic traps along the folding pathway. Taken together, the data suggest that the rate-limiting step for folding of group II intron ai5γ occurs early along the reaction pathway. We propose that this behavior resembles protein folding that is limited in rate by high contact order, or the need to form key tertiary interactions from partners that are located far apart in the primary or secondary structure.

Original languageEnglish (US)
Pages (from-to)297-310
Number of pages14
JournalJournal of Molecular Biology
Volume315
Issue number3
DOIs
StatePublished - Jan 1 2002

Keywords

  • Footprinting
  • Group II ribozyme
  • Kinetics
  • Magnesium titration
  • RNA folding

ASJC Scopus subject areas

  • Structural Biology
  • Molecular Biology

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