Folding mechanism of the Tetrahymena ribozyme P4-P6 domain

M. L. Deras, Michael D. Brenowitz, C. Y. Ralston, M. R. Chance, S. A. Woodson

Research output: Contribution to journalArticle

81 Citations (Scopus)

Abstract

Synchrotron X-ray-dependent hydroxyl radical footprinting was used to probe the folding kinetics of the P4-P6 domain of the Tetrahymena group I ribozyme, which forms a stable, closely packed tertiary structure. The 160-nt domain folds independently at a similar rate (~2 s-1>) as it does in the ribozyme, when folding is measured in 10 mM sodium cacodylate and 10 mM MgCl2. Surprisingly, tertiary interactions around a three-helix junction (P5abc) within the P4-P6 domain fold at least 25 times more rapidly (k ≥ 50 s-1>) in isolation, than when part of the wild-type P4-P6 RNA. This difference implies that long-range interactions in the P4-P6 domain can interfere with folding of P5abc. P4-P6 was observed to fold much faster at higher ionic strength than in 10 mM sodium cacodylate. Analytical centrifugation was used to measure the sedimentation and diffusion coefficients of the unfolded RNA. The hydrodynamic radius of the RNA decreased from 58 to 46 Å over the range of 0-100 mM NaCl. We propose that at low ionic strength, the addition of Mg2+ causes the domain to collapse to a compact intermediate where P5abc is trapped in a non-native structure. At high ionic strength, the RNA rapidly collapses to the native structure. Faster folding most likely results from a different average initial conformation of the RNA in higher salt conditions.

Original languageEnglish (US)
Pages (from-to)10975-10985
Number of pages11
JournalBiochemistry
Volume39
Issue number36
DOIs
StatePublished - Sep 12 2000

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Tetrahymena
Catalytic RNA
Cacodylic Acid
Osmolar Concentration
RNA
Ionic strength
Sodium
RNA Folding
Nucleic Acid Conformation
Synchrotrons
Magnesium Chloride
Hydrodynamics
Centrifugation
Hydroxyl Radical
Salts
X-Rays
Sedimentation
Conformations
X rays
Kinetics

ASJC Scopus subject areas

  • Biochemistry

Cite this

Deras, M. L., Brenowitz, M. D., Ralston, C. Y., Chance, M. R., & Woodson, S. A. (2000). Folding mechanism of the Tetrahymena ribozyme P4-P6 domain. Biochemistry, 39(36), 10975-10985. https://doi.org/10.1021/bi0010118

Folding mechanism of the Tetrahymena ribozyme P4-P6 domain. / Deras, M. L.; Brenowitz, Michael D.; Ralston, C. Y.; Chance, M. R.; Woodson, S. A.

In: Biochemistry, Vol. 39, No. 36, 12.09.2000, p. 10975-10985.

Research output: Contribution to journalArticle

Deras, ML, Brenowitz, MD, Ralston, CY, Chance, MR & Woodson, SA 2000, 'Folding mechanism of the Tetrahymena ribozyme P4-P6 domain', Biochemistry, vol. 39, no. 36, pp. 10975-10985. https://doi.org/10.1021/bi0010118
Deras ML, Brenowitz MD, Ralston CY, Chance MR, Woodson SA. Folding mechanism of the Tetrahymena ribozyme P4-P6 domain. Biochemistry. 2000 Sep 12;39(36):10975-10985. https://doi.org/10.1021/bi0010118
Deras, M. L. ; Brenowitz, Michael D. ; Ralston, C. Y. ; Chance, M. R. ; Woodson, S. A. / Folding mechanism of the Tetrahymena ribozyme P4-P6 domain. In: Biochemistry. 2000 ; Vol. 39, No. 36. pp. 10975-10985.
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