Monitoring RNA base structure and dynamics using site-directed spin labeling

Peter Z. Qin, K. Hideg, Juli Feigon, Wayne L. Hubbell

Research output: Contribution to journalArticle

90 Citations (Scopus)

Abstract

Site-directed spin labeling utilizes site-specific attachment of a stable nitroxide radical to probe the structure and dynamics of macromolecules. In the present study, a 4-thiouridine base is introduced at each of six different positions in a 23-nucleotide RNA molecule. The 4-thiouridine derivatives were subsequently modified with one of three methanethiosulfonate nitroxide reagents to introduce a spin label at specific sites. The electron paramagnetic resonance spectra of the labeled RNAs were analyzed in terms of nitroxide motion and the RNA solution structure. At a base-paired site in the RNA helix, where the nitroxide has weak or no local interactions, motion of the nitroxide is apparently dominated by rotation about bonds within the probe. The motion is similar to that found for a structurally related probe on helical sites in proteins, suggesting a similar mode of motion. At other sites that are hydrogen bonded and stacked within the helix, local interactions within the RNA molecule modulate the nitroxide motion in a manner consistent with expectations based on the known structure. For a base that is not structurally constrained, the mobility is higher than at any other site, presumably due to motion of the base itself. These results demonstrate the general utility of the 4-thiouridine/methanethiosulfonate coupling method to introduce nitroxide spin labels into RNA and the ability of the resulting label to probe local structure and dynamics.

Original languageEnglish
Pages (from-to)6772-6783
Number of pages12
JournalBiochemistry
Volume42
Issue number22
DOIs
Publication statusPublished - Jun 10 2003

Fingerprint

Labeling
Thiouridine
RNA
Monitoring
Spin Labels
Molecules
Electron Spin Resonance Spectroscopy
Macromolecules
Paramagnetic resonance
Labels
Hydrogen
Nucleotides
Derivatives
Proteins
methanethiosulfonate

ASJC Scopus subject areas

  • Biochemistry

Cite this

Monitoring RNA base structure and dynamics using site-directed spin labeling. / Qin, Peter Z.; Hideg, K.; Feigon, Juli; Hubbell, Wayne L.

In: Biochemistry, Vol. 42, No. 22, 10.06.2003, p. 6772-6783.

Research output: Contribution to journalArticle

Qin, Peter Z. ; Hideg, K. ; Feigon, Juli ; Hubbell, Wayne L. / Monitoring RNA base structure and dynamics using site-directed spin labeling. In: Biochemistry. 2003 ; Vol. 42, No. 22. pp. 6772-6783.
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