A nucleotide-independent cyclic nitroxide label for monitoring segmental motions in nucleic acids

Phuong H. Nguyen, Anna M. Popova, K. Hideg, Peter Z. Qin

Research output: Contribution to journalArticle

11 Citations (Scopus)

Abstract

Background: Spin labels, which are chemically stable radicals attached at specific sites of a bio-molecule, enable investigations on structure and dynamics of proteins and nucleic acids using techniques such as site-directed spin labeling and paramagnetic NMR. Among spin labels developed, the class of rigid labels have limited or no independent motions between the radical bearing moiety and the target, and afford a number of advantages in measuring distances and monitoring local dynamics within the parent bio-molecule. However, a general method for attaching a rigid label to nucleic acids in a nucleotide-independent manner has not been reported. Results: We developed an approach for installing a nearly rigid nitroxide spin label, designated as R5c, at a specific site of the nucleic acid backbone in a nucleotide-independent manner. The method uses a post-synthesis approach to covalently attach the nitroxide moiety in a cyclic fashion to phosphorothioate groups introduced at two consecutive nucleotides of the target strand. R5c-labeled nucleic acids are capable of pairing with their respective complementary strands, and the cyclic nature of R5c attachment significantly reduced independence motions of the label with respect to the parent duplex, although it may cause distortion of the local environment at the site of labeling. R5c yields enhanced sensitivity to the collective motions of the duplex, as demonstrated by its capability to reveal changes in collective motions of the substrate recognition duplex of the 120-kDa Tetrahymena group I ribozyme, which elude detection by a flexible label. Conclusions: The cyclic R5c nitroxide can be efficiently attached to a target nucleic acid site using a post-synthetic coupling approach conducted under mild biochemical conditions, and serves as a viable label for experimental investigation of segmental motions in nucleic acids, including large folded RNAs.

Original languageEnglish
Article number6
JournalBMC Biophysics
Volume8
Issue number1
DOIs
Publication statusPublished - 2015

Fingerprint

Cyclic Nucleotides
Nucleic Acids
Spin Labels
Nucleotides
Tetrahymena
RNA
Proteins

Keywords

  • Dynamics
  • EPR
  • Nucleic acids
  • Rigid label
  • Site-directed spin labeling

ASJC Scopus subject areas

  • Biophysics

Cite this

A nucleotide-independent cyclic nitroxide label for monitoring segmental motions in nucleic acids. / Nguyen, Phuong H.; Popova, Anna M.; Hideg, K.; Qin, Peter Z.

In: BMC Biophysics, Vol. 8, No. 1, 6, 2015.

Research output: Contribution to journalArticle

Nguyen, Phuong H. ; Popova, Anna M. ; Hideg, K. ; Qin, Peter Z. / A nucleotide-independent cyclic nitroxide label for monitoring segmental motions in nucleic acids. In: BMC Biophysics. 2015 ; Vol. 8, No. 1.
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