Structural probing techniques on natural aptamers

Catherine A. Wakeman, Wade C. Winkler

Research output: Chapter in Book/Report/Conference proceedingChapter

6 Scopus citations

Abstract

RNA sequences fold in a hierarchical manner to form complex structures. This folding pathway proceeds first with formation of secondary structure elements followed by the compilation of tertiary contacts. Although bioinformatics-based tools are commonly used to predict secondary structure models, it is notoriously difficult to achieve a high degree of accuracy via these approaches alone. Therefore, a diverse assortment of biochemical and biophysical techniques are regularly used to investigate the structural arrangements of biological RNAs. Among these different experimental techniques are structural probing methods, which are often times employed to determine which nucleotides for a given RNA polymer are paired or unpaired. Yet other probing methods assess whether certain RNA structures undergo dynamical structure changes. In this chapter we outline a general protocol for in-line probing, a method for analyzing secondary structure (and backbone flexibility) and describe a basic experimental protocol for hydroxyl radical footprinting as a method of investigating RNA folding.

Original languageEnglish
Title of host publicationNucleic Acid and Peptide Aptamers
Subtitle of host publicationMethods and Protocols
EditorsGunter Mayer
Pages115-133
Number of pages19
DOIs
StatePublished - 2009

Publication series

NameMethods in Molecular Biology
Volume535
ISSN (Print)1064-3745

Keywords

  • In-line probing
  • RNA folding
  • RNA secondary and tertiary structure
  • hydroxyl radical footprinting
  • riboswitch

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    Wakeman, C. A., & Winkler, W. C. (2009). Structural probing techniques on natural aptamers. In G. Mayer (Ed.), Nucleic Acid and Peptide Aptamers: Methods and Protocols (pp. 115-133). (Methods in Molecular Biology; Vol. 535). https://doi.org/10.1007/978-1-59745-557-2_8