Characterization of the kinetics of RNA annealing and strand displacement activities of the E. coli DEAD-box helicase CsdA

RNA Biol. 2013 Jan;10(1):149-56. doi: 10.4161/rna.23475. Epub 2013 Jan 1.


CsdA is one of five E. coli DEAD-box helicases and as a cold-shock protein assists RNA structural remodeling at low temperatures. The helicase has been shown to catalyze duplex unwinding in an ATP-dependent way and accelerate annealing of complementary RNAs, but detailed kinetic analyses are missing. Therefore, we performed kinetic measurements using a coupled annealing and strand displacement assay with high temporal resolution to analyze how CsdA balances the two converse activities. We furthermore tested the hypothesis that the unwinding activity of DEAD-box helicases is largely determined by the substrate's thermodynamic stability using full-length CsdA and a set of RNAs with constant length, but increasing GC content. The rate constants for strand displacement did indeed decrease with increasing duplex stability, with a calculated free energy between -31.3 and -40 kcal/mol being the limit for helix unwinding. Thus, our data generally support the above hypothesis, showing that for CsdA substrate thermal stability is an important rate limiting factor.

Keywords: CsdA; DEAD-box helicase; DeaD; FRET; RNA annealing; RNA chaperone; StpA; kinetics; strand displacement.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Base Composition
  • Base Pairing
  • Base Sequence
  • DEAD-box RNA Helicases / metabolism*
  • DNA Helicases / metabolism*
  • Escherichia coli / genetics*
  • Escherichia coli / metabolism*
  • Escherichia coli Proteins / metabolism*
  • Kinetics
  • RNA / chemistry
  • RNA / metabolism*
  • RNA Folding
  • RNA Stability
  • RNA, Double-Stranded / chemistry
  • RNA, Double-Stranded / metabolism
  • Substrate Specificity
  • Thermodynamics


  • Escherichia coli Proteins
  • RNA, Double-Stranded
  • RNA
  • deaD protein, E coli
  • DNA Helicases
  • DEAD-box RNA Helicases