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Detection of scalar couplings involving 2'-hydroxyl protons across hydrogen bonds in a frameshifting mRNA pseudoknot

David P Giedroc1, Peter V Cornish, Mirko Hennig

  • 1Department of Biochemistry, Center for Advanced Biomolecular Research, Texas A&M University, College Station, Texas 77843-2128, USA. giedroc@tamu.edu

Journal of the American Chemical Society
|April 17, 2003
PubMed

Insights

This study reveals critical RNA triple helix interactions in the pea enation mosaic virus-1 (PEMV-1) frameshift-stimulating pseudoknot. It details noncanonical hydrogen bonds essential for RNA structure and function.

Area of Science:

  • Molecular Biology
  • Structural Biology
  • Virology

Background:

  • The pea enation mosaic virus-1 (PEMV-1) frameshift-stimulating pseudoknot is crucial for viral replication.
  • This RNA structure is predominantly composed of an RNA triple helix.

Purpose of the Study:

  • To investigate the noncanonical O-H.N hydrogen bonding interactions within the PEMV-1 RNA triple helix.
  • To characterize the role of specific ribose 2'-OH protons and adenine N1 atoms in stabilizing the pseudoknot structure.

Main Methods:

  • Utilized a tailored nonrefocused (1H,15N) CPMG HSQC experiment on uniformly (13C,15N)-labeled PEMV-1 RNA.
  • Employed quantitative 1D (15N) spin-echo difference experiments to measure cross-hydrogen bond scalar coupling constants.

Main Results:

  • A direct correlation was observed between the 2'-OH proton of C15 and the N1 nitrogen of A27, mediated by a cross-hydrogen bond scalar coupling.
  • The (1H)J(2'OH,N) coupling constant for the C15/A27 interaction was determined to be 1.7 ± 0.1 Hz.
  • A larger coupling constant of 3.5 ± 0.3 Hz was measured for the C16/A25 interaction, despite the absence of a direct correlation signal.

Conclusions:

  • These findings provide a detailed understanding of critical noncanonical hydrogen bonding in RNA triple helices.
  • The identified loop-stem interactions are essential for the stability and function of the PEMV-1 pseudoknot.
  • The study highlights the importance of these interactions in viral RNA structures.

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