The structural basis for RNA-mediated gene regulation and for the de novo design of riboswitches: The example of the synthetic neomycin-sensing riboswitch

• Applicants: Professorin Dr. Beatrix Süß; Professor Dr. Jens Wöhnert
• Subject Area: Structural Biology
• Term: from 2009 to 2015
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 127262111

• Final Report Year: 2015

Final Report Abstract

Overall, the results obtained during this project establish the neomycin riboswitch as one of the best studied model systems for RNA-based regulatory systems. We could demonstrate that this riboswitch presents a structural novel architecture to bind aminoglycosides with very high specificity using a conformational capture mechanism of ligand recognition. Ligand specificity is achieved solely through the modulation of the global conformational dynamics of the complex by very small differences in localized intermolecular interactions. However, the structural and dynamical insights into ligand recognition and function of this riboswitch could not be successfully utilized to design riboswitches from inactive aptamers. Together with the observation that unguided saturation mutagenesis of key functional elements of this riboswitch only led to a moderate improvement of regulatory activity suggests that this small regulatory RNA-element is already very close to functional optimality.

Publications

• Highly modular structure and ligand binding by conformational capture in a minimalistic riboswitch. Angew. Chem. Int. Ed. Engl. 2010, 49: 6216-6219
  E. Duchardt-Ferner, J. E. Weigand, S. R. Schmidtke, O. Ohlenschläger, B. Suess, J. Wöhnert
  
• NMR resonance assignments of an engineered neomycin-sensing riboswitch RNA bound to ribostamycin and tobramycin. Biomol. NMR Assign. 2010, 4: 115-118
  S. R. Schmidtke, E. Duchardt-Ferner, J. E. Weigand, B. Suess, J. Wöhnert
  
• PELDOR spectroscopy reveals preorganization of the neomycin-responsive riboswitch tertiary structure. J Am Chem Soc 2010 132:1454-5
  I. Krstic, O. Frolow, D. Sezer, B. Endeward, J.E. Weigand, B. Suess, J.W. Engels, F.T. Prisner
  
• Mechanistic insights into an engineered riboswitch: a switching element which confers riboswitch activity. Nucleic Acids Res. 2011, 39: 3363-3372
  J. E. Weigand, S. R. Schmidtke, T. J. Will, E. Duchardt-Ferner, C. Hammann, J. Wöhnert, B. Suess
  
• Rapid identification of noncanonical RNA structure elements by direct detection of OH⋅⋅⋅O-P, NH⋅⋅⋅O-P, and NH2⋅⋅⋅O-P hydrogen bonds in solution NMR-spectroscopy. Angew. Chem. Int. Ed. Engl. 2011, 50: 7927-7930
  E. Duchardt-Ferner, J. Ferner, J. Wöhnert
  
• Building a stable RNA U-turn with a protonated cytidine. RNA 2014, 20: 1163-1172
  S. R. Gottstein-Schmidtke, E. Duchardt-Ferner, J. E. Weigand, F. Groher, B. Suess, J. Wöhnert
  
• Sequence elements distal to the ligand binding pocket tune the efficiency of a synthetic riboswitch. Chembiochem 2014, 11: 1627-1637
  J. E. Weigand, S. R. Gottstein-Schmidtke, S. Demolli, F. Groher, E. Duchardt-Ferner, J. Wöhnert, B. Suess