Synthesis of Selenium-modified RNA by engineered RNA Polymerases

Final Report Abstract

Modifications within RNA are used for studying RNA structure and dynamics. The 2’-position of the sugar moiety is a desirable position for introducing modifications within the RNA target because modifications attached to this position provide the RNA with desired properties such as increased stability. Additionally, the 2’-position is important because it can be advantageous for introducing selenium into RNA for use in X-ray crystallographic structure determination. 2’-Methylseleno (2’-SeCH3)-modified RNA was accessible by chemical synthesis before this project started. In this study, the enzymatic synthesis of 2’-SeCH3- modified RNA has been elaborated by using engineered variants of T7 RNA polymerase in combination with 2’-SeCH3-modified nucleoside triphosphates (NTPs). This approach provides access to long selenium modified RNA, which cannot be accomplished by chemical synthesis alone. These investigations were conducted in collaboration with Prof. Dr. Ronald Micura from Innsbruck University in Austria, where his research team synthesized the four 2’-SeCH3-NTPs.

Publications

• “Enzymatic synthesis of 2’-methylseleno-modified RNA” Chem. Sci. 2011, 2, 2224-2231
  V. Siegmund, T. Santner, R. Micura, A. Marx
  
• “Enzymatic synthesis of 8-vinyland 8-styryl-2’-deoxyguanosine modified DNA - novel fluorescent molecular probes” Bioorg. Med. Chem. Lett. 2012, 22, 3136-3139
  B. Holzberger, J. Strohmeier, V. Siegmund, U. Diederichsen, A. Marx
  
• “Screening mutant libraries of T7 RNA polymerase for candidates with increased acceptance of 2’-modified nucleotides” Chem. Commun. 2012, 9870-2
  V. Siegmund, T. Santner, R. Micura, A. Marx
  
• “Solution Structure and Conformational Dynamics of Deoxyxylonucleic Acid (dXNA): An Orthogonal Nucleic Acid Candidate” Chem. Eur. J. 2012, 18, 869-79
  M. Maiti, V. Siegmund, M. Abramov, E. Lescrinier, H. Rosemeyer, M. Froeyen, A. Ramaswamy, A. Ceulemans, A. Marx, P. Herdewijn
  
• “The synthesis of 2’-methylseleno adenosine and guanosine 5’-triphosphates” Bioorg. Med. Chem. Lett. 2012, 20, 2416-8
  T. Santner, V. Siegmund, A. Marx, R. Micura