We synthesized a small library of eighteen 5-substituted pyrimidine or 7-substituted 7-deazapurine nucleoside triphosphates bearing methyl, ethynyl, phenyl, benzofuryl or dibenzofuryl groups through cross-coupling reactions of nucleosides followed by triphosphorylation or through direct cross-coupling reactions of halogenated nucleoside triphosphates. We systematically studied the influence of the modification on the efficiency of T7 RNA polymerase catalyzed synthesis of modified RNA and found that modified ATP, UTP and CTP analogues bearing smaller modifications were good substrates and building blocks for the RNA synthesis even in difficult sequences incorporating multiple modified nucleotides. Bulky dibenzofuryl derivatives of ATP and GTP were not substrates for the RNA polymerase. In the case of modified GTP analogues, a modified procedure using a special promoter and GMP as initiator needed to be used to obtain efficient RNA synthesis. The T7 RNA polymerase synthesis of modified RNA can be very efficiently used for synthesis of modified RNA but the method has constraints in the sequence of the first three nucleotides of the transcript, which must contain a non-modified G in the +1 position.

Institute of Organic Chemistry and Biochemistry Czech Academy of Sciences Flemingovo nam 2 CZ 16610 Prague 6 Czech Republic

Citace poskytuje Crossref.org

Enzymatic Synthesis of Modified RNA Containing 5-Methyl- or 5-Ethylpyrimidines or Substituted 7-Deazapurines and Influence of the Modifications on Stability, Translation, and CRISPR-Cas9 Cleavage

Enzymatic synthesis of reactive RNA probes containing squaramate-linked cytidine or adenosine for bioconjugations and cross-linking with lysine-containing peptides and proteins

Synthesis and Biological Profiling of Quinolino-Fused 7-Deazapurine Nucleosides

Expedient production of site specifically nucleobase-labelled or hypermodified RNA with engineered thermophilic DNA polymerases

Chloroacetamide-Modified Nucleotide and RNA for Bioconjugations and Cross-Linking with RNA-Binding Proteins

Design, Synthesis, and Biochemical and Biological Evaluation of Novel 7-Deazapurine Cyclic Dinucleotide Analogues as STING Receptor Agonists