Complete Chemical Synthesis of Minimal Messenger RNA by Efficient Chemical Capping Reaction

ACS Chem Biol. 2022 Jun 17;17(6):1308-1314. doi: 10.1021/acschembio.1c00996. Epub 2022 May 24.

Abstract

Site-specific chemical modification of mRNA can improve its translational efficiency and stability. For this purpose, it is desirable to develop a complete chemical synthesis method for chemically modified mRNA. The key is a chemical reaction that introduces a cap structure into the chemically synthesized RNA. In this study, we developed a fast and quantitative chemical capping reaction between 5'-phosphorylated RNA and N7-methylated GDP imidazolide in the presence of 1-methylimidazole in the organic solvent dimethyl sulfoxide. It enabled quantitative preparation of capping RNA within 3 h. We prepared chemically modified 107-nucleotide mRNAs, including N6-methyladenosine, insertion of non-nucleotide linkers, and 2'-O-methylated nucleotides at the 5' end and evaluated their effects on translational activity in cultured HeLa cells. The results showed that mRNAs with non-nucleotide linkers in the untranslated regions were sufficiently tolerant to translation and that mRNAs with the Cap_2 structure had higher translational activity than those with the Cap_0 structure.

MeSH terms

  • HeLa Cells
  • Humans
  • Nucleotides*
  • Protein Biosynthesis
  • RNA Caps* / genetics
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism

Substances

  • Nucleotides
  • RNA Caps
  • RNA, Messenger