Phosphorothioate Modification of mRNA Accelerates the Rate of Translation Initiation to Provide More Efficient Protein Synthesis

• Published in: Angewandte Chemie International Edition Vol. 59; no. 40; pp. 17403 - 17407
• Main Authors: Kawaguchi, Daisuke, Kodama, Ayumi, Abe, Naoko, Takebuchi, Kei, Hashiya, Fumitaka, Tomoike, Fumiaki, Nakamoto, Kosuke, Kimura, Yasuaki, Shimizu, Yoshihiro, Abe, Hiroshi
• Format: Journal Article
• Language: English
• Published: Germany Wiley Subscription Services, Inc 28.09.2020
• Edition: International ed. in English
• Subjects: backbone modification; Complex formation; E coli; Epidermal Growth Factor - genetics; Epidermal Growth Factor - metabolism; Escherichia coli - metabolism; mRNA; mRNA turnover; Peptide Chain Initiation, Translational; Phosphorothioate; phosphorothioate oligonucleotides; Phosphorothioate Oligonucleotides - chemistry; Protein Biosynthesis; protein expression; Protein folding; Protein synthesis; Proteins; Ribosomes; Ribosomes - metabolism; RNA; RNA modification; RNA, Messenger - chemistry; RNA, Messenger - metabolism; Transcription; Translation; Translation initiation; mRNA; phosphorothioate oligonucleotides; RNA; protein expression; backbone modification
• ISSN: 1433-7851; 1521-3773
• DOI: 10.1002/anie.202007111

Messenger RNAs (mRNAs) with phosphorothioate modification (PS‐mRNA) to the phosphate site of A, G, C, and U with all 16 possible combinations were prepared, and the translation reaction was evaluated using an E. coli cell‐free translation system. Protein synthesis from PS‐mRNA increased in 12 of 15 patterns when compared with that of unmodified mRNA. The protein yield increased 22‐fold when the phosphorothioate modification at A/C sites was introduced into the region from the 5′‐end to the initiation codon. Single‐turnover analysis of PS‐mRNA translation showed that phosphorothioate modification increases the number of translating ribosomes, thus suggesting that the rate of translation initiation (rate of ribosome complex formation) is positively affected by the modification. The method provides a new strategy for improving translation by using non‐natural mRNA. Phosphorothioate (PS) modification of mRNA leads to enhanced protein expression in a reconstituted E. coli translation system. Single‐turnover analysis of PS‐mRNA translation showed that phosphorothioate modification increases the number of translating ribosomes, thus suggesting that translation initiation is more efficient on the modified mRNA.