Wang Yiming, Wang Xiaoxue, Li Wenchao, Chen Xinjie, Lu Yuan
Department of Chemical Engineering, Tsinghua University, Beijing, 100084, China.
Key Laboratory of Industrial Biocatalysis, Ministry of Education, Tsinghua University, Beijing, 100084, China.
Biotechnol Notes. 2024 Nov 20;5:165-172. doi: 10.1016/j.biotno.2024.11.005. eCollection 2024.
With the wide application of messenger RNA (mRNA) technology in medicine and vaccine fields, higher requirements are put forward for mRNA expression efficiency . Since the 5' cap structure can spatially protect mRNA from exonuclease degradation and enhance the initiation of translation reactions, mRNA caps are a promising option to improve the efficiency of mRNA expression . In order to obtain more efficient mRNA capping enzymes, seven mRNA capping enzymes from different viral sources were explored in this study. Eukaryotic and prokaryotic cells were used for the heterologous expression of the cap enzymes, and was identified as the most suitable host cell for heterologous expression. In addition, in order to improve the solubility of the capping enzyme, four kinds of soluble labels were screened, among which maltose-binding protein had the best effect and the widest applicability. The mRNA was then transfected into the human cells, and the highest transfection efficiency was achieved using the bluetongue virus capping enzyme. Its effect was 38 % higher than that of the previously widely used vaccinia virus capping enzyme. This work will promote the development of mRNA technology and expand its application space.
随着信使核糖核酸（mRNA）技术在医学和疫苗领域的广泛应用，对mRNA表达效率提出了更高的要求。由于5'帽结构可以在空间上保护mRNA免受核酸外切酶降解，并增强翻译反应的起始，mRNA帽是提高mRNA表达效率的一个有前景的选择。为了获得更高效的mRNA加帽酶，本研究探索了七种来自不同病毒来源的mRNA加帽酶。真核细胞和原核细胞被用于帽酶的异源表达，并且被确定为最适合异源表达的宿主细胞。此外，为了提高加帽酶的溶解性，筛选了四种可溶性标签，其中麦芽糖结合蛋白效果最佳且适用性最广。然后将mRNA转染到人类细胞中，使用蓝舌病毒加帽酶实现了最高的转染效率。其效果比先前广泛使用的痘苗病毒加帽酶高38%。这项工作将推动mRNA技术的发展并扩大其应用空间。
