Abstract
With the wide application of messenger RNA (mRNA) technology in medicine and vaccine fields, higher requirements are put forward for mRNA expression efficiency in vivo. Since the 5' cap structure can spatially protect mRNA from exonuclease degradation and enhance the initiation of translation reactions, in vitro mRNA caps are a promising option to improve the efficiency of mRNA expression in vivo. 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 Escherichia coli 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.