文献类型：期刊文献

英文题名：Exploring genetic codon expansion for unnatural amino acid incorporation in filamentous fungus Aspergillus nidulans

作者：Li, Xueying[1];Wang, Jing[1];Li, Jingyi[1];Zhou, Yao[1];Huang, Xiaofei[1];Guo, Lingyan[1];Liu, Renning[1];Luo, Yiqing[1];Tan, Xinyu[1];Hu, Xiaotao[1];Gao, Yan[1];Yu, Bingzi[1];Fu, Mingxin[1];Wang, Ping[2];Zhou, Shengmin[1]

机构：[1]East China Univ Sci & Technol, Sch Biotechnol, State Key Lab Bioreactor Engn, Shanghai 200237, Peoples R China;[2]Univ Minnesota, Dept Bioprod & Biosyst Engn, St Paul, MN 55108 USA

年份：2024

卷号：393

起止页码：91

外文期刊名：JOURNAL OF BIOTECHNOLOGY

收录：;EI(收录号：20243116782444);WOS:【SCI-EXPANDED(收录号:WOS:001282048200001)】；

基金：This work was supported by the National Natural Science Foundation of China (22077032 and 21672065) .

语种：英文

外文关键词：Unnatural amino acid; Amber suppression; Filamentous fungus; Aspergillus; Genetic codon expansion

摘要：Genetic code expansion technology allows the incorporation of unnatural amino acids (UAAs) into proteins, which is useful in protein engineering, synthetic biology, and gene therapy. Despite its potential applications in various species, filamentous fungi remain unexplored. This study aims to address this gap by developing these techniques in Aspergillus nidulans. We introduced an amber stop codon into a specific sequence within the reporter gene expressed in A. nidulans and replaced the anticodon of the fungal tRNATyr with CUA. This resulted in the synthesis of the target protein, confirming the occurrence of amber suppression in the fungus. When exogenous E. coli tRNATyrCUA (Ec. tRNATyr successfully synthesized the target protein via amber suppression and were shown to be orthogonal to the fungal translation system. By replacing the wild-type Ec.TyrRS with a mutant with a higher affinity for the UAA Omethyl-L-tyrosine, the fungal system was able to initiate the synthesis of the UAA-labeled protein (UAA-protein). We further increased the expression level of the UAA-protein through several rational modifications. The successful development of a genetic code expansion technique for A. nidulans has introduced a potentially valuable approach to the study of fungal protein structure and function.