基于TPT3/NaM型非天然碱基核苷酸的无细胞基因表达研究

非天然碱基是一类能够模拟天然碱基功能，在源头上对遗传信息进行编码、存储和提取的分子。但是，目前在细胞体系中进行的非天然碱基基因表达不可避免地受到细胞膜的屏障和细胞生长的限制，复杂的细胞表达系统严重阻碍了非天然碱基研究及其生物医学应用。为此，本课题拟以无细胞体系为研究对象，通过化学生物学交叉研究方法，探索构筑新型简单、高效的非天然碱基无细胞基因表达途径。我们将从前期发现的具有最佳体外复制活性的TPT3/NaM型非天然碱基结构改造入手，在分子层面研究和阐明非天然碱基核苷酸进行高效无细胞蛋白表达的化学结构特征；在密码子层面研究和揭示非天然碱基与天然碱基组合编码非天然氨基酸的构效关系和互作机制；在整体层面上，建立非天然碱基无细胞基因表达系统的基本构筑原则和优化策略。本研究将从源头上为非天然碱基谱写遗传密码提供新的研究平台，为蛋白质改造及生物大分子药物创制提供前瞻性技术储备。

Unnatural base pairs are a class of promising nucleotide derivatives, which can be used to store, code and retrieve gene informaton by mimicing the fucntions of natural base pairs (A/T(U), G/C). However, the cellular gene expression systems for unnatural base pairs suffer from the obstruction of cellular membranes and the constraint of sustaining cell growth. The complexity of cellular gene expression systems have severely limited the synthetic biological study of unnatural base pairs and their further applications in the biomedical fields. In the project, based on the our previously researches in finding TPT3/NaM unnatural base pair bearing the highest replicate efficiecy and fidelity, we will focus on the cell-free gene expression systems, and attempt to build a new efficient and concise unnatural-base-pair cell-free gene expression system by use of chemical biological methods. In the level of molecules, we will study which kind of TPT3/NaM-type nucleotides can promote the cell-free gene expression with optimal biological activities, and then illustrate their chemical features. In the level of gene code, we will study how to use the unnatural bases to design extended gene code via combinations with natural bases, and then investigate possible interactions of the unnatural gene codes under cell-free gene expression systems. In the integrity level, we will build the principle of making efficient and concise TPT3/NaM-unnatural-base-pair cell-free gene expression system, and then indicate how to improve the properties of such gene expression systems. These researches will offer a new platform from the orgin to explore how the unnatural base pairs extend the gene codes, and lay a foundation for developments of forward-looking technologies in fields of protein drugs and other branches of biomedicine.