美国斯克里普斯研究所Ahmed H. Badran团队实现无需宿主基因组改造的高效遗传密码扩展。这一研究成果于2024年9月11日在线发表在国际学术期刊《自然—生物技术》上。

研究人员发现密码子使用是影响利用非天然密码子进行高效遗传密码扩展的一个此前未被认可的因素。依靠仅含天然核糖体的常规大肠杆菌菌株，研究人员开发了一种基于质粒的密码子压缩策略，能够减少上下文依赖性并提高非典型氨基酸（ncAA）在四联体密码子中的引入效率。

研究人员确认该策略与所有已知的遗传密码扩展资源兼容，进而鉴定出12对相互正交的tRNA-合成酶对。在此基础上，研究人员演化并优化了5对tRNA-合成酶，能够在正交四联体密码子上引入广泛的ncAA库。

最后，研究人员将这些资源扩展到体内生物合成平台，能够轻松生成超过100种包含多达三个独特ncAA的新型肽大环化合物。该方法将加速多重遗传密码扩展的创新，并推动化学多样性生物分子的发现。

据介绍，通过补充ncAA的翻译，可以产生具有自然界中不存在的新功能的蛋白质序列，但现有的ncAA引入策略效率低下且依赖上下文。

附：英文原文

Title: Efficient genetic code expansion without host genome modifications

Author: Costello, Alan, Peterson, Alexander A., Lanster, David L., Li, Zhiyi, Carver, Gavriela D., Badran, Ahmed H.

Issue&Volume: 2024-09-11

Abstract: Supplementing translation with noncanonical amino acids (ncAAs) can yield protein sequences with new-to-nature functions but existing ncAA incorporation strategies suffer from low efficiency and context dependence. We uncover codon usage as a previously unrecognized contributor to efficient genetic code expansion using non-native codons. Relying only on conventional Escherichia coli strains with native ribosomes, we develop a plasmid-based codon compression strategy that minimizes context dependence and improves ncAA incorporation at quadruplet codons. We confirm that this strategy is compatible with all known genetic code expansion resources, which allowed us to identify 12 mutually orthogonal transfer RNA (tRNA)–synthetase pairs. Enabled by these findings, we evolved and optimized five tRNA–synthetase pairs to incorporate a broad repertoire of ncAAs at orthogonal quadruplet codons. Lastly, we extend these resources to an in vivo biosynthesis platform that can readily create >100 new-to-nature peptide macrocycles bearing up to three unique ncAAs. Our approach will accelerate innovations in multiplexed genetic code expansion and the discovery of chemically diverse biomolecules.

DOI: 10.1038/s41587-024-02385-y

Source: https://www.nature.com/articles/s41587-024-02385-y