State Key Laboratory of Coordination Chemistry, School of Chemistry and Chemical Engineering, Nanjing University, 210023, Nanjing, China.
State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University, 210023, Nanjing, China.
Angew Chem Int Ed Engl. 2023 May 15;62(21):e202216115. doi: 10.1002/anie.202216115. Epub 2023 Apr 13.
Site-specific functionalization of natural amino acid-containing biological nanopores is pivotal in single molecule sensing. However, pore engineering methodologies are restricted to a limited choice and introduction of unnatural chemical components is extremely difficult. Herein we report the genetic code expansion (GCE) strategy to introduce unnatural amino acid (UAA) to an octameric Mycobacterium smegmatis porin A (MspA) nanopore. GCE allows for rapid and efficient introduction of bioorthogonal reactive site (i.e., azide) to the pore rim, and conjugation of single stranded DNA or lysozyme was demonstrated. The lysozyme-conjugated pore was further used for the discrimination of different oligosaccharides, demonstrating a sensing capacity that a bare MspA nanopore does not possess. GCE with bioorthogonal handles, which has never been previously applied in the preparation of nanopores, is a versatile strategy for pore engineering and may further expand the application scenarios of nanopores.
在单分子传感中，特异性地修饰含天然氨基酸的生物纳米孔对于其功能化至关重要。然而，目前的纳米孔工程方法受到了极大的限制，只能选择有限的方法，并且极难引入非天然的化学组分。在此，我们报告了遗传密码扩展（GCE）策略，用于将非天然氨基酸（UAA）引入八聚体分枝杆菌 smegmatis 孔蛋白 A（MspA）纳米孔中。GCE 允许快速有效地在孔边缘引入生物正交反应性位点（即叠氮化物），并证明了单链 DNA 或溶菌酶的连接。进一步将溶菌酶偶联的孔用于区分不同的寡糖，证明了裸 MspA 纳米孔所不具备的传感能力。带有生物正交接头的 GCE 以前从未应用于纳米孔的制备，是一种通用的纳米孔工程策略，可能会进一步扩展纳米孔的应用场景。
