Wang Ruowen, Lu Danqing, Bai Huarong, Jin Cheng, Yan Guobei, Ye Mao, Qiu Liping, Chang Rongshan, Cui Cheng, Liang Hao, Tan Weihong
Molecular Sciences and Biomedicine Laboratory, State Key Laboratory for Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering and College of Biology, Collaborative Innovation Center for Molecular Engineering and Theranostics, Hunan University, Changsha 410082, China; Departments of Chemistry and Department of Physiology and Functional Genomics, Center for Research at the Bio/Nano Interface, Shands Cancer Center, UF Genetics Institute and McKnight Brain Institute, University of Florida, Gainesville, Florida32611-7200, United States.
Molecular Sciences and Biomedicine Laboratory, State Key Laboratory for Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering and College of Biology, Collaborative Innovation Center for Molecular Engineering and Theranostics, Hunan University, Changsha 410082, China.
Chem Sci. 2016 Mar 1;7(3):2157-2161. doi: 10.1039/C5SC02631H. Epub 2015 Dec 10.
Conjugation of DNAs to defined locations on a protein surface will offer powerful tools for positioning functional groups and molecules in biological and biomedical studies. However, tagging protein with DNA is challenging in physiological environments, which requires a bioorthogonal approach. Here we report a chemical solution to selectively conjugate DNA aptamer with a protein by protein-aptamer template (PAT)-directed reactions. Since protein-aptamer interactions are bioorthogonal, we exploit PAT as a unique platform for specific DNA-protein cross-linking. We develop a series of modified oligonucleotides for PAT-directed reactions and screen out F-carboxyl as a suitable functionality for selective and site-specific conjugation. The functionality is incorporated into aptamers by our F-carboxyl phosphoramidite with easy synthesis. We also demonstrate the necessity of a linker between the reactive functionality and the aptamer sequences.
将DNA与蛋白质表面的特定位置进行共轭连接，将为生物和生物医学研究中定位官能团和分子提供强大的工具。然而，在生理环境中用DNA标记蛋白质具有挑战性，这需要一种生物正交方法。在此，我们报告了一种化学解决方案，通过蛋白质-适配体模板（PAT）导向反应将DNA适配体与蛋白质选择性地共轭连接。由于蛋白质-适配体相互作用是生物正交的，我们利用PAT作为特异性DNA-蛋白质交联的独特平台。我们开发了一系列用于PAT导向反应的修饰寡核苷酸，并筛选出F-羧基作为选择性和位点特异性共轭连接的合适官能团。通过我们易于合成的F-羧基亚磷酰胺，该官能团被引入到适配体中。我们还证明了反应性官能团与适配体序列之间连接子的必要性。
