Department of Biomedical Engineering, Cornell University, Ithaca, New York 14853, USA.
Bioconjug Chem. 2009 Aug 19;20(8):1482-9. doi: 10.1021/bc9000118. Epub 2009 Aug 3.
We report a highly specific, robust, and generic method for noncovalent labeling of cellular proteins with highly fluorescent core-shell silica nanoparticles termed C dots. Our approach uses short genetically engineered peptides with affinity for silica (GEPS) that are site-specifically introduced at the termini or in loops of cellular proteins. Because GEPS are absent from native cell surface proteins, GEPS-tagged recombinant proteins can be selectively and rapidly labeled with fluorescent C dots. To demonstrate the versatility of our method, we targeted 30 nm C dots to two structurally distinct integral outer membrane proteins in Escherichia coli, FhuA and OmpX. Efficient labeling was achieved in 15 min or less and was observed to be highly sensitive and specific. This strategy provides a powerful technique, comparable to other chemical and biological labeling strategies, for efficient and quantitative investigation of protein function in live biological cells.
我们报道了一种高度特异、稳健且通用的方法，用于通过称为 C 点的高度荧光核壳二氧化硅纳米颗粒对细胞蛋白进行非共价标记。我们的方法使用对硅具有亲和力的短基因工程肽（GEPS），这些肽在细胞蛋白的末端或环上进行特异性引入。由于 GEPS 不存在于天然细胞表面蛋白中，因此可以选择性地快速用荧光 C 点标记 GEPS 标记的重组蛋白。为了证明我们方法的多功能性，我们将 30nm 的 C 点靶向到大肠杆菌中两种结构不同的完整外膜蛋白 FhuA 和 OmpX。在 15 分钟或更短的时间内实现了有效的标记，并且观察到其具有高度的灵敏性和特异性。该策略提供了一种强大的技术，可与其他化学和生物学标记策略相媲美，用于在活生物细胞中高效且定量地研究蛋白质功能。
