School of Chemistry and Chemical Engineering, Shanghai Key Laboratory of Electrical Insulation and Thermal Aging, State Key Laboratory of Metal Matrix Composites , Shanghai Jiao Tong University , 800 Dongchuan Road , Shanghai 200240 , People's Republic of China.
J Phys Chem B. 2020 Feb 6;124(5):871-880. doi: 10.1021/acs.jpcb.9b11329. Epub 2020 Jan 22.
The tremendous gap of fluorescence emission of synthetic green fluorescent protein (GFP) chromophore to the protein itself makes it impossible to use for applications in molecular and cellular imaging. Here, we developed an efficient methodology to enhance the photoluminescence response of synthetic GFP chromophore analogues by constructing dual-self-restricted chromophores. Single self-restricted chromophores were first generated with 2,5-dimethoxy substitution on the aromatic ring, which were further modified with phenyl or 2,5-dimethoxy phenyl to form dual-self-restricted chromophores. These two chromophores showed an obvious solvatofluorochromic color palette across blue to yellow with a maximum emission Stokes shift of 95 nm and dramatically enhanced fluorescence emission in various aprotic solvents, especially in hexane, where the QY reached around 0.6. Importantly, in acetonitrile and dimethyl sulfoxide, the fluorescence QYs of both chromophores were over 0.22, which were the highest reported so far in high polar organic solvents. Meanwhile, the fluorescence lifetimes also improved obviously with the maximum of around 4.5 ns. Theoretical calculations revealed a more favorable Mülliken atomic charge translocation over the double-bond bridge and illustrated much higher energy barriers for the Z/E photoisomerization together with larger bond orders compared with the GFP core chromophore, -HBDI. Our work significantly improved the fluorescence emission of synthetic GFP chromophore analogues in polar solvents while reserved the multicolor emitting function, providing a solid molecular motif for engineering high-performance fluorescent probes.
合成绿色荧光蛋白（GFP）发色团与蛋白质本身之间巨大的荧光发射差距使得其无法应用于分子和细胞成像。在这里，我们开发了一种有效的方法，通过构建双自限制发色团来增强合成 GFP 发色团类似物的光致发光响应。首先，在芳环上进行 2,5-二甲氧基取代，生成单自限制发色团，然后用苯基或 2,5-二甲氧基苯基进一步修饰，形成双自限制发色团。这两种发色团在蓝色到黄色之间表现出明显的溶剂化变色，最大发射斯托克斯位移为 95nm，并在各种非质子溶剂中显著增强了荧光发射，特别是在己烷中，QY 达到约 0.6。重要的是，在乙腈和二甲基亚砜中，两种发色团的荧光 QY 均超过 0.22，这是迄今为止在高极性有机溶剂中报道的最高值。同时，荧光寿命也明显提高，最大值约为 4.5ns。理论计算揭示了双键桥更有利于 Mülliken 原子电荷迁移，并说明了与 GFP 核心发色团 -HBDI 相比，Z/E 光致异构化的能量势垒更高，键序更大。我们的工作显著提高了极性溶剂中合成 GFP 发色团类似物的荧光发射，同时保留了多色发射功能，为工程高性能荧光探针提供了坚实的分子基元。
