Shedding light on ultrafast ring-twisting pathways of halogenated GFP chromophores from the excited to ground state
Shedding light on ultrafast ring-twisting pathways of halogenated GFP chromophores from the excited to ground state
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DOI：
10.1039/d1cp02140k
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发表时间：
2021-06-28
影响因子：
3.3
通讯作者：
Fang, Chong
中科院分区：
文献类型：
--
作者：
Boulanger, Sean A.;Chen, Cheng;Fang, Chong
由于大约三十年来绿色荧光蛋白（GFP）彻底改变了分子和细胞生物学，人们对理解、设计和控制GFP发色团的荧光特性（即，HBDI）衍生物从蛋白质基质到溶液中。在这些跨学科的努力中，HBDI衍生物的激发态动力学的阐明是将光诱导过程和荧光量子产率（QYY）相关联的关键。本文采用稳态电子光谱、飞秒瞬态吸收（fs-TA）、飞秒受激拉曼光谱（FSRS）和量子计算等方法，研究了一系列单卤代和二卤代HBDI衍生物（X = F，Cl，Br，2F，2Cl，2Br）在碱性水溶液中的反应，获得了对反应物理坐标的新认识.在激发态下，卤代“松软”发色团表现出反重原子效应，这反映在电子自旋Y与弗兰克-康登能量（E-FC）或斯托克斯位移之间的强相关性，以及k（nr）与E-FC之间的强相关性，以及迅速分叉为I环（主要）和P环（次要）扭转运动。在基态，这两个环扭转运动变得更容易受到立体和表现出光谱签名从卤素依赖热基态吸收带衰减TA数据。我们设想这种类型的卤代HBDI衍生物的系统分析，以提供指导原则的GFP发色团的位点特异性修饰，并扩大设计空间，在整个光循环中具有可辨别的光谱特征的水溶液中的更明亮的和潜在的光开关的有机化学探针。
Since green fluorescent protein (GFP) has revolutionized molecular and cellular biology for about three decades, there has been a keen interest in understanding, designing, and controlling the fluorescence properties of GFP chromophore (i.e., HBDI) derivatives from the protein matrix to solution. Amongst these cross-disciplinary efforts, the elucidation of excited-state dynamics of HBDI derivatives holds the key to correlating the light-induced processes and fluorescence quantum yield (FQY). Herein, we implement steady-state electronic spectroscopy, femtosecond transient absorption (fs-TA), femtosecond stimulated Raman spectroscopy (FSRS), and quantum calculations to study a series of mono- and dihalogenated HBDI derivatives (X = F, Cl, Br, 2F, 2Cl, and 2Br) in basic aqueous solution, gaining new insights into the photophysical reaction coordinates. In the excited state, the halogenated "floppy" chromophores exhibit an anti-heavy atom effect, reflected by strong correlations between FQY vs. Franck-Condon energy (E-FC) or Stokes shift, and k(nr)vs. E-FC, as well as a swift bifurcation into the I-ring (major) and P-ring (minor) twisting motions. In the ground state, both ring-twisting motions become more susceptible to sterics and exhibit spectral signatures from the halogen-dependent hot ground-state absorption band decay in TA data. We envision this type of systematic analysis of the halogenated HBDI derivatives to provide guiding principles for the site-specific modification of GFP chromophores, and expand design space for brighter and potentially photoswitchable organic chemical probes in aqueous solution with discernible spectral signatures throughout the photocycle.