Photophysics of the red-form Kaede chromophore.
Photophysics of the red-form Kaede chromophore.
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DOI：
10.1039/d3sc00368j
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发表时间：
2023-04-05
期刊：
影响因子：
8.4
通讯作者：
中科院分区：
文献类型：
--
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绿色荧光蛋白（GFP）推动了生物成像的革命性进展。光转换荧光蛋白（Photoconvertible fluorescent proteins，PCFPs）是FP家族的一个重要分支，Kaede是其原型。独特的是，PCFP可以在紫外线照射下永久地从绿色转换为红色发射形式，从而促进了位点特异性光标记和蛋白质追踪的应用。FP的优化和利用需要了解发色团的光物理和光化学行为。因此，主要的GFP发色团一直是激烈的实验和理论研究的主题。相比之下，红色发光PCFP发色团的光物理学基本上未被研究。在这里，我们提出了一个详细的调查的激发态性质的Kaede发色团在溶液中，利用稳态测量，超快时间分辨电子和振动光谱，和电子结构理论。其激发态动力学与亲本GFP的激发态动力学非常不同。最值得注意的是，PCFP发色团具有高度复杂的波长依赖性荧光衰减和比GFP发色团长一个数量级的平均寿命。瞬态电子和振动光谱表明，这些动力学产生的激发态构象的光谱和动力学不同，但化学相似的范围。这些构象填充直接通过激发一个单一的构象的基态结构的广泛的热分布，这表明一个激发态的潜在表面与几个极小。温度依赖性证实了激发态表面势垒的存在，并揭示了无辐射衰变机制是内转换。这些实验观察结果与假设简单基态势能面访问具有多个最小值的复杂激发态的模型一致。负责光学突出蛋白Kaede中颜色切换的发色团具有出乎意料的复杂激发态动力学，在这里进行测量和分析。这将为开发新的成像蛋白提供信息。
The green fluorescent protein (GFP) drove revolutionary progress in bioimaging. Photoconvertible fluorescent proteins (PCFPs) are an important branch of the FP family, of which Kaede is the prototype. Uniquely, PCFPs can be permanently switched from green to red emitting forms on UV irradiation, facilitating applications in site-specific photolabelling and protein tracking. Optimisation and exploitation of FPs requires understanding of the photophysical and photochemical behaviour of the chromophore. Accordingly, the principal GFP chromophore has been the subject of intense experimental and theoretical investigation. In contrast, the photophysics of the red emitting PCFP chromophore are largely unstudied. Here we present a detailed investigation of the excited-state properties of the Kaede chromophore in solution, utilising steady state measurements, ultrafast time-resolved electronic and vibrational spectroscopies, and electronic structure theory. Its excited state dynamics are very different to those of the parent GFP. Most remarkably, the PCFP chromophore has highly complex wavelength-dependent fluorescence decays and a mean lifetime an order of magnitude longer than the GFP chromophore. Transient electronic and vibrational spectroscopies suggest that these dynamics arise from a range of excited-state conformers that are spectrally and kinetically distinct but chemically similar. These conformers are populated directly by excitation of a broad thermal distribution of ground state structures about a single conformer, suggesting an excited-state potential surface with several minima. Temperature-dependence confirms the existence of barriers on the excited-state surface and reveals the radiationless decay mechanism to be internal conversion. These experimental observations are consistent with a model assuming a simple ground state potential energy surface accessing a complex excited state possessing multiple minima. The chromophore responsible for colour switching in the optical highlighting protein Kaede has unexpectedly complicated excited state dynamics, which are measured and analysed here. This will inform the development of new imaging proteins.
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影响因子：
2.8
作者：
Baleeva, Nadezhda S.;Myannik, Ksenia A.;Baranov, Mikhail S.
通讯作者：
Baranov, Mikhail S.
影响因子：
15
作者：
Baranov, Mikhail S.;Lukyanov, Konstantin A.;Solntsev, Kyril M.
通讯作者：
Solntsev, Kyril M.
DOI：
10.1073/pnas.0500489102
发表时间：
2005-07-05
影响因子：
11.1
作者：
Habuchi, S;Ando, R;Hofkens, J
通讯作者：
Hofkens, J
荧光蛋白红色 Kaede 发色团；一步法高产合成及在太阳能电池中的潜在应用
影响因子：
4.9
作者：
Chuang, Wei-Ti;Chen, Bo-So;Chou, Pi-Tai
通讯作者：
Chou, Pi-Tai
影响因子：
5.6
作者：
Hayashi, Ikuko;Mizuno, Hideaki;Ikura, Mitsuhiko
通讯作者：
Ikura, Mitsuhiko