Radical-Triggered Reaction Mechanism of the Green-to-Red Photoconversion of EosFP
Radical-Triggered Reaction Mechanism of the Green-to-Red Photoconversion of EosFP
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EosFP 绿光红光转换的自由基触发反应机制
DOI：
10.1021/acs.jpcb.0c04587
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发表时间：
2020
期刊：
影响因子：
--
通讯作者：
Fare C
中科院分区：
文献类型：
--
作者：
Fare C
在绿色到红色的光致变色荧光蛋白EosFP的光转换的反应中间体已被观察到使用高强度连续蓝色照明。通过光诱导的积累，继续转化为绿色的形式，在随后的黑暗中，puronecular含有酪氨酰基自由基，虽然在电子光谱和FTIR光谱中的有害移动的功能，确定了中间体。将pH降低至5.5显著延迟了该酪氨酰中间体的衰变，这伴随着反应物和产物的电子光谱中的斯塔克位移特征。反应中间体的高频和指纹FTIR光谱区域的振动模式归属支持了一个拟议的事件序列，其中新形成的Cα C β烯键先于His-62咪唑环上的修饰，并证实了Phe-61上的C O（NH 2）产物基团。我们提出了一种反应机制，涉及酪氨酰生成通过单重态激发态介导的氧化，随后引发的共价反应的氧化的绿色生色团。
Reaction intermediates in the green-to-red photoconversion of the photochromic fluorescent protein EosFP have been observed using high-intensity continuous blue illumination. An intermediate was identified through light-induced accumulation that continues to convert the green form in subsequent darkness, putatively containing a tyrosyl radical, albeit with anomalously shifted features in both the electronic and FTIR spectra. Lowering the pH to 5.5 significantly delays the decay of this tyrosyl intermediate, which is accompanied by Stark-shifted features in the electronic spectra of reactants and products. Vibrational mode assignments for the high-frequency and fingerprint FTIR spectral regions of the reaction intermediates support a proposed sequence of events where the newly formed Cα═Cβethylenic bond precedes modifications on the His-62 imidazole ring and confirms a C═O(NH2) product group on Phe-61. We propose a reaction mechanism that involves tyrosyl generation via singlet excited-state-mediated oxidation which subsequently triggers the covalent reactions by oxidation of the green chromophore.