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首页> 外文期刊>Proceedings of the National Academy of Sciences of the United States of America >EosFP, a fluorescent marker protein with UV-inducible green-to-red fluorescence conversion.
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EosFP, a fluorescent marker protein with UV-inducible green-to-red fluorescence conversion.

机译：EosFP，一种荧光标记蛋白，具有紫外线诱导的绿色到红色荧光转换。

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摘要

A gene encoding a fluorescent protein from the stony coral Lobophyllia hemprichii has been cloned in Escherichia coli and characterized by biochemical and biophysical methods. The protein, which we named EosFP, emits strong green fluorescence (516 nm) that changes to red (581 nm) upon near-UV irradiation at approximately 390 nm because of a photo-induced modification involving a break in the peptide backbone next to the chromophore. Single-molecule fluorescence spectroscopy shows that the wild type of EosFP is tetrameric, with strong Forster resonance coupling among the individual fluorophores. We succeeded in breaking up the tetramer into AB and AC subunit dimers by introducing the single point mutations V123T and T158H, respectively, and the combination of both mutations yielded functional monomers. Fusion constructs with a variety of proteins were prepared and expressed in human cells, showing that normal biological functions were retained. The possibility to locally change the emission wavelength by focused UV light makes EosFP a superb marker for experiments aimed at tracking the movements of biomolecules within the living cell.
机译：已经从大肠杆菌中克隆了一种编码来自石质珊瑚海豚的荧光蛋白的基因的基因，并通过生化和生物物理方法对其进行了表征。我们将这种蛋白质命名为EosFP，它发出强绿色荧光（516 nm），在大约390 nm的近紫外光照射下，由于光诱导的修饰，包括邻近肽骨架的肽断裂，该荧光会变为红色（581 nm）。发色团。单分子荧光光谱表明，野生型EosFP是四聚体，在各个荧光团之间具有很强的Forster共振耦合。通过分别引入单点突变V123T和T158H，我们成功地将四聚体分解为AB和AC亚基二聚体，并且两个突变的组合产生了功能单体。制备了具有多种蛋白质的融合构建体并在人细胞中表达，表明保留了正常的生物学功能。 EosFP可以通过聚焦的紫外线局部改变发射波长，从而使EosFP成为旨在跟踪活细胞内生物分子运动的实验的绝佳标记。

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