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首页> 外文期刊>Proceedings of the National Academy of Sciences of the United States of America >Reversible single-molecule photoswitching in the GFP-like fluorescent protein Dronpa
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Reversible single-molecule photoswitching in the GFP-like fluorescent protein Dronpa

机译：GFP样荧光蛋白Dronpa中的可逆单分子光开关

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摘要

Reversible photoswitching of individual molecules has been demonstrated fora number of mutants of the green fluorescent protein (GFP). To date, however, a limited number of switching events with slow response to light have been achieved at the single-molecule level. Here, we report reversible photoswitching characteristics observed in individual molecules of Dronpa, a mutant of a GFP-like fluorescent protein that was cloned from a coral Pectiniidae. Ensemble spectroscopy shows that intense irradiation at 488 nm changes Dronpa to a dim protonated form, but even weak irradiation at 405 nm restores it to the bright deprotonated form. Although Dronpa exists in an acid-base equilibrium, only the photoincluced protonated form shows the switching behavior. At the single-molecule level, 488- and 405-nm lights can be used to drive the molecule back and forth between the bright and dim states. Such reversible photoswitching could be repeated > 100 times. The response speed to irradiation depends almost linearly on the irradiation power, with the response time being in the order of milliseconds. The perfect reversibility of the Dronpa photoswitching allows us to propose a detailed model, which quantitatively describes interconversion among the various states. The fast response of Dronpa to light holds great promise for following fast diffusion or transport of signaling molecules in live cells.
机译：对于绿色荧光蛋白（GFP）的许多突变体，已经证明了单个分子的可逆光开关。然而，迄今为止，在单分子水平上已经实现了有限数量的对光的响应较慢的开关事件。在这里，我们报告在Dronpa的单个分子中观察到的可逆光开关特性，Dronpa是从珊瑚果蝇科克隆的GFP样荧光蛋白的突变体。集成光谱显示，在488 nm处的强辐射使Dronpa变成了暗淡的质子化形式，但是即使在405 nm处的弱照射也将其还原为明亮的去质子化形式。尽管Dronpa处于酸碱平衡状态，但仅光吸收质子化形式显示出转换行为。在单分子水平上，可以使用488 nm和405 nm的光在明亮状态和暗淡状态之间来回驱动分子。这种可逆的光开关可以重复> 100次。辐照的响应速度几乎线性地取决于辐照功率，响应时间约为毫秒。 Dronpa光电开关的完美可逆性使我们能够提出一个详细的模型，该模型定量描述了各种状态之间的相互转换。 Dronpa对光的快速响应为信号分子在活细胞中的快速扩散或运输提供了广阔的前景。

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