Ultrafast fluorescence depolarisation in green fluorescence protein tandem dimers as hydrophobic environment sensitive probes.
Ultrafast fluorescence depolarisation in green fluorescence protein tandem dimers as hydrophobic environment sensitive probes.
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DOI：
10.1039/d3cp01765f
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发表时间：
2023-07-26
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--
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超快光子学的进步使得能够在亚纳秒时间尺度上监测生物化学相互作用。此外，荧光蛋白分子间能量转移的皮秒动力学已被观察到。在这里，我们提出了一种基因编码的荧光传感器，可以检测疏水性的变化，通过监测超快荧光去极化的发展。我们的传感器是由一对二聚体增强型绿色荧光蛋白（dEGFPs）连接的灵活的氨基酸接头。我们表明，二聚化被干扰的两种蛋白质的疏水相互作用的甘油的加入。时间分辨荧光各向异性揭示了系统的衰减超快荧光去极化时，传感器暴露于增加甘油浓度。这表明随着疏水性增加，串联二聚体内的dEGFP配对减少。蛋白质荧光团的解配对显著改变了蛋白质之间的能量转移速率，导致传感器的极限各向异性增加。增加甘油浓度导致dEGFP-TD中超快荧光去极化的系统衰减。
Advances in ultra-fast photonics have enabled monitoring of biochemical interactions on a sub nano-second time scale. In addition, picosecond dynamics of intermolecular energy transfer in fluorescent proteins has been observed. Here, we present the development of a genetically encoded fluorescent sensor that can detect changes in hydrophobicity by monitoring ultrafast fluorescence depolarisation. Our sensor is composed of a pair of dimeric enhanced green fluorescent proteins (dEGFPs) linked by a flexible amino-acid linker. We show dimerisation is perturbed by the addition of glycerol which interferes with the hydrophobic interaction of the two proteins. Time-resolved fluorescence anisotropy revealed a systematic attenuation of ultrafast fluorescence depolarisation when the sensor was exposed to increasing glycerol concentrations. This suggests that as hydrophobicity increases, dEGFP pairing decreases within a tandem dimer. Un-pairing of the protein fluorophores dramatically alters the rate of energy transfer between the proteins, resulting in an increase in the limiting anisotropy of the sensor. Increasing glycerol concentrations led to a systematic attenuation of ultrafast fluorescence depolarisation in dEGFP-TD.
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影响因子：
3.5
作者：
Cormack, BP;Valdivia, RH;Falkow, S
通讯作者：
Falkow, S
影响因子：
3.7
作者：
Arpino JA;Rizkallah PJ;Jones DD
通讯作者：
Jones DD
影响因子：
4.4
作者：
STRICKLER, SJ;BERG, RA
通讯作者：
BERG, RA
DOI：
10.1074/jbc.m111.331967
发表时间：
2012-05-18
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
Hunt J;Keeble AH;Dale RE;Corbett MK;Beavil RL;Levitt J;Swann MJ;Suhling K;Ameer-Beg S;Sutton BJ;Beavil AJ
通讯作者：
Beavil AJ
影响因子：
48
作者：
Cranfill PJ;Sell BR;Baird MA;Allen JR;Lavagnino Z;de Gruiter HM;Kremers GJ;Davidson MW;Ustione A;Piston DW
通讯作者：
Piston DW