Triple FRET: A tool for studying long-range molecular interactions
Triple FRET: A tool for studying long-range molecular interactions
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DOI：
10.1002/cphc.200200634
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发表时间：
2003-07-14
期刊：
影响因子：
2.9
通讯作者：
Schwille, P
中科院分区：
文献类型：
--
作者：
Haustein, E;Jahnz, M;Schwille, P
大约20年前，荧光共振能量转移（FRET）被扩展到生物学问题，并被用作测量分子距离的光谱标尺。[1]然而，这种技术的实用性被限制在10到大约75 ä之间的极短距离，[2] 100 ä是最大限度。不幸的是，许多涉及蛋白质-蛋白质相互作用和大分子复合物的生物学问题远远超出了这些狭窄的维度。克服这些限制的最直观的方法是简单地添加第三个生色团，从而将该原理扩展到三色FRET（triFRET）。这种FRET级联的效率已经成功地证明了在树枝状聚合物和ssDNA中非常短的发色团间距离。[3，4]但对于更长的距离和构象更刚性的系统，这是可能的吗？传统的FRET技术有很多困难，那么能量转移效率又有多大呢？在这里，我们证明了triFRET不仅是可行的，而且有效距离可以很容易地增加到100 ä或更远;这是以前无法用传统的FRET测量。
Extended to biological questions about 20 years ago, fluorescence resonance energy transfer (FRET) has been used as a spectroscopic ruler for measuring molecular distances.[1] However, the utility of this technique has been restricted to extremely short distances between 10 to about 75 ä,[2] with 100 ä being the utmost limit. Unfortunately, many biological problems that involve protein-protein interactions and large macromolecular complexes far exceed these narrow dimensions. The most intuitive way to overcome these limitations would consist of simply adding a third chromophore, thus extending this principle to triple-colour FRET (triFRET). The efficiency of such a FRET cascade has already been successfully demonstrated for very short interchromophoric distances both in dendrimers and in ssDNA.[3, 4]But is this possible for longer distances and conformationally more rigid systems? With all the difficulties of conventional FRET techniques, what energy-transfer efficiency can be expected? Here, we show that triFRET is not only feasible, but the effective distance could easily be increased to 100ä and beyond; this was previously inaccessible with conventional FRET measurements.