Clapp Aaron R, Medintz Igor L, Mauro J Matthew, Fisher Brent R, Bawendi Moungi G, Mattoussi Hedi
Optical Sciences Division, Code 5611, U.S. Naval Research Laboratory, Washington, D.C. 20375, USA.
J Am Chem Soc. 2004 Jan 14;126(1):301-10. doi: 10.1021/ja037088b.
We used luminescent CdSe-ZnS core-shell quantum dots (QDs) as energy donors in fluorescent resonance energy transfer (FRET) assays. Engineered maltose binding protein (MBP) appended with an oligohistidine tail and labeled with an acceptor dye (Cy3) was immobilized on the nanocrystals via a noncovalent self-assembly scheme. This configuration allowed accurate control of the donor-acceptor separation distance to a range smaller than 100 A and provided a good model system to explore FRET phenomena in QD-protein-dye conjugates. This QD-MBP conjugate presents two advantages: (1) it permits one to tune the degree of spectral overlap between donor and acceptor and (2) provides a unique configuration where a single donor can interact with several acceptors simultaneously. The FRET signal was measured for these complexes as a function of both degree of spectral overlap and fraction of dye-labeled proteins in the QD conjugate. Data showed that substantial acceptor signals were measured upon conjugate formation, indicating efficient nonradiative exciton transfer between QD donors and dye-labeled protein acceptors. FRET efficiency can be controlled either by tuning the QD photoemission or by adjusting the number of dye-labeled proteins immobilized on the QD center. Results showed a clear dependence of the efficiency on the spectral overlap between the QD donor and dye acceptor. Apparent donor-acceptor distances were determined from efficiency measurements and corresponding Förster distances, and these results agreed with QD bioconjugate dimensions extracted from structural data and core size variations among QD populations.
我们使用发光的CdSe-ZnS核壳量子点（QDs）作为荧光共振能量转移（FRET）分析中的能量供体。通过非共价自组装方案，将带有寡聚组氨酸尾巴并标记有受体染料（Cy3）的工程化麦芽糖结合蛋白（MBP）固定在纳米晶体上。这种配置允许将供体-受体分离距离精确控制在小于100埃的范围内，并提供了一个良好的模型系统来探索量子点-蛋白质-染料共轭物中的FRET现象。这种量子点-MBP共轭物具有两个优点：（1）它允许调节供体和受体之间的光谱重叠程度；（2）提供了一种独特的配置，其中单个供体可以同时与多个受体相互作用。针对这些复合物测量了FRET信号，该信号是光谱重叠程度和量子点共轭物中染料标记蛋白质比例的函数。数据表明，共轭物形成后测量到了大量的受体信号，表明量子点供体和染料标记的蛋白质受体之间发生了有效的非辐射激子转移。FRET效率可以通过调节量子点的光发射或通过调整固定在量子点中心的染料标记蛋白质的数量来控制。结果表明，效率明显依赖于量子点供体和染料受体之间的光谱重叠。根据效率测量结果和相应的Förster距离确定了表观供体-受体距离，这些结果与从结构数据中提取的量子点生物共轭物尺寸以及量子点群体之间的核心尺寸变化一致。
