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Abstract：
Understanding the kinetics of protein interactions plays a key role in biology with significant implications for the design of analytical methods for disease monitoring and diagnosis in medical care, research and industrial applications. Herein, we introduce a novel plasmonic approach to study the binding kinetics of protein-ligand interactions following the formation of silver nanoparticles (Ag NPs) dimers by UV-Vis spectroscopy that can be used as probes for antigen detection and quantification. To illustrate and test the method, the kinetics of the prototype biotin-streptavidin (Biot-STV) pair interaction was studied. Controlled aggregates (dimers) of STV functionalized Ag NPs were produced by adding stoichiometric quantities of gliadin-specific biotinylated antibodies (IgG-Biot). The dimerization kinetics was studied in a systematic way as a function of Ag NPs size and at different concentrations of IgG-Biot. The kinetics data have shown to be consistent with a complex reaction mechanism in which only the Ag NPs attached to the IgG-Biot located in a specific STV site are able to form dimers. These results help in elucidating a complex reaction mechanism involved in the dimerization kinetics of functionalized Ag NPs, which can serve as probes in surface plasmon resonance-based bioassays for the detection and quantification of different biomarkers or analytes of interest.
摘要：
了解蛋白质相互作用的动力学在生物学中起着关键作用，对医疗保健中疾病监测和诊断的分析方法的设计具有重要意义。研究和工业应用。在这里，我们介绍了一种新的等离子体方法来研究蛋白质-配体相互作用的结合动力学后形成的银纳米颗粒（AgNP）二聚体的紫外-可见光谱，可用作探针抗原检测和定量。为了说明和测试方法，研究了原型生物素-链霉亲和素（Biot-STV）对相互作用的动力学。通过添加化学计量的麦醇溶蛋白特异性生物素化抗体(IgG-Biot)产生STV官能化AgNP的受控聚集体(二聚体)。在不同浓度的IgG-Biot下，以系统的方式研究了二聚化动力学与AgNP大小的关系。动力学数据已显示与复杂反应机制一致,其中只有连接到位于特定STV位点的IgG-Biot的AgNP能够形成二聚体。这些结果有助于阐明与功能化AgNP的二聚动力学有关的复杂反应机理，其可以用作基于表面等离子体共振的生物测定中的探针，用于检测和定量不同的生物标志物或感兴趣的分析物。
