Page Asher N, George Nicholas P
Department of Biochemistry, University of Wisconsin, Madison, WI, USA.
Methods Mol Biol. 2012;922:169-74. doi: 10.1007/978-1-62703-032-8_12.
Surface plasmon resonance (SPR) is a widely employed technique for studying protein-protein interactions. Here, we describe a method for the analysis of single-stranded DNA binding protein (SSB)-heterologous protein interactions by SPR. This method avoids several pitfalls often associated with SPR, particularly difficulties in immobilizing the protein while still allowing for facile regeneration of the sensor chip surface for subsequent experiments. Essentially, the method entails immobilizing a biotinylated single-stranded DNA oligo onto the chip surface, which is then bound by SSB prior to analyte addition to the SSB-coated chip. This allows for rapid qualitative and detailed quantitative analysis of both equilibrium and kinetic parameters of the SSB-protein interaction.
表面等离子体共振（SPR）是一种广泛应用于研究蛋白质-蛋白质相互作用的技术。在此，我们描述了一种通过SPR分析单链DNA结合蛋白（SSB）与异源蛋白相互作用的方法。该方法避免了一些通常与SPR相关的陷阱，特别是在固定蛋白质时遇到的困难，同时仍能使传感器芯片表面便于再生以用于后续实验。从本质上讲，该方法需要将生物素化的单链DNA寡核苷酸固定在芯片表面，然后在向包被有SSB的芯片中添加分析物之前，使SSB与之结合。这使得能够对SSB-蛋白质相互作用的平衡和动力学参数进行快速定性和详细定量分析。
