Velazquez-Campoy Adrian, Leavitt Stephanie A, Freire Ernesto
Institute of Biocomputation and Physics of Complex Systems (BIFI), Joint Unit IQFR-CSIC-BIFI, Universidad de Zaragoza, Zaragoza, Spain,
Methods Mol Biol. 2015;1278:183-204. doi: 10.1007/978-1-4939-2425-7_11.
The analysis of protein-protein interactions has attracted the attention of many researchers from both a fundamental point of view and a practical point of view. From a fundamental point of view, the development of an understanding of the signaling events triggered by the interaction of two or more proteins provides key information to elucidate the functioning of many cell processes. From a practical point of view, understanding protein-protein interactions at a quantitative level provides the foundation for the development of antagonists or agonists of those interactions. Isothermal Titration Calorimetry (ITC) is the only technique with the capability of measuring not only binding affinity but the enthalpic and entropic components that define affinity. Over the years, isothermal titration calorimeters have evolved in sensitivity and accuracy. Today, TA Instruments and MicroCal market instruments with the performance required to evaluate protein-protein interactions. In this methods paper, we describe general procedures to analyze heterodimeric (porcine pancreatic trypsin binding to soybean trypsin inhibitor) and homodimeric (bovine pancreatic α-chymotrypsin) protein associations by ITC.
从基础和实际两个角度来看，蛋白质-蛋白质相互作用的分析都吸引了众多研究人员的关注。从基础角度而言，深入了解由两种或更多种蛋白质相互作用引发的信号事件，可为阐明许多细胞过程的功能提供关键信息。从实际角度来说，在定量水平上理解蛋白质-蛋白质相互作用，为开发这些相互作用的拮抗剂或激动剂奠定了基础。等温滴定量热法（ITC）是唯一不仅能够测量结合亲和力，还能测量定义亲和力的焓和熵成分的技术。多年来，等温滴定量热仪在灵敏度和准确性方面不断发展。如今，TA仪器公司和MicroCal公司销售的仪器具备评估蛋白质-蛋白质相互作用所需的性能。在这篇方法论文中，我们描述了通过ITC分析异源二聚体（猪胰蛋白酶与大豆胰蛋白酶抑制剂结合）和同源二聚体（牛胰α-糜蛋白酶）蛋白质缔合的一般程序。
