通过 NMR 光谱法测定天然无序蛋白质的结合动力学。
Determining Binding Kinetics of Intrinsically Disordered Proteins by NMR Spectroscopy.
机构信息
Department of Integrative Structural and Computational Biology, The Scripps Research Institute, La Jolla, CA, USA.
Department of Life Sciences, Graduate School of Arts and Sciences, The University of Tokyo, Tokyo, Japan.
出版信息
Methods Mol Biol. 2020;2141:663-681. doi: 10.1007/978-1-0716-0524-0_34.
The unique structural flexibility of intrinsically disordered proteins (IDPs) is central to their diverse functions in cellular processes. Protein-protein interactions involving IDPs are frequently transient and dynamic in nature. Nuclear magnetic resonance (NMR) spectroscopy is an especially powerful tool for characterizing the structural propensities, dynamics, and interactions of IDPs. Here we describe applications of the Carr-Purcell-Meiboom-Gill (CPMG) relaxation dispersion experiment in combination with NMR titrations to characterize the kinetics and mechanisms of interactions between intrinsically disordered proteins and their targets. We illustrate the method with reference to interactions between the activation domain of the human T-cell leukemia virus type-I (HTLV-1) basic leucine zipper protein (HBZ) and its cellular binding partner, the KIX domain of the transcriptional coactivator CBP.
无规卷曲蛋白（IDPs）独特的结构灵活性是其在细胞过程中发挥多种功能的核心。涉及 IDPs 的蛋白质-蛋白质相互作用通常具有瞬时和动态的性质。核磁共振（NMR）光谱是表征 IDPs 的结构倾向、动态和相互作用的特别强大的工具。在这里，我们描述了 Carr-Purcell-Meiboom-Gill（CPMG）弛豫分散实验与 NMR 滴定的结合在表征 IDPs 与其靶标之间相互作用的动力学和机制中的应用。我们参考人类 T 细胞白血病病毒 I 型（HTLV-1）碱性亮氨酸拉链蛋白（HBZ）的激活结构域与其细胞结合伴侣转录共激活因子 CBP 的 KIX 结构域之间的相互作用来说明该方法。
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