Analysis of Multivalent IDP Interactions: Stoichiometry, Affinity, and Local Concentration Effect Measurements.
Analysis of Multivalent IDP Interactions: Stoichiometry, Affinity, and Local Concentration Effect Measurements.
复制标题
DOI：
10.1007/978-1-0716-0524-0_23
复制
发表时间：
2020
影响因子：
--
通讯作者：
Samuel Sparks;R. Hayama;M. Rout;D. Cowburn
中科院分区：
文献类型：
--
作者：
Samuel Sparks;R. Hayama;M. Rout;D. Cowburn
核磁共振（NMR）滴定法和等温滴定量热法可以结合起来提供多价内在无序蛋白（IDP）相互作用如何涉及焓熵平衡的评估。在这里，我们描述了潜在的技术细节和额外的方法，如动态光散射分析，需要评估这些反应。我们将此应用于一个中心相互作用，涉及含有多个苯丙氨酸-甘氨酸重复序列的聚核孔蛋白（gf - nups）的无序区域，这是特别感兴趣的，因为它们与核转运因子（NTRs）的相互作用是核孔复合物（NPC）介导的大分子的矛盾快速但高度选择性运输的基础。这些分析表明，低每FG基序亲和力和焓熵平衡的结合阻止了FG- nups和NTRs之间的高亲和相互作用，而大量的FG基序促进了FG- ntr之间的频繁接触，从而提高了选择性。
Nuclear magnetic resonance (NMR) titration and isothermal titration calorimetry can be combined to provide an assessment of how multivalent intrinsically disordered protein (IDP) interactions can involve enthalpy–entropy balance. Here, we describe the underlying technical details and additional methods, such as dynamic light scattering analysis, needed to assess these reactions. We apply this to a central interaction involving the disordered regions of phe–gly nucleoporins (FG-Nups) that contain multiple phenylalanine–glycine repeats which are of particular interest, as their interactions with nuclear transport factors (NTRs) underlie the paradoxically rapid yet also highly selective transport of macromolecules mediated by the nuclear pore complex (NPC). These analyses revealed that a combination of low per-FG motif affinity and the enthalpy–entropy balance prevents high-avidity interaction between FG-Nups and NTRs while the large number of FG motifs promotes frequent FG–NTR contacts, resulting in enhanced selectivity.