Department of Organic Chemistry, Faculty of Chemistry, Jagiellonian University, Gronostajowa 2, 30-387 Krakow, Poland.
Max Planck Institute for Biochemistry, Am Klopferspitz 18, 82152 Martinsried, Germany.
Molecules. 2020 Jul 1;25(13):3017. doi: 10.3390/molecules25133017.
Screening for small-molecule fragments that can lead to potent inhibitors of protein-protein interactions (PPIs) is often a laborious step as the fragments cannot dissociate the targeted PPI due to their low μM-mM affinities. Here, we describe an NMR competition assay called w-AIDA-NMR (weak-antagonist induced dissociation assay-NMR), which is sensitive to weak μM-mM ligand-protein interactions and which can be used in initial fragment screening campaigns. By introducing point mutations in the complex's protein that is not targeted by the inhibitor, we lower the effective affinity of the complex, allowing for short fragments to dissociate the complex. We illustrate the method with the compounds that block the Mdm2/X-p53 and PD-1/PD-L1 oncogenic interactions. Targeting the PD-/PD-L1 PPI has profoundly advanced the treatment of different types of cancers.
筛选能够导致蛋白-蛋白相互作用（PPIs）强效抑制剂的小分子片段通常是一项艰巨的任务，因为这些片段由于其低微摩尔-毫摩尔亲和力而无法解离靶向的 PPI。在这里，我们描述了一种称为 w-AIDA-NMR（弱拮抗剂诱导解离测定-NMR）的 NMR 竞争测定法，该方法对弱微摩尔-毫摩尔配体-蛋白相互作用敏感，可用于初步的片段筛选。通过在复合物的蛋白质中引入不被抑制剂靶向的点突变，我们降低了复合物的有效亲和力，从而使短片段能够解离复合物。我们用阻断 Mdm2/X-p53 和 PD-1/PD-L1 致癌相互作用的化合物来说明该方法。针对 PD-/PD-L1 PPI 的治疗已极大地推进了不同类型癌症的治疗。
