Covalent Inhibitors of Protein–Protein Interactions Targeting Lysine, Tyrosine, or Histidine Residues

We have recently reported a series of Lys-covalent agents targeting the BIR3 domain of the X-linked inhibitor of apoptosis protein (XIAP) using a benzamide-sulfonyl fluoride warhead. Using XIAP as a model system, we further investigated a variety of additional warheads that can be easily incorporated into binding peptides and analyzed their ability to form covalent adducts with lysine and other amino acids, including tyrosine, histidine, serine, and threonine, using biochemical and biophysical assays. Moreover, we tested aqueous, plasma stability, cell permeability, and cellular efficacy of the most effective agents. These studies identified aryl-fluoro sulfates as likely the most suitable electrophiles to effectively form covalent adducts with Lys, Tyr, and His residues, given that these agents were cell permeable and stable in aqueous buffer and in plasma. Our studies contain a number of general findings that open new possible avenues for the design of potent covalent protein–protein interaction antagonists.本研究团队近期报道了一系列以苯甲酰胺磺酰氟为亲电弹头，靶向X连锁凋亡抑制蛋白（X-linked inhibitor of apoptosis protein, XIAP）BIR3结构域的赖氨酸共价结合剂。以XIAP为模型体系，我们进一步探究了多种可便捷整合至结合肽中的新型亲电弹头，并通过生化与生物物理实验分析了这些弹头与赖氨酸（Lys）、酪氨酸（Tyr）、组氨酸（His）、丝氨酸及苏氨酸等氨基酸形成共价加合物的能力。此外，我们还对筛选得到的最优制剂的水溶液稳定性、血浆稳定性、细胞通透性及细胞活性进行了检测。研究发现，芳基氟硫酸盐或许是与赖氨酸（Lys）、酪氨酸（Tyr）及组氨酸（His）残基形成高效共价加合物的最优亲电试剂，这类制剂兼具细胞通透性，且在水溶液缓冲液与血浆中均保持稳定。本研究所得的多项一般性结论，可为强效共价蛋白质-蛋白质相互作用拮抗剂的设计开辟全新方向。