Mitochondrial dysfunction underlies the etiology of a broad spectrum of diseases including heart disease, cancer, neurodegenerative diseases, and the general aging process. Therapeutics that restore healthy mitochondrial function hold promise for treatment of these conditions. The synthetic tetrapeptide, elamipretide (SS-31), improves mitochondrial function, but mechanistic details of its pharmacological effects are unknown. Reportedly, SS-31 primarily interacts with the phospholipid cardiolipin in the inner mitochondrial membrane. Here we utilize chemical cross-linking with mass spectrometry to identify protein interactors of SS-31 in mitochondria. The SS-31-interacting proteins, all known cardiolipin binders, fall into two groups, those involved in ATP production through the oxidative phosphorylation pathway and those involved in 2-oxoglutarate metabolic processes. Residues cross-linked with SS-31 reveal binding regions that in many cases, are proximal to cardiolipin–protein interacting regions. These results offer a glimpse of the protein interaction landscape of SS-31 and provide mechanistic insight relevant to SS-31 mitochondrial therapy.

线粒体功能障碍是多种疾病的病因，包括心脏病，癌症，神经退行性疾病和一般衰老过程。恢复健康的线粒体功能的疗法有望治疗这些疾病。合成的四肽Elamipretide（SS-31）可改善线粒体功能，但尚不清楚其药理作用的机理细节。据报道，SS-31主要与线粒体内膜中的磷脂心磷脂相互作用。在这里，我们利用质谱进行化学交联来鉴定线粒体中SS-31的蛋白质相互作用物。与SS-31相互作用的蛋白（所有已知的心磷脂结合剂）分为两组，那些通过氧化磷酸化途径参与ATP产生的物质，以及那些与2-氧戊二酸酯代谢过程有关的物质。与SS-31交联的残基揭示了在许多情况下靠近心磷脂-蛋白质相互作用区域的结合区域。这些结果提供了对SS-31蛋白质相互作用情况的一瞥，并提供了与SS-31线粒体疗法相关的机理性见解。