Combining mass spectrometry–based chemical cross-linking and complexome profiling, we analyzed the interactome of heart mitochondria. We focused on complexes of oxidative phosphorylation and found that dimeric apoptosis-inducing factor 1 (AIFM1) forms a defined complex with ∼10% of monomeric cytochrome c oxidase (COX) but hardly interacts with respiratory chain supercomplexes. Multiple AIFM1 intercross-links engaging six different COX subunits provided structural restraints to build a detailed atomic model of the COX-AIFM1₂ complex (PDBDEV_00000092). An application of two complementary proteomic approaches thus provided unexpected insight into the macromolecular organization of the mitochondrial complexome. Our structural model excludes direct electron transfer between AIFM1 and COX. Notably, however, the binding site of cytochrome c remains accessible, allowing formation of a ternary complex. The discovery of the previously overlooked COX-AIFM1₂ complex and clues provided by the structural model hint at potential roles of AIFM1 in oxidative phosphorylation biogenesis and in programmed cell death.

结合基于质谱的化学交联和复合体分析，我们分析了心脏线粒体的相互作用组。我们专注于氧化磷酸化复合物，发现二聚体凋亡诱导因子 1 (AIFM1) 与约 10% 的单体细胞色素c氧化酶 (COX)形成确定的复合物，但几乎不与呼吸链超复合物相互作用。涉及六个不同 COX 亚基的多个 AIFM1 交联提供了结构限制，以构建 COX-AIFM1 ₂的详细原子模型复杂 (PDBDEV_00000092)。因此，两种互补的蛋白质组学方法的应用为线粒体复合体的大分子组织提供了意想不到的见解。我们的结构模型不包括 AIFM1 和 COX 之间的直接电子转移。然而，值得注意的是，细胞色素c的结合位点仍然可以访问，从而形成三元复合物。先前被忽视的 COX-AIFM1 ₂复合物的发现和结构模型提供的线索暗示了 AIFM1 在氧化磷酸化生物发生和程序性细胞死亡中的潜在作用。