A serious consequence of myocardial ischemia-reperfusion injury (I/R) is oxidative damage, which causes mitochondrial dysfunction. The cascading ROS can propagate and potentially induce heme bleaching and protein cysteine sulfonation (PrSO₃H) of the mitochondrial electron transport chain. Herein we studied the mechanism of I/R-mediated irreversible oxidative injury of complex III in mitochondria from rat hearts subjected to 30-min of ischemia and 24-h of reperfusion in vivo. In the I/R region, the catalytic activity of complex III was significantly impaired. Spectroscopic analysis indicated that I/R mediated the destruction of hemes b and c + c₁ in the mitochondria, supporting I/R-mediated complex III impairment. However, no significant impairment of complex III activity and heme damage were observed in mitochondria from the risk region of rat hearts subjected only to 30-min ischemia, despite a decreased state 3 respiration. In the I/R mitochondria, carbamidomethylated C₁₂₂/C₁₂₅ of cytochrome c₁ via alkylating complex III with a down regulation of HCCS was exclusively detected, supporting I/R-mediated thioether defect of heme c₁. LC-MS/MS analysis showed that I/R mitochondria had intensely increased complex III PrSO₃H levels at the C₂₃₆ ligand of the [2Fe2S] cluster of the Rieske iron‑sulfur protein (uqcrfs1), thus impairing the electron transport activity. MS analysis also indicated increased PrSO₃H of the hinge protein at C₆₅ and of cytochrome c₁ at C₁₄₀ and C₂₂₀, which are confined in the intermembrane space. MS analysis also showed that I/R extensively enhanced the PrSO₃H of the core 1 (uqcrc1) and core 2 (uqcrc2) subunits in the matrix compartment, thus supporting the conclusion that complex III releases ROS to both sides of the inner membrane during reperfusion. Analysis of ischemic mitochondria indicated a modest reduction from the basal level of complex III PrSO₃H detected in the mitochondria of sham control hearts, suggesting that the physiologic hyperoxygenation and ROS overproduction during reperfusion mediated the enhancement of complex III PrSO₃H. In conclusion, reperfusion-mediated heme damage with increased PrSO₃H controls oxidative injury to complex III and aggravates mitochondrial dysfunction in the post-ischemic heart.

心肌缺血再灌注损伤 (I/R) 的一个严重后果是氧化损伤，导致线粒体功能障碍。级联的 ROS 可以传播并可能诱导线粒体电子传递链的血红素漂白和蛋白质半胱氨酸磺化 (PrSO ₃ H)。在此，我们研究了在体内缺血 30 分钟和再灌注 24 小时的大鼠心脏线粒体中 I/R 介导的复合物 III 的不可逆氧化损伤的机制。在 I/R 区域，复合物 III 的催化活性显着受损。光谱分析表明 I/R 介导了血红素b和c  +  c _1的破坏在线粒体中，支持 I/R 介导的复合物 III 损伤。然而，尽管状态 3 呼吸减少，但仅在缺血 30 分钟的大鼠心脏危险区域的线粒体中未观察到复合物 III 活性和血红素损伤的显着损害。在 I/R 线粒体中，仅检测到通过烷基化复合物 III 与 HCCS 下调的细胞色素c _{1的氨基甲酰甲基化 C 122} /C _{125 ，支持 I/R 介导的血红素}c ₁硫醚缺陷。LC-MS/MS 分析表明，I/R_线粒体在[ _2FeRieske 铁硫蛋白 ( uqcrfs1 ) 的 2S] 簇，从而削弱了电子传递活性。_{MS 分析还表明，在 C 65}处铰链蛋白的PrSO ₃ H和在 C ₁₄₀和 C ₂₂₀处的细胞色素c ₁增加，它们被限制在膜间空间中。MS 分析还表明，I/R 广泛增强了核心 1 ( uqcrc1 ) 和核心 2 ( uqcrc2 ) 的 PrSO _{3 H}) 基质隔室中的亚基，从而支持复合物 III 在再灌注过程中向内膜两侧释放 ROS 的结论。_{缺血性线粒体的分析表明，在假对照心脏的线粒体中检测到的复合物 III PrSO 3} H的基础水平适度降低，这表明再灌注期间的生理性高氧合和 ROS 过度产生介导了复合物 III PrSO ₃ H 的增强。 总之，再灌注介导的血红素损伤与增加的 PrSO ₃ H 控制复合物 III 的氧化损伤并加重缺血后心脏的线粒体功能障碍。