Department of Chemistry, Middle Tennessee State University, 1301 East Main Street, Murfreesboro, TN 37132, USA.
Department of Chemistry, Middle Tennessee State University, 1301 East Main Street, Murfreesboro, TN 37132, USA.
J Mol Biol. 2019 Mar 15;431(6):1250-1266. doi: 10.1016/j.jmb.2019.01.039. Epub 2019 Feb 5.
Oxidative stress is a common challenge to mitochondrial function where reactive oxygen species are capable of significant organelle damage. The generation of mitochondrial reactive oxygen species occurs in the inner membrane and matrix compartments as a consequence of subunit function in the electron transport chain and citric acid cycle, respectively. Maintenance of mitochondrial proteostasis and stress response is facilitated by compartmentalized proteases that couple the energy of ATP hydrolysis to unfolding and the regulated removal of damaged, misfolded, or aggregated proteins. The mitochondrial protease YME1L functions in the maintenance of proteostasis in the intermembrane space. YME1L is an inner membrane-anchored hexameric protease with distinct N-terminal, transmembrane, AAA+ (ATPases associated with various cellular activities), and C-terminal M41 zinc-dependent protease domains. The effect of oxidative stress on enzymes such as YME1L tasked with maintaining proteostasis is currently unclear. We report here that recombinant YME1L undergoes a reversible conformational change in response to oxidative stress that involves the interaction of one hydrogen peroxide molecule per YME1L monomer with affinities equal to 31 ± 2 and 26 ± 1 mM for conditions lacking or including nucleotide, respectively. Our data also reveal that oxidative stress does not significantly impact nucleotide binding equilibria, but does stimulate a 2-fold increase in the rate constant for high-affinity ATP binding from (8.9 ± 0.2) × 10 M s to (1.5 ± 0.1) × 10 M s. Taken together, these data may suggest a mechanism for the regulated processing of YME1L by other inner membrane proteases such as OMA1.
氧化应激是线粒体功能的常见挑战，其中活性氧能够对细胞器造成严重损伤。线粒体活性氧的产生发生在内膜和基质隔室中，分别是电子传递链和柠檬酸循环亚基功能的结果。线粒体蛋白质稳态和应激反应的维持是通过将 ATP 水解的能量与展开以及受损、错误折叠或聚集蛋白的有调节去除偶联的分隔蛋白酶来实现的。线粒体蛋白酶 YME1L 在线粒体蛋白稳态的维持中发挥作用。YME1L 是一种具有独特的 N 端、跨膜、AAA+（与各种细胞活动相关的 ATP 酶）和 C 端 M41 锌依赖性蛋白酶结构域的膜内六聚体蛋白酶。目前尚不清楚氧化应激对 YME1L 等负责维持蛋白质稳态的酶的影响。我们在这里报告，重组 YME1L 在氧化应激下发生可逆构象变化，涉及每个 YME1L 单体与一个过氧化氢分子的相互作用，亲和力分别等于 31±2 和 26±1mM，分别为无核苷酸和有核苷酸条件下的亲和力。我们的数据还表明，氧化应激不会显著影响核苷酸结合平衡，但确实会刺激高亲和力 ATP 结合的速率常数增加 2 倍，从（8.9±0.2）×10M s增加到（1.5±0.1）×10M s。综上所述，这些数据可能表明了 YME1L 被其他内膜蛋白酶（如 OMA1）有调节地加工的机制。
