Max Perutz Labs, Medical University of Vienna, Vienna BioCenter (VBC), Dr. Bohr-Gasse 9/2, 1030 Vienna, Austria.
Institute of Biochemistry, Graz University of Technology, Petersgasse 12/2, 8010 Graz, Austria; Department of Medical Biochemistry and Biophysics, Karolinska Institutet, 171 77 Stockholm, Sweden.
Mol Cell. 2021 Jun 17;81(12):2520-2532.e16. doi: 10.1016/j.molcel.2021.04.007. Epub 2021 Apr 29.
The tRNA ligase complex (tRNA-LC) splices precursor tRNAs (pre-tRNA), and Xbp1-mRNA during the unfolded protein response (UPR). In aerobic conditions, a cysteine residue bound to two metal ions in its ancient, catalytic subunit RTCB could make the tRNA-LC susceptible to oxidative inactivation. Here, we confirm this hypothesis and reveal a co-evolutionary association between the tRNA-LC and PYROXD1, a conserved and essential oxidoreductase. We reveal that PYROXD1 preserves the activity of the mammalian tRNA-LC in pre-tRNA splicing and UPR. PYROXD1 binds the tRNA-LC in the presence of NAD(P)H and converts RTCB-bound NAD(P)H into NAD(P), a typical oxidative co-enzyme. However, NAD(P) here acts as an antioxidant and protects the tRNA-LC from oxidative inactivation, which is dependent on copper ions. Genetic variants of PYROXD1 that cause human myopathies only partially support tRNA-LC activity. Thus, we establish the tRNA-LC as an oxidation-sensitive metalloenzyme, safeguarded by the flavoprotein PYROXD1 through an unexpected redox mechanism.
tRNA 连接酶复合物（tRNA-LC）在未折叠蛋白反应（UPR）期间剪接前体 tRNA（pre-tRNA）和 Xbp1-mRNA。在有氧条件下，其古老的催化亚基 RTCB 中结合两个金属离子的半胱氨酸残基可能使 tRNA-LC 容易发生氧化失活。在这里，我们证实了这一假设，并揭示了 tRNA-LC 与 PYROXD1 之间的共同进化关联，PYROXD1 是一种保守且必需的氧化还原酶。我们揭示了 PYROXD1 在 pre-tRNA 剪接和 UPR 中保持哺乳动物 tRNA-LC 的活性。PYROXD1 在 NAD(P)H 的存在下结合 tRNA-LC，并将 RTCB 结合的 NAD(P)H 转化为 NAD(P)，这是一种典型的氧化辅酶。然而，这里的 NAD(P) 充当抗氧化剂，保护 tRNA-LC 免受氧化失活，这依赖于铜离子。导致人类肌病的 PYROXD1 遗传变异仅部分支持 tRNA-LC 活性。因此，我们将 tRNA-LC 确立为一种氧化敏感的金属酶，通过一种意想不到的氧化还原机制由黄素蛋白 PYROXD1 来保护。
