School of Life Science and Technology, ShanghaiTech University, Shanghai, China.
Elife. 2022 Mar 14;11:e76107. doi: 10.7554/eLife.76107.
The bifunctional enzyme Δ-pyrroline-5-carboxylate synthase (P5CS) is vital to the synthesis of proline and ornithine, playing an essential role in human health and agriculture. Pathogenic mutations in the P5CS gene (ALDH18A1) lead to neurocutaneous syndrome and skin relaxation connective tissue disease in humans, and P5CS deficiency seriously damages the ability to resist adversity in plants. We have recently found that P5CS forms cytoophidia in vivo and filaments in vitro. However, it is difficult to appreciate the function of P5CS filamentation without precise structures. Using cryo-electron microscopy, here we solve the structures of full-length P5CS in three states at resolution from 3.1 to 4.3 Å. We observe distinct ligand-binding states and conformational changes for the GK and GPR domains, respectively. Divergent helical filaments are assembled by P5CS tetramers and stabilized by multiple interfaces. Point mutations disturbing those interfaces prevent P5CS filamentation and greatly reduce the enzymatic activity. Our findings reveal that filamentation is crucial for the coordination between the GK and GPR domains, providing a structural basis for the catalytic function of P5CS filaments.
双功能酶 Δ-吡咯啉-5-羧酸合成酶 (P5CS) 对脯氨酸和鸟氨酸的合成至关重要，在人类健康和农业中发挥着重要作用。P5CS 基因 (ALDH18A1) 的致病突变导致人类神经皮肤综合征和皮肤松弛结缔组织病，而 P5CS 缺乏严重损害植物抵抗逆境的能力。我们最近发现 P5CS 在体内形成细胞纤维，在体外形成纤维丝。然而，如果没有精确的结构，就很难理解 P5CS 纤维丝形成的功能。使用冷冻电子显微镜，我们在这里以 3.1 到 4.3 Å 的分辨率解决了全长 P5CS 在三种状态下的结构。我们观察到 GK 和 GPR 结构域分别具有不同的配体结合状态和构象变化。P5CS 四聚体组装出不同的螺旋纤维丝，并通过多个界面稳定。破坏这些界面的点突变阻止了 P5CS 纤维丝的形成，并大大降低了酶的活性。我们的发现表明纤维丝的形成对于 GK 和 GPR 结构域之间的协调至关重要，为 P5CS 纤维丝的催化功能提供了结构基础。
