Smc5/6 is a member of the eukaryotic structural maintenance of chromosomes (SMC) family of complexes with important roles in genome maintenance and viral restriction. However, limited structural understanding of Smc5/6 hinders the elucidation of its diverse functions. Here, we report cryo-EM structures of the budding yeast Smc5/6 complex in eight-subunit, six-subunit and five-subunit states. Structural maps throughout the entire length of these complexes reveal modularity and key elements in complex assembly. We show that the non-SMC element (Nse)2 subunit supports the overall shape of the complex and uses a wedge motif to aid the stability and function of the complex. The Nse6 subunit features a flexible hook region for attachment to the Smc5 and Smc6 arm regions, contributing to the DNA repair roles of the complex. Our results also suggest a structural basis for the opposite effects of the Nse1–3–4 and Nse5–6 subcomplexes in regulating Smc5/6 ATPase activity. Collectively, our integrated structural and functional data provide a framework for understanding Smc5/6 assembly and function.

Smc5/6 是真核生物染色体结构维护 （SMC） 复合物家族的成员，在基因组维护和病毒限制中起重要作用。然而，对 Smc5/6 的结构理解有限，阻碍了对其不同功能的阐明。在这里，我们报道了出芽酵母 Smc5/6 复合物在八个亚基、六个亚基和五个亚基状态下的冷冻电镜结构。这些综合体整个长度的结构图揭示了复杂装配体的模块化和关键要素。我们表明非 SMC 元件 （Nse）2 亚基支持复合物的整体形状，并使用楔形基序来帮助复合物的稳定性和功能。Nse6 亚基具有一个灵活的钩状区域，用于附着在 Smc5 和 Smc6 臂区域，有助于复合物的 DNA 修复作用。我们的结果还表明 Nse1-3-4 和 Nse5-6 亚复合物在调节 Smc5/6 ATP 酶活性方面具有相反作用的结构基础。总的来说，我们的综合结构和功能数据为理解 Smc5/6 组装和功能提供了一个框架。