Humans have three different proliferating cell nuclear antigen (PCNA) clamp-loading complexes: RFC and CTF18-RFC load PCNA onto DNA, but ATAD5-RFC can only unload PCNA from DNA. The underlying structural basis of ATAD5-RFC unloading is unknown. We show here that ATAD5 has two unique locking loops that appear to tie the complex into a rigid structure, and together with a domain that plugs the DNA-binding chamber, prevent conformation changes required for DNA binding, likely explaining why ATAD5-RFC is exclusively a PCNA unloader. These features are conserved in the yeast PCNA unloader Elg1-RFC. We observe intermediates in which PCNA bound to ATAD5-RFC exists as a closed planar ring, a cracked spiral or a gapped spiral. Surprisingly, ATAD5-RFC can open a PCNA gap between PCNA protomers 2 and 3, different from the PCNA protomers 1 and 3 gap observed in all previously characterized clamp loaders.

人类有三种不同的增殖细胞核抗原 （PCNA） 夹装复合物：RFC 和 CTF18-RFC 将 PCNA 加载到 DNA 上，但 ATAD5-RFC 只能从 DNA 中卸载 PCNA。ATAD5-RFC 卸载的底层结构基础尚不清楚。我们在这里表明，ATAD5 有两个独特的锁定环，似乎将复合物连接到刚性结构中，并且与一个堵塞 DNA 结合室的结构域一起，防止了 DNA 结合所需的构象变化，这可能解释了为什么 ATAD5-RFC 完全是 PCNA 卸载器。这些特征在酵母 PCNA 卸载器 Elg1-RFC 中是保守的。我们观察到与 ATAD5-RFC 结合的 PCNA 以闭合平面环、破裂螺旋或间隙螺旋的形式存在的中间体。令人惊讶的是，ATAD5-RFC 可以在 PCNA 前聚体 2 和 3 之间打开一个 PCNA 间隙，这与在所有先前表征的夹式加载器中观察到的 PCNA 前聚体 1 和 3 间隙不同。