The yeast SWR1 complex catalyses the exchange of histone H2A–H2B dimers in nucleosomes, with Htz1–H2B dimers^1,2,3. Here we used single-molecule analysis to demonstrate two-step double exchange of the two H2A–H2B dimers in a canonical yeast nucleosome with Htz1–H2B dimers, and showed that double exchange can be processive without release of the nucleosome from the SWR1 complex. Further analysis showed that bound nucleosomes flip between two states, with each presenting a different face, and hence histone dimer, to SWR1. The bound dwell time is longer when an H2A–H2B dimer is presented for exchange than when presented with an Htz1–H2B dimer. A hexasome intermediate in the reaction is bound to the SWR1 complex in a single orientation with the ‘empty’ site presented for dimer insertion. Cryo-electron microscopy analysis revealed different populations of complexes showing nucleosomes caught ‘flipping’ between different conformations without release, each placing a different dimer into position for exchange, with the Swc2 subunit having a key role in this process. Together, the data reveal a processive mechanism for double dimer exchange that explains how SWR1 can ‘proofread’ the dimer identities within nucleosomes.

酵母 SWR1 复合物催化核小体中组蛋白 H2A-H2B 二聚体与 Htz1-H2B 二聚体^1,2,3 的交换。在这里，我们使用单分子分析来证明经典酵母核小体中两个 H2A-H2B 二聚体与 Htz1-H2B 二聚体的两步双重交换，并表明双重交换可以在不从 SWR1 复合物中释放核小体的情况下进行。进一步分析表明，结合的核小体在两种状态之间翻转，每种状态都呈现不同的面孔，因此组蛋白二聚体与 SWR1 不同。当 H2A-H2B 二聚体呈递进行交换时，结合的停留时间比呈递 Htz1-H2B 二聚体时长。反应中的六六体中间体以单个方向与 SWR1 复合物结合，“空”位点用于二聚体插入。冷冻电子显微镜分析揭示了不同的复合物群，显示核小体在不同构象之间“翻转”而不释放，每个构象将不同的二聚体置于交换位置，其中 Swc2 亚基在此过程中起关键作用。总之，这些数据揭示了双二聚体交换的进行机制，解释了 SWR1 如何“校对”核小体内的二聚体身份。