Department of Molecular Biology, University of Geneva, Geneva, Switzerland.
Department of Physiology, University of California, San Francisco, San Francisco, CA, USA.
Nature. 2021 Aug;596(7870):138-142. doi: 10.1038/s41586-021-03764-0. Epub 2021 Jul 21.
In early mitosis, the duplicated chromosomes are held together by the ring-shaped cohesin complex. Separation of chromosomes during anaphase is triggered by separase-a large cysteine endopeptidase that cleaves the cohesin subunit SCC1 (also known as RAD21). Separase is activated by degradation of its inhibitors, securin and cyclin B, but the molecular mechanisms of separase regulation are not clear. Here we used cryogenic electron microscopy to determine the structures of human separase in complex with either securin or CDK1-cyclin B1-CKS1. In both complexes, separase is inhibited by pseudosubstrate motifs that block substrate binding at the catalytic site and at nearby docking sites. As in Caenorhabditis elegans and yeast, human securin contains its own pseudosubstrate motifs. By contrast, CDK1-cyclin B1 inhibits separase by deploying pseudosubstrate motifs from intrinsically disordered loops in separase itself. One autoinhibitory loop is oriented by CDK1-cyclin B1 to block the catalytic sites of both separase and CDK1. Another autoinhibitory loop blocks substrate docking in a cleft adjacent to the separase catalytic site. A third separase loop contains a phosphoserine that promotes complex assembly by binding to a conserved phosphate-binding pocket in cyclin B1. Our study reveals the diverse array of mechanisms by which securin and CDK1-cyclin B1 bind and inhibit separase, providing the molecular basis for the robust control of chromosome segregation.
在有丝分裂早期，复制的染色体被环形的黏合复合物连接在一起。后期分裂时，染色体的分离是由分离酶（一种大型半胱氨酸内肽酶，可切割黏合亚基 SCC1（也称为 RAD21））触发的。分离酶的活性被其抑制剂 securin 和 cyclin B 的降解所激活，但分离酶调节的分子机制尚不清楚。在这里，我们使用低温电子显微镜来确定与人分离酶与 securin 或 CDK1-cyclin B1-CKS1 复合物的结构。在这两种复合物中，分离酶被假底物基序抑制，这些基序阻断了催化部位和附近对接部位的底物结合。与秀丽隐杆线虫和酵母一样，人 securin 含有其自身的假底物基序。相比之下，CDK1-cyclin B1 通过将自身无规则环中的假底物基序部署到分离酶中来抑制分离酶。一个自动抑制环由 CDK1-cyclin B1 定向，以阻断分离酶和 CDK1 的催化部位。另一个自动抑制环阻塞了与分离酶催化部位相邻的裂缝中的底物对接。第三个分离酶环包含一个磷酸丝氨酸，通过与 cyclin B1 中的保守磷酸结合口袋结合，促进复合物组装。我们的研究揭示了 securin 和 CDK1-cyclin B1 结合和抑制分离酶的多种机制，为染色体分离的强大控制提供了分子基础。
