Boland Andreas, Martin Thomas G, Zhang Ziguo, Yang Jing, Bai Xiao-Chen, Chang Leifu, Scheres Sjors H W, Barford David
MRC Laboratory of Molecular Biology, Cambridge, UK.
Nat Struct Mol Biol. 2017 Apr;24(4):414-418. doi: 10.1038/nsmb.3386. Epub 2017 Mar 6.
Separase is a caspase-family protease that initiates chromatid segregation by cleaving the kleisin subunits (Scc1 and Rec8) of cohesin, and regulates centrosome duplication and mitotic spindle function through cleavage of kendrin and Slk19. To understand the mechanisms of securin regulation of separase, we used single-particle cryo-electron microscopy (cryo-EM) to determine a near-atomic-resolution structure of the Caenorhabditis elegans separase-securin complex. Separase adopts a triangular-shaped bilobal architecture comprising an N-terminal tetratricopeptide repeat (TPR)-like α-solenoid domain docked onto the conserved C-terminal protease domain. Securin engages separase in an extended antiparallel conformation, interacting with both lobes. It inhibits separase by interacting with the catalytic site through a pseudosubstrate mechanism, thus revealing that in the inhibited separase-securin complex, the catalytic site adopts a conformation compatible with substrate binding. Securin is protected from cleavage because an aliphatic side chain at the P1 position represses protease activity by disrupting the organization of catalytic site residues.
分离酶是一种半胱天冬酶家族蛋白酶，它通过切割黏连蛋白的kleisin亚基（Scc1和Rec8）启动染色单体分离，并通过切割kendrin和Slk19来调节中心体复制和有丝分裂纺锤体功能。为了理解securin对分离酶的调控机制，我们使用单颗粒冷冻电子显微镜（cryo-EM）确定了秀丽隐杆线虫分离酶-securin复合物的近原子分辨率结构。分离酶采用三角形双叶结构，由一个对接在保守的C端蛋白酶结构域上的N端四肽重复（TPR）样α-螺旋结构域组成。Securin以延伸的反平行构象与分离酶结合，与两个叶都相互作用。它通过假底物机制与催化位点相互作用来抑制分离酶，从而揭示在受抑制的分离酶-securin复合物中，催化位点采用与底物结合兼容的构象。Securin受到保护不被切割，因为P1位置的脂肪族侧链通过破坏催化位点残基的组织来抑制蛋白酶活性。
