Division of Structural Biology, Institute of Cancer Research, Chester Beatty Laboratories, 237 Fulham Road, London SW3 6JB, UK.
J Mol Biol. 2013 Nov 15;425(22):4236-48. doi: 10.1016/j.jmb.2013.04.004. Epub 2013 Apr 11.
The anaphase-promoting complex or cyclosome (APC/C) is a large E3 RING-cullin ubiquitin ligase composed of between 14 and 15 individual proteins. A striking feature of the APC/C is that only four proteins are involved in directly recognizing target proteins and catalyzing the assembly of a polyubiquitin chain. All other subunits, which account for >80% of the mass of the APC/C, provide scaffolding functions. A major proportion of these scaffolding subunits are structurally related. In metazoans, there are four canonical tetratricopeptide repeat (TPR) proteins that form homo-dimers (Apc3/Cdc27, Apc6/Cdc16, Apc7 and Apc8/Cdc23). Here, we describe the crystal structure of the N-terminal homo-dimerization domain of Schizosaccharomyces pombe Cdc23 (Cdc23(Nterm)). Cdc23(Nterm) is composed of seven contiguous TPR motifs that self-associate through a related mechanism to those of Cdc16 and Cdc27. Using the Cdc23(Nterm) structure, we generated a model of full-length Cdc23. The resultant "V"-shaped molecule docks into the Cdc23-assigned density of the human APC/C structure determined using negative stain electron microscopy (EM). Based on sequence conservation, we propose that Apc7 forms a homo-dimeric structure equivalent to those of Cdc16, Cdc23 and Cdc27. The model is consistent with the Apc7-assigned density of the human APC/C EM structure. The four canonical homo-dimeric TPR proteins of human APC/C stack in parallel on one side of the complex. Remarkably, the uniform relative packing of neighboring TPR proteins generates a novel left-handed suprahelical TPR assembly. This finding has implications for understanding the assembly of other TPR-containing multimeric complexes.
后期促进复合物或细胞周期蛋白(APC/C)是一种由 14 到 15 个单独蛋白组成的大型 E3 RING-cullin 泛素连接酶。APC/C 的一个显著特点是只有四个蛋白直接参与识别靶蛋白并催化多泛素链的组装。所有其他亚基,占 APC/C 质量的>80%,提供支架功能。这些支架亚基中的大部分在结构上是相关的。在后生动物中,有四个典型的四肽重复(TPR)蛋白形成同源二聚体(Apc3/Cdc27、Apc6/Cdc16、Apc7 和 Apc8/Cdc23)。在这里,我们描述了酿酒酵母 Cdc23(Cdc23(Nterm))N 端同源二聚化结构域的晶体结构。Cdc23(Nterm)由七个连续的 TPR 基序组成,通过与 Cdc16 和 Cdc27 相似的机制自我组装。利用 Cdc23(Nterm)结构,我们生成了全长 Cdc23 的模型。所得的“V”形分子与使用负染电子显微镜(EM)确定的人 APC/C 结构中 Cdc23 分配的密度对接。基于序列保守性,我们提出 Apc7 形成与 Cdc16、Cdc23 和 Cdc27 相当的同源二聚体结构。该模型与人类 APC/C EM 结构中 Apc7 分配的密度一致。人 APC/C 的四个典型同源二聚体 TPR 蛋白在复合物的一侧平行堆叠。值得注意的是,相邻 TPR 蛋白的均匀相对包装产生了一种新的左手超螺旋 TPR 组装。这一发现对理解其他含有 TPR 的多聚体复合物的组装具有重要意义。