Structure of spastin bound to a glutamate-rich peptide implies a hand-over-hand mechanism of substrate translocation
Structure of spastin bound to a glutamate-rich peptide implies a hand-over-hand mechanism of substrate translocation
复制标题
DOI：
10.1074/jbc.ac119.009890
复制
发表时间：
2020-01-10
影响因子：
4.8
通讯作者：
Hill, Christopher P.
中科院分区：
文献类型：
--
作者：
Han, Han;Schubert, Heidi L.;Hill, Christopher P.
AAA+ ATP酶家族的许多成员作为展开其蛋白质底物的六聚体起作用。这些AAA解折叠酶包括痉挛蛋白，其通过驱动微管的重塑和切断在真核细胞的结构中起关键作用，微管是微管蛋白亚基的细胞骨架聚合物。在这里，我们证明了一个人痉挛弱结合到未修饰的肽从C-末端段的人微管蛋白？1 A/B。包含交替的谷氨酸和酪氨酸残基的肽结合更紧密，这与已知的谷氨酸化对痉挛蛋白微管切断活性的重要性一致。阿cryo-EM结构的spastin肽复合物在4.2？解析揭示了不对称的六聚体，其中五个痉挛蛋白亚基采用螺旋、螺旋楼梯构型，其在中心孔内结合肽，而六聚体的第六个亚基从肽/底物移位，就好像从螺旋的一端过渡到另一端。这种结构不同于最近发表的果蝇spastin的结构，它形成了一个六亚基螺旋，没有一个过渡亚基。我们的结构类似于其他最近报道的AAA解折叠酶，包括减数分裂进化枝相对Vps 4，并支持一个模型，其中痉挛利用一个手在手的微管蛋白易位和微管重塑机制。
Many members of the AAA+ ATPase family function as hexamers that unfold their protein substrates. These AAA unfoldases include spastin, which plays a critical role in the architecture of eukaryotic cells by driving the remodeling and severing of microtubules, which are cytoskeletal polymers of tubulin subunits. Here, we demonstrate that a human spastin binds weakly to unmodified peptides from the C-terminal segment of human tubulin ?1A/B. A peptide comprising alternating glutamate and tyrosine residues binds more tightly, which is consistent with the known importance of glutamylation for spastin microtubule severing activity. A cryo-EM structure of the spastin-peptide complex at 4.2 ? resolution revealed an asymmetric hexamer in which five spastin subunits adopt a helical, spiral staircase configuration that binds the peptide within the central pore, whereas the sixth subunit of the hexamer is displaced from the peptide/substrate, as if transitioning from one end of the helix to the other. This configuration differs from a recently published structure of spastin from Drosophila melanogaster, which forms a six-subunit spiral without a transitioning subunit. Our structure resembles other recently reported AAA unfoldases, including the meiotic clade relative Vps4, and supports a model in which spastin utilizes a hand-over-hand mechanism of tubulin translocation and microtubule remodeling.