Correlation of CRM1-NES affinity with nuclear export activity.
Correlation of CRM1-NES affinity with nuclear export activity.
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DOI：
10.1091/mbc.e18-02-0096
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发表时间：
2018-08-15
影响因子：
3.3
通讯作者：
Chook YM
中科院分区：
文献类型：
--
作者：
Fu SC;Fung HYJ;Cağatay T;Baumhardt J;Chook YM
CRM1 （Exportin1/XPO1）通过结合经典的核输出信号（NESs）将数百种功能广泛的蛋白质输出到细胞核外。8- 15个氨基酸长的NESs含有4 - 5个疏水残基，在序列和crm1结合结构上高度多样化。在这里，我们研究了细胞中24种不同的NES肽的核输出活性与其CRM1-NES亲和力之间的关系。我们发现，结合亲和力和核输出活性与NESs的解离常数（Kds）在几十纳摩尔到几十微摩尔之间呈线性相关。Kds超出这一范围的国家大大减少了核出口活动。其中包括两种异常紧密结合的肽，一种来自小鼠微小病毒的非结构蛋白2 (MVM NS2)，另一种来自蛋白激酶a抑制剂（PKI） NES的突变体。结合CRM1的MVM NS2NES的晶体结构表明，异常紧密的CRM1结合源于NES内的分子内接触，这可能稳定了游离肽中CRM1结合的构象。这种机制的理解导致了两种新型肽抑制剂的设计，它们以皮摩尔亲和力结合CRM1。
CRM1 (Exportin1/XPO1) exports hundreds of broadly functioning protein cargoes out of the cell nucleus by binding to their classical nuclear export signals (NESs). The 8- to 15-amino-acid-long NESs contain four to five hydrophobic residues and are highly diverse in both sequence and CRM1-bound structure. Here we examine the relationship between nuclear export activities of 24 different NES peptides in cells and their CRM1-NES affinities. We found that binding affinity and nuclear export activity are linearly correlated for NESs with dissociation constants (Kds) between tens of nanomolar to tens of micromolar. NESs with Kds outside this range have significantly reduced nuclear export activities. These include two unusually tight-binding peptides, one from the nonstructural protein 2 of murine minute virus (MVM NS2) and the other a mutant of the protein kinase A inhibitor (PKI) NES. The crystal structure of CRM1-bound MVM NS2NES suggests that extraordinarily tight CRM1 binding arises from intramolecular contacts within the NES that likely stabilizes the CRM1-bound conformation in free peptides. This mechanistic understanding led to the design of two novel peptide inhibitors that bind CRM1 with picomolar affinity.