The human EMAP-like protein-70 (ELP70) is a microtubule destabilizer that localizes to the mitotic apparatus
The human EMAP-like protein-70 (ELP70) is a microtubule destabilizer that localizes to the mitotic apparatus
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DOI：
10.1074/jbc.m106628200
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发表时间：
2002-01-11
影响因子：
4.8
通讯作者：
Suprenant, KA
中科院分区：
文献类型：
--
作者：
Eichenmüller, B;Everley, P;Suprenant, KA
在这篇报道中，我们发现棘皮动物的杆状微管（MT）下的杆状相关蛋白（EMAP）下的棘皮动物（m）和杆状蛋白（ELP）下的杆状相关蛋白（E）下的杆状相关蛋白（MAP-）下的棘皮动物（l）和杆状蛋白（ELP）下的棘皮动物（p）有一个相似的结构域，即杆状疏水蛋白（ELP）下的高度保守的杆状蛋白（HELP）结构域和一个大的天冬氨酸（WD）重复结构域。为了确定哺乳动物ELP的功能，我们产生了针对70 kDa的人ELP的抗体，并显示ELP 70与HeLa细胞提取物中的Mrs共组装，并与有丝分裂器中的MT共定位。为了确定ELP 70是否直接与MT结合，从杆状病毒感染的Sf 9细胞表达人ELP 70并纯化至均一。纯化的ELP 70与纯化的MT结合，化学计量为0.40 +/-0.04 mol ELP 70/mol微管蛋白二聚体，固有解离常数为0.44 +/-0.13 μ m。使用成核组装测定和视频增强差分干涉对比显微镜，我们证明，ELP 70减少种子成核，降低生长速率，并促进MT灾难的浓度依赖性的方式。结果，含有ELP 70的MT显著短于仅由微管蛋白组装的MT。这些数据表明，ELP 70是一种新的MT去稳定剂。提出了一个侧向失稳模型来描述ELP 70对微管的影响。
In this report, we show that the echinoderm (m) under bar icrotubule (MT)(a) under bar ssociated protein (EMAP) and related (E) under bar MAP-(l) under bar ike (p) under bar roteins (ELPs) share a similar domain organization with a highly conserved (h) under bar ydrophobic (ELP) under bar (HELP) domain and a large tryptophan-aspartic acid (WD) repeat domain. To determine the function of mammalian ELPs, we generated antibodies against a 70-kDa human ELP and showed that ELP70 coassembled with Mrs in HeLa cell extracts and colocalized with MTs in the mitotic apparatus. To determine whether ELP70 bound to MTs directly, human ELP70 was expressed and purified to homogeneity from baculovirus-infected Sf9 cells. Purified ELP70 bound to purified MTs with a stoichiometry of 0.40 +/- 0.04 mol of ELP70/mol of tubulin dimer and with an intrinsic dissociation constant of 0.44 +/- 0.13 mum. Using a nucleated assembly assay and video-enhanced differential interference contrast microscopy, we demonstrated that ELP70 reduced seeded nucleation, reduced the growth rate, and promoted MT catastrophes in a concentration-dependent manner. As a result, ELP70-containing MTs were significantly shorter than MTs assembled from tubulin alone. These data indicate that ELP70 is a novel MT destabilizer. A lateral destabilization model is presented to describe ELP70's effects on microtubules.