The C-terminal tail of UNC-60B (actin depolymerizing factor/cofilin) is critical for maintaining its stable association with F-actin and is implicated in the second actin-binding site
The C-terminal tail of UNC-60B (actin depolymerizing factor/cofilin) is critical for maintaining its stable association with F-actin and is implicated in the second actin-binding site
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DOI：
10.1074/jbc.m007563200
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发表时间：
2001-02-23
影响因子：
4.8
通讯作者：
Weeds, AG
中科院分区：
文献类型：
--
作者：
Ono, S;McGough, A;Weeds, AG
肌动蛋白解聚因子（ADF）/cofilin通过结合两个纵向相关的肌动蛋白亚基来改变肌动蛋白丝的扭曲。在缺乏ADF/cofilin-F-肌动蛋白复合物的原子模型的情况下，我们已经确定了ADF/cofilin中对于丝结合至关重要的残基。在这里，我们对UNC-60 B的C末端尾部进行了表征用羧肽酶A去除C端异亮氨酸（Ile（152））或诱变截短Ile（152），可消除F-肌动蛋白结合活性，但大大增强肌动蛋白解聚活性，用Ala取代Ile（152）也有类似效果，但不太明显; F-肌动蛋白结合减弱，解聚活性略有增强。Arg（151）和Ile（152）的截断或Arg（151）被Ala取代也消除了F-肌动蛋白结合并增强了解聚活性。这些突变体中F-肌动蛋白结合的丧失伴随着切断活性的丧失或大大降低。所有的突变体都与G-肌动蛋白相互作用，与野生型没有区别。冷冻电子显微镜显示，ESTA-60 B改变了F-肌动蛋白的扭曲到类似的程度，脊椎动物ADF/cofilins。螺旋重建和结构建模的α-60 B-F-actin复合物揭示了α-60 B的C端可能参与两个肌动蛋白结合位点之一。
Actin depolymerizing factor (ADF)/cofilin changes the twist of actin filaments by binding two longitudinally associated actin subunits, In the absence of an atomic model of the ADF/cofilin-F-actin complex, we have identified residues in ADF/cofilin that are essential for filament binding. Here, we have characterized the C-terminal tail of UNC-60B (a nematode ADF/cofilin isoform) as a novel determinant for its association with F-actin, Removal of the C-terminal isoleucine (Ile(152)) by carboxypeptidase A or truncation by mutagenesis eliminated F-actin binding activity but strongly enhanced actin depolymerizing activity, Replacement of Ile(152) by Ala had a similar but less marked effect; F-actin binding was weakened and depolymerizing activity slightly enhanced. Truncation of both Arg(151) and Ile(152) or replacement of Arg(151) with Ala also abolished F-actin binding and enhanced depolymerizing activity. Loss of F-actin binding in these mutants was accompanied by loss or greatly decreased severing activity. All of the variants of UNC-60B interacted with G-actin in an indistinguishable manner from wild type. Cryoelectron microscopy showed that UNC-60B changed the twist of F-actin to a similar extent to vertebrate ADF/cofilins. Helical reconstruction and structural modeling of UNC-60B-F-actin complex reveal how the C terminus of UNC-60B might be involved in one of the two actin-binding sites.