How to arm a supervillin: designing F-actin binding activity into supervillin headpiece.
How to arm a supervillin: designing F-actin binding activity into supervillin headpiece.
复制标题
DOI：
10.1016/j.jmb.2009.08.018
复制
发表时间：
2009-10-30
影响因子：
5.6
通讯作者：
McKnight, C. James
中科院分区：
文献类型：
--
作者：
Brown, Jeffrey W.;Vardar-Ulu, Didem;McKnight, C. James
关键词：
Villin型头部结构域是紧凑的基序，已广泛用作蛋白质折叠的模型系统。虽然大多数头片段结构域结合肌动蛋白，但也有一些缺乏这种活性。在这里，我们提出了第一个核磁共振溶液的结构和15 N-弛豫分析的绒毛型头部结构域天然缺乏F-肌动蛋白结合活性，即超绒毛蛋白头部（SVHP）。发现该结构类似于结合F-肌动蛋白的其他头片段结构域。我们的NMR分析表明，supervillin头片段缺乏构象灵活的区域（V-环），存在于所有其他villin型头片段结构域，这是必不可少的磷酰基调节dematin头片段。在比较的静电表面电位图的SVHP，其他绒毛型头片段结构域与F-肌动蛋白的显着亲和力，我们确定了一个积极的表面电位保守的头片段结构域之间的结合F-肌动蛋白，但没有从SVHP。SVHP中的单点突变（L38 K）产生类似的正表面电位，赋予SVHP对F-肌动蛋白的特异性亲和力，其在最紧密结合头片段结构域的数量级内。我们认为，这种效应可能是由一个特定的埋盐桥之间的头盔和肌动蛋白。由于没有高分辨率的结构信息存在的绒毛型头片段：F-肌动蛋白复合物，我们的研究结果表明，通过正诱变，它是可能的设计到同源蛋白的结合活性，而没有结构信息的对应物的结合表面。
Villin-type headpiece domains are compact motifs that have been used extensively as model systems for protein folding. Although the majority of headpiece domains bind actin, there are some that lack this activity. Here, we present the first NMR solution structure and 15N-relaxation analysis of a villin-type headpiece domain natively devoid of F-actin binding activity, that of supervillin headpiece (SVHP). The structure was found to be similar to other headpiece domains that bind F-actin. Our NMR analysis demonstrates that supervillin headpiece lacks a conformationally flexible region (V-loop), present in all other villin-type headpiece domains, and which is essential to the phosphoryl regulation of dematin headpiece. In comparing the electrostatic surface potential map of SVHP, to that of other villin-type headpiece domains with significant affinity for F-actin, we identified a positive surface potential conserved among headpiece domains that bind F-actin, but absent from SVHP. A single point mutation (L38K) in SVHP, which creates a similar positive surface potential, endowed SVHP with specific affinity for F-actin that is within an order of magnitude of the tightest binding headpiece domains. We propose that this effect is likely conferred by a specific buried salt-bridge between headpiece and actin. As no high-resolution structural information exists for the villin-type headpiece: F-actin complex, our results demonstrate that through positive mutagenesis, it is possible to design binding activity into homologous proteins without structural information of the counterpart's binding surface.
登录
查看更多内容
影响因子：
8
作者：
Frank, BS;Vardar, D;McKnight, CJ
通讯作者：
McKnight, CJ
影响因子：
2.9
作者：
FORMANKAY, JD;GRONENBORN, AM;CLORE, GM
通讯作者：
CLORE, GM
DOI：
10.1107/s0907444998003254
发表时间：
1998-09-01
期刊：
ACTA CRYSTALLOGRAPHICA SECTION D-BIOLOGICAL CRYSTALLOGRAPHY
影响因子：
--
作者：
Brunger, AT;Adams, PD;Warren, GL
通讯作者：
Warren, GL
影响因子：
5.6
作者：
ARSENIEV, A;SCHULTZE, P;WUTHRICH, K
通讯作者：
WUTHRICH, K
影响因子：
2.7
作者：
DELAGLIO, F;GRZESIEK, S;BAX, A
通讯作者：
BAX, A