Single-molecule detection of phosphorylation-induced plasticity changes during ezrin activation
Single-molecule detection of phosphorylation-induced plasticity changes during ezrin activation
复制标题
单分子检测埃兹蛋白激活过程中磷酸化诱导的可塑性变化
DOI：
10.1016/j.febslet.2007.06.071
复制
发表时间：
2007-07-24
期刊：
影响因子：
3.5
通讯作者：
Yao, Xuebiao
中科院分区：
文献类型：
--
作者：
Liu, Dan;Ge, Ling;Yao, Xuebiao
Ezrin-radixin-moesin蛋白家族在皮层肌动蛋白细胞骨架和质膜之间提供了受调节的连接。埃兹蛋白的磷酸化在功能上与膜动力学和可塑性有关。我们最近的研究表明，磷酸化的ezrin保守的T567残基改变胃壁细胞的生理。然而，磷酸化诱导ezrin激活的分子机制仍然难以捉摸。在这里，我们使用原子力显微镜（AFM）探测磷酸化介导的激活ezrin在单分子。产生磷酸模拟和不可磷酸化的突变埃兹蛋白并纯化至同质。通过AFM对两种ezrin突变体的比较分析证明了磷酸活化后N-和C-末端结构域的解折叠。为了测量机械展开过程中域间接触的物理力，我们使用涂覆在AFM针尖上的N-末端埃兹蛋白探测埃兹蛋白的限定区域。比较力测量表明，T567磷酸化诱导的ezrin的展开有利于分子间的关联。两者合计，这些结果提供了磷酸化引起的ERM蛋白的功能激活的分子例证，并表明刺激诱导的蛋白质构象变化可以用作协调细胞动力学的信号传导机制。(c)2007年由Elsevier B. V.代表欧洲生物化学学会联合会出版。
Ezrin-radixin-moesin protein family provides a regulated link between the cortical actin cytoskeleton and the plasma membrane. Phosphorylation of ezrin has been functionally linked to membrane dynamics and plasticity. Our recent study demonstrated that phosphorylation of the conserved T567 residue of ezrin alters the physiology of gastric parietal cells. However, the molecular mechanism of phosphorylation-induced ezrin activation has remained elusive. Here we use atomic force microscopy (AFM) to probe phosphorylation-mediated activation of ezrin in single molecules. The phospho-mimicking and non-phosphorylatable mutant ezrin proteins were generated and purified to homogeneity. Comparative analyses of two ezrin rnutants by AFM demonstrate the unfolding of the N- and C-terminal domains upon the phospho-activation. To measure the physical force underlying the inter-domain contact during mechanical unfolding, we probed the defined region of ezrin using the N-terminal ezrin coated onto the AFM tip. Comparative force measurements indicate that T567 phosphorylation-induced unfolding of ezrin favors the inter-molecular association. Taken together, these results provide molecular illustration of phosphorylation elicited functional activation of ERM proteins and indicate that stimulus-induced protein conformational change can be used as a signaling mechanism orchestrating cellular dynamics. (c) 2007 Published by Elsevier B.V. on behalf of the Federation of European Biochemical Societies.