Site-specific labeling of proteins for electron microscopy.
Site-specific labeling of proteins for electron microscopy.
复制标题
DOI：
10.1016/j.jsb.2015.09.010
复制
发表时间：
2015-11
影响因子：
3
通讯作者：
Lander GC
中科院分区：
文献类型：
--
作者：
Dambacher CM;Lander GC
电子显微镜通常用于确定大分子组装体的亚基组织。然而，该领域缺乏可靠的分子标记方法来明确识别组成亚基。我们提出了一种策略，利用非天然氨基酸的独特性质，以便能够在大分子表面上任何暴露于溶剂的位置上对单个、易于识别的蛋白质标签进行位点特异性附着。使用这种方法，我们清楚地标记了 19S 蛋白酶体盖子复合物内的亚基，该亚基不适合用传统方法进行标记。
Electron microscopy is commonly employed to determine the subunit organization of large macromolecular assemblies. However, the field lacks a robust molecular labeling methodology for unambiguous identification of constituent subunits. We present a strategy that exploits the unique properties of an unnatural amino acid in order to enable site-specific attachment of a single, readily identifiable protein label at any solvent-exposed position on the macromolecular surface. Using this method, we show clear labeling of a subunit within the 19S proteasome lid subcomplex that has not been amenable to labeling by traditional approaches.