基于亲和网格的核糖体上 PKC 与 RACK1 结合的冷冻电镜研究。
Affinity grid-based cryo-EM of PKC binding to RACK1 on the ribosome.
机构信息
Department of Biochemistry and Molecular Biophysics, Columbia University, New York City, NY 10032, USA.
出版信息
J Struct Biol. 2013 Feb;181(2):190-4. doi: 10.1016/j.jsb.2012.11.006. Epub 2012 Dec 8.
Abstract
Affinity grids (AG) are specialized EM grids that bind macromolecular complexes containing tagged proteins to obtain maximum occupancy for structural analysis through single-particle EM. In this study, utilizing AG, we show that His-tagged activated PKC βII binds to the small ribosomal subunit (40S). We reconstructed a cryo-EM map which shows that PKC βII interacts with RACK1, a seven-bladed β-propeller protein present on the 40S and binds in two different regions close to blades 3 and 4 of RACK1. This study is a first step in understanding the molecular framework of PKC βII/RACK1 interaction and its role in translation.
摘要
亲和网格（AG）是专门的 EM 网格，可将含有标记蛋白的大分子复合物结合在一起，通过单颗粒 EM 获得最大的结构分析占有率。在这项研究中，我们利用 AG 表明，His 标记的激活型 PKCβII 与小核糖体亚基（40S）结合。我们重建了一个冷冻电镜图谱，显示 PKCβII 与 RACK1 相互作用，RACK1 是存在于 40S 上的七叶β-推进器蛋白，与 RACK1 的第 3 和第 4 叶附近的两个不同区域结合。这项研究是理解 PKCβII/RACK1 相互作用的分子框架及其在翻译中的作用的第一步。
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