Agrawal R K, Linde J, Sengupta J, Nierhaus K H, Frank J
Wadsworth Center, Empire State Plaza, Albany, NY 12201-0509, USA.
J Mol Biol. 2001 Aug 24;311(4):777-87. doi: 10.1006/jmbi.2001.4907.
L11 protein is located at the base of the L7/L12 stalk of the 50 S subunit of the Escherichia coli ribosome. Because of the flexible nature of the region, recent X-ray crystallographic studies of the 50 S subunit failed to locate the N-terminal domain of the protein. We have determined the position of the complete L11 protein by comparing a three-dimensional cryo-EM reconstruction of the 70 S ribosome, isolated from a mutant lacking ribosomal protein L11, with the three-dimensional map of the wild-type ribosome. Fitting of the X-ray coordinates of L11-23 S RNA complex and EF-G into the cryo-EM maps combined with molecular modeling, reveals that, following EF-G-dependent GTP hydrolysis, domain V of EF-G intrudes into the cleft between the 23 S ribosomal RNA and the N-terminal domain of L11 (where the antibiotic thiostrepton binds), causing the N-terminal domain to move and thereby inducing the formation of the arc-like connection with the G' domain of EF-G. The results provide a new insight into the mechanism of EF-G-dependent translocation.
L11蛋白位于大肠杆菌核糖体50 S亚基的L7/L12茎的基部。由于该区域的柔性性质，最近对50 S亚基的X射线晶体学研究未能定位该蛋白的N端结构域。我们通过比较从缺乏核糖体蛋白L11的突变体中分离出的70 S核糖体的三维冷冻电镜重建与野生型核糖体的三维图谱，确定了完整L11蛋白的位置。将L11-23 S RNA复合物和EF-G的X射线坐标拟合到冷冻电镜图谱中并结合分子建模，结果表明，在EF-G依赖的GTP水解后，EF-G的结构域V侵入23 S核糖体RNA与L11的N端结构域之间的裂隙（抗生素硫链丝菌肽结合的部位），导致N端结构域移动，从而诱导与EF-G的G'结构域形成弧形连接。这些结果为EF-G依赖的转位机制提供了新的见解。
