Oleinikov A V, Jokhadze G G, Traut R R
Department of Pediatrics, School of Medicine, University of California, Davis, CA 95616, USA.
Proc Natl Acad Sci U S A. 1998 Apr 14;95(8):4215-8. doi: 10.1073/pnas.95.8.4215.
During protein synthesis, the two elongation factors Tu and G alternately bind to the 50S ribosomal subunit at a site of which the protein L7/L12 is an essential component. L7/L12 is present in each 50S subunit in four copies organized as two dimers. Each dimer consists of distinct domains: a single N-terminal ("tail") domain that is responsible for both dimerization and binding to the ribosome via interaction with the protein L10 and two independent globular C-terminal domains ("heads") that are required for binding of elongation factors to ribosomes. The two heads are connected by flexible hinge sequences to the N-terminal domain. Important questions concerning the mechanism by which L7/L12 interacts with elongation factors are posed by us in response to the presence of two dimers, two heads per dimer, and their dynamic, mobile properties. In an attempt to answer these questions, we constructed a single-headed dimer of L7/L12 by using recombinant DNA techniques and chemical cross-linking. This chimeric molecule was added to inactive core particles lacking wild-type L7/L12 and shown to restore activity to a level approaching that of wild-type two-headed L7/L12.
在蛋白质合成过程中，两种延伸因子Tu和G交替结合到50S核糖体亚基上的一个位点，蛋白质L7/L12是该位点的重要组成部分。L7/L12以两个二聚体的形式存在于每个50S亚基中，共四个拷贝。每个二聚体由不同的结构域组成：一个单一的N端（“尾部”）结构域，负责二聚化并通过与蛋白质L10相互作用结合到核糖体上；两个独立的球状C端结构域（“头部”），延伸因子与核糖体结合需要它们。两个头部通过柔性铰链序列连接到N端结构域。鉴于存在两个二聚体、每个二聚体有两个头部以及它们的动态、可移动特性，我们提出了关于L7/L12与延伸因子相互作用机制的重要问题。为了回答这些问题，我们利用重组DNA技术和化学交联构建了L7/L12的单头二聚体。将这种嵌合分子添加到缺乏野生型L7/L12的无活性核心颗粒中，结果显示其活性恢复到接近野生型双头L7/L12的水平。
