Lee Jeonghwa, Wang Fei, Schnell Danny J
Department of Biochemistry and Molecular Biology and Program in Plant Biology, University of Massachusetts, Amherst, Massachusetts 01003, USA.
J Biol Chem. 2009 Nov 6;284(45):31130-41. doi: 10.1074/jbc.M109.053751. Epub 2009 Sep 10.
The post-translational import of nucleus-encoded preproteins into chloroplasts occurs through multimeric translocons in the outer (Toc) and inner (Tic) membranes. The high fidelity of the protein import process is maintained by specific recognition of the transit peptide of preproteins by the coordinate activities of two homologous GTPase Toc receptors, Toc34 and Toc159. Structural and biochemical studies suggest that dimerization of the Toc receptors functions as a component of the mechanism to control access of preproteins to the membrane translocation channel of the translocon. We show that specific mutations that disrupted receptor dimerization in vitro reduced the rate of protein import in transgenic Arabidopsis compared with the wild type receptor. The mutations did not affect the GTPase activities of the receptors. Interestingly, these mutations did not decrease the initial preprotein binding at the receptors, but they reduced the efficiency of the transition from preprotein binding to membrane translocation. These data indicate that dimerization of receptors has a direct role in protein import and support a hypothesis in which receptor-receptor interactions participate in the initiation of membrane translocation of chloroplast preproteins as part of the molecular mechanism of GTP-regulated protein import.
细胞核编码的前体蛋白向叶绿体的翻译后导入是通过外膜（Toc）和内膜（Tic）中的多聚体转运体进行的。蛋白导入过程的高保真度是由两种同源GTP酶Toc受体Toc34和Toc159的协同活性对前体蛋白转运肽的特异性识别来维持的。结构和生化研究表明，Toc受体的二聚化作为控制前体蛋白进入转运体膜转运通道机制的一个组成部分发挥作用。我们发现，与野生型受体相比，体外破坏受体二聚化的特定突变降低了转基因拟南芥中的蛋白导入速率。这些突变不影响受体的GTP酶活性。有趣的是，这些突变并没有降低前体蛋白在受体处的初始结合，但它们降低了从前体蛋白结合到膜转运转变的效率。这些数据表明，受体二聚化在蛋白导入中具有直接作用，并支持一种假说，即受体-受体相互作用作为GTP调节蛋白导入分子机制的一部分，参与叶绿体前体蛋白膜转运的起始。
