Cline K, Henry R, Li C, Yuan J
Horticultural Sciences Department, University of Florida, Gainesville 32611.
EMBO J. 1993 Nov;12(11):4105-14. doi: 10.1002/j.1460-2075.1993.tb06094.x.
Many thylakoid proteins are cytosolically synthesized and have to cross the two chloroplast envelope membranes as well as the thylakoid membrane en route to their functional locations. In order to investigate the localization pathways of these proteins, we over-expressed precursor proteins in Escherichia coli and used them in competition studies. Competition was conducted for import into the chloroplast and for transport into or across isolated thylakoids. We also developed a novel in organello method whereby competition for thylakoid transport occurred within intact chloroplasts. Import of all precursors into chloroplasts was similarly inhibited by saturating concentrations of the precursor to the OE23 protein. In contrast, competition for thylakoid transport revealed three distinct precursor specificity groups. Lumen-resident proteins OE23 and OE17 constitute one group, lumenal proteins plastocyanin and OE33 a second, and the membrane protein LHCP a third. The specificity determined by competition correlates with previously determined protein-specific energy requirements for thylakoid transport. Taken together, these results suggest that thylakoid precursor proteins are imported into chloroplasts on a common import apparatus, whereupon they enter one of several precursor-specific thylakoid transport pathways.
许多类囊体蛋白是在细胞质中合成的，在到达其功能位点的途中必须穿过叶绿体的两层包膜膜以及类囊体膜。为了研究这些蛋白的定位途径，我们在大肠杆菌中过表达前体蛋白，并将其用于竞争研究。竞争实验分别针对叶绿体导入以及向分离的类囊体中转运或穿过类囊体进行。我们还开发了一种新的体内方法，使类囊体转运的竞争在完整的叶绿体中进行。OE23蛋白前体的饱和浓度同样抑制了所有前体向叶绿体的导入。相比之下，类囊体转运竞争揭示了三个不同的前体特异性组。腔驻留蛋白OE23和OE17构成一组，腔蛋白质体蓝素和OE33构成第二组，膜蛋白LHCP构成第三组。通过竞争确定的特异性与先前确定的类囊体转运的蛋白质特异性能量需求相关。综上所述，这些结果表明类囊体前体蛋白通过共同的导入装置导入叶绿体，随后进入几种前体特异性类囊体转运途径之一。
