Moore Misty, Goforth Robyn L, Mori Hiroki, Henry Ralph
Department of Biological Sciences, University of Arkansas, Fayetteville, AR 72701, USA.
J Cell Biol. 2003 Sep 29;162(7):1245-54. doi: 10.1083/jcb.200307067.
Integration of thylakoid proteins by the chloroplast signal recognition particle (cpSRP) posttranslational transport pathway requires the cpSRP, an SRP receptor homologue (cpFtsY), and the membrane protein ALB3. Similarly, Escherichia coli uses an SRP and FtsY to cotranslationally target membrane proteins to the SecYEG translocase, which contains an ALB3 homologue, YidC. In neither system are the interactions between soluble and membrane components well understood. We show that complexes containing cpSRP, cpFtsY, and ALB3 can be precipitated using affinity tags on cpSRP or cpFtsY. Stabilization of this complex with GMP-PNP specifically blocks subsequent integration of substrate (light harvesting chl a/b-binding protein [LHCP]), indicating that the complex occupies functional ALB3 translocation sites. Surprisingly, neither substrate nor cpSRP43, a component of cpSRP, was necessary to form a complex with ALB3. Complexes also contained cpSecY, but its removal did not inhibit ALB3 function. Furthermore, antibody bound to ALB3 prevented ALB3 association with cpSRP and cpFtsY and inhibited LHCP integration suggesting that a complex containing cpSRP, cpFtsY, and ALB3 must form for proper LHCP integration.
叶绿体信号识别颗粒（cpSRP）翻译后转运途径对类囊体蛋白的整合需要cpSRP、一种SRP受体同源物（cpFtsY）以及膜蛋白ALB3。同样，大肠杆菌利用一种SRP和FtsY将膜蛋白共翻译靶向至SecYEG转位酶，该转位酶含有一种ALB3同源物YidC。在这两个系统中，可溶性成分与膜成分之间的相互作用都尚未得到充分理解。我们发现，含有cpSRP、cpFtsY和ALB3的复合物可以利用cpSRP或cpFtsY上的亲和标签进行沉淀。用GMP-PNP稳定该复合物会特异性地阻断底物（捕光叶绿素a/b结合蛋白[LHCP]）随后的整合，这表明该复合物占据了功能性的ALB3转位位点。令人惊讶的是，底物和cpSRP的一个组分cpSRP43对于与ALB3形成复合物都不是必需的。复合物中还含有cpSecY，但其去除并不抑制ALB3的功能。此外，与ALB3结合的抗体阻止了ALB3与cpSRP和cpFtsY的结合，并抑制了LHCP的整合，这表明含有cpSRP、cpFtsY和ALB3的复合物必须形成才能实现LHCP的正常整合。
