Mant A, Nielsen V S, Knott T G, Møller B L, Robinson C
Department of Biological Sciences, University of Warwick, Coventry, United Kingdom.
J Biol Chem. 1994 Nov 4;269(44):27303-9.
The photosystem I (PSI) complex in higher plants contains eight nuclear-encoded subunits, of which two (PSI-F and -N) are synthesized with bipartite presequences containing cleavable thylakoid transfer sequences. Previous studies on four other chloroplast proteins bearing bipartite presequences have shown that they are transported across the thylakoid membrane by two distinct mechanisms. One mechanism is delta pH-dependent and hence sensitive to uncouplers, whereas the other is inhibited by azide. We show that PSI-F is targeted by the latter pathway, since its translocation across the thylakoid membrane is inhibited by azide but not by nigericin. Translocation is furthermore unaffected by the presence of high concentrations of the lumenal 23-kDa photosystem II (PSII) protein, which is known to be transported by the delta pH-dependent pathway. In contrast, translocation of PSI-N across the thylakoid membrane is completely blocked by saturating concentrations of pre-23-kDa protein. Three proteins are now known to be synthesized with thylakoid transfer signals in both higher plants and cyanobacteria (PSI-F, plastocyanin, and the 33-kDa PSII protein), and all three are transported by the azide-sensitive, possibly sec-dependent pathway. In contrast, PSI-N and the 23-kDa and 16-kDa PSII proteins (transported by the delta pH-driven pathway in higher plants) are all absent in cyanobacteria. These data suggest that the delta pH-dependent translocation mechanism for these proteins may also have arisen relatively recently during the evolution of the chloroplast. Three additional PSI proteins (PSI-H, -K, and -L) are synthesized in the cytosol with stroma-targeting presequences and hence integrate into the thylakoid membrane by means of information in the mature proteins. We show that the integration mechanisms are insensitive to azide in each case, and nigericin causes only a slight inhibition of integration in each case. We therefore suggest that these proteins are targeted into the thylakoid membrane by a separate pathway(s).
高等植物中的光系统I（PSI）复合体包含八个核编码亚基，其中两个亚基（PSI-F和-N）在合成时带有可裂解类囊体转运序列的双功能前导序列。先前对其他四种带有双功能前导序列的叶绿体蛋白的研究表明，它们通过两种不同的机制穿过类囊体膜。一种机制是依赖ΔpH，因此对解偶联剂敏感，而另一种机制则被叠氮化物抑制。我们发现PSI-F是通过后一种途径靶向的，因为它穿过类囊体膜的转运受到叠氮化物的抑制，而不受尼日利亚菌素的抑制。此外，转运不受高浓度的腔内23 kDa光系统II（PSII）蛋白的影响，已知该蛋白是通过依赖ΔpH的途径转运的。相比之下，PSI-N穿过类囊体膜的转运被饱和浓度的前23 kDa蛋白完全阻断。现在已知在高等植物和蓝细菌中，有三种蛋白在合成时带有类囊体转运信号（PSI-F、质体蓝素和33 kDa的PSII蛋白），并且这三种蛋白都是通过对叠氮化物敏感、可能依赖Sec的途径转运的。相比之下，PSI-N以及23 kDa和16 kDa的PSII蛋白（在高等植物中通过ΔpH驱动的途径转运）在蓝细菌中均不存在。这些数据表明，这些蛋白依赖ΔpH的转运机制可能也是在叶绿体进化过程中相对较晚出现的。另外三种PSI蛋白（PSI-H、-K和-L）在细胞质中合成时带有靶向基质的前导序列，因此通过成熟蛋白中的信息整合到类囊体膜中。我们发现，在每种情况下，整合机制对叠氮化物均不敏感，并且尼日利亚菌素在每种情况下仅对整合产生轻微抑制。因此，我们认为这些蛋白是通过一条独立的途径靶向到类囊体膜中的。
