In Vitro Reconstitution of Insertion and Processing of Cytochrome f in a Homologous Chloroplast Translation System*
In Vitro Reconstitution of Insertion and Processing of Cytochrome f in a Homologous Chloroplast Translation System*
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同源叶绿体翻译系统中细胞色素 f 的插入和加工的体外重建*
DOI：
10.1074/jbc.m103005200
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发表时间：
2001
期刊：
影响因子：
--
通讯作者：
K. V. van Wijk
中科院分区：
文献类型：
--
作者：
T. Röhl;K. V. van Wijk
使用同源叶绿体翻译系统，我们重建了叶绿体编码的类囊体蛋白细胞色素f(pCytf)的插入和加工。交联证明 pCyt 新生链在连接到 70 S 核糖体时与 cpSecA 紧密相互作用，但这严格依赖于类囊体膜和功能性信号肽。这表明 cpSecA 仅在与膜结合时才在 pCytf 生物发生中发挥作用，很可能作为 Sec 易位子的一部分。没有检测到叶绿体信号识别颗粒 (cpSRP) 的 54 kDa 亚基与 pCytf 新生链之间相互作用的证据，这表明与多胞体 D1 蛋白相比，pCytf 不需要 cpSRP 进行靶向。 pCytf 的插入仅发生在共翻译过程中，导致加工后的信号肽和成熟蛋白在类囊体中进行加工和积累。这种共翻译膜插入和加工需要功能性信号肽并被叠氮化物抑制，这表明 cpSecA 对于可溶性管腔结构域的易位至关重要。 pCytf 也在翻译后与类囊体相关，但可溶性 N 末端结构域不能易位到管腔中。这是第一项利用叶绿体翻译机制从外源转录本重建叶绿体编码的类囊体膜蛋白的合成、靶向和插入的研究。
Using a homologous chloroplast translation system, we have reconstituted insertion and processing of the chloroplast-encoded thylakoid protein cytochrome f(pCytf). Cross-linking demonstrated that pCytfnascent chains when attached to the 70 S ribosome tightly interact with cpSecA, but this is strictly dependent on thylakoid membranes and a functional signal peptide. This indicates that cpSecA is only operative in pCytf biogenesis when it is bound to the membrane, most likely as part of the Sec translocon. No evidence for interaction between the 54-kDa subunit of the chloroplast signal recognition particle (cpSRP) and the pCytf nascent chain could be detected, suggesting that pCytf, in contrast to the polytopic D1 protein, does not require cpSRP for targeting. Insertion of pCytf occurred only co-translationally, resulting in processing and accumulation of both the processed signal peptide and the mature protein in the thylakoid. This co-translational membrane insertion and processing required a functional signal peptide and was inhibited by azide, demonstrating that cpSecA is essential for translocation of the soluble luminal domain. pCytfalso associated post-translationally with thylakoids, but the soluble N-terminal domain could not be translocated into the lumen. This is the first study in which synthesis, targeting, and insertion of a chloroplast-encoded thylakoid membrane protein is reconstituted from exogenous transcripts and using the chloroplast translational machinery.
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DOI：
10.1073/pnas.87.21.8227
发表时间：
1990-11-01
影响因子：
11.1
作者：
OLIVER, DB;CABELLI, RJ;JAROSIK, GP
通讯作者：
JAROSIK, GP
血红素在细胞色素 c6 生物合成中的作用。
DOI：
--
发表时间：
1994
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
Howe,G;Merchant,S
通讯作者：
Merchant,S
影响因子：
56.9
作者：
Jianguo Yuan;Ralph Henry;M. Mccaffery;Kenneth Cline
通讯作者：
Jianguo Yuan;Ralph Henry;M. Mccaffery;Kenneth Cline
影响因子：
3.3
作者：
R. Voelker;J. Mendel-Hartvig;A. Barkan
通讯作者：
R. Voelker;J. Mendel-Hartvig;A. Barkan
影响因子：
11.4
作者：
VOELKER, R;BARKAN, A
通讯作者：
BARKAN, A