Identification of the Pore-forming Region of the Outer Chloroplast Envelope Protein OEP16*
Identification of the Pore-forming Region of the Outer Chloroplast Envelope Protein OEP16*
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叶绿体外膜蛋白 OEP16* 成孔区域的鉴定
DOI：
10.1074/jbc.275.16.11758
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发表时间：
2000
期刊：
影响因子：
--
通讯作者：
J. Soll
中科院分区：
文献类型：
--
作者：
T. Steinkamp;K. Hill;S. Hinnah;R. Wagner;T. Röhl;K. Pohlmeyer;J. Soll
叶绿体外被膜蛋白OEP 16形成阳离子选择性高电导通道，具有对胺和氨基酸的渗透性。直接参与通道形成的OEP 16区域已经通过对选择的重组OEP 16突变体的电生理分析来鉴定。由于这些突变体的分析依赖于重组蛋白的使用，我们评估了直接从豌豆叶绿体中分离的OEP 16和在大肠杆菌中产生的重组蛋白的电生理特性。结果表明，由天然豌豆OEP 16形成的通道的基本性质如电导率、选择性和开放概率与由该蛋白质的重组来源形成的通道活性相当。OEP 16突变体的电生理学分析后，我们发现，点突变和插入额外的氨基酸残基在推定的螺旋1（Glu 73到Val 91）的区域没有改变的OEP 16通道的特性。唯一的例外是Cys 71 →Ser突变，这导致通道的CuCl 2敏感性丧失。对OEP 16的N-和C-末端缺失突变体和含有确定的改组结构域的突变体的分析表明，能够在脂质体中形成通道的OEP 16的最小连续区域位于氨基酸残基21和93之间的蛋白质的前半部分。
The chloroplast outer envelope protein OEP16 forms a cation-selective high conductance channel with permeability to amines and amino acids. The region of OEP16 directly involved in channel formation has been identified by electrophysiological analysis of a selection of reconstituted OEP16 mutants. Because analysis of these mutants depended on the use of recombinant protein, we evaluated the electrophysiological properties of OEP16 isolated directly from pea chloroplasts and of the recombinant protein produced in Escherichia coli. The results show that the basic properties like conductance, selectivity, and open probability of the channel formed by native pea OEP16 are comparable with the channel activity formed by the recombinant source of the protein. Following electrophysiological analysis of OEP16 mutants we found that point mutations and insertion of additional amino acid residues in the region of the putative helix 1 (Glu73 to Val91) did not change the properties of the OEP16 channel. The only exception was a Cys71→Ser mutation, which led to a loss of the CuCl2 sensitivity of the channel. Analysis of N- and C-terminal deletion mutants of OEP16 and mutants containing defined shuffled domains indicated that the minimal continuous region of OEP16, which is able to form a channel in liposomes, lies in the first half of the protein between amino acid residues 21 and 93.
DOI：
10.1016/s0021-9258(17)41716-9
发表时间：
1994-02
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
Hiroshi Nikaido
通讯作者：
Hiroshi Nikaido
DOI：
10.1016/s0021-9258(18)45908-x
发表时间：
1992-02
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
Etsuko SugawaraS;Hiroshi Nikaido
通讯作者：
Etsuko SugawaraS;Hiroshi Nikaido
蛋白质在通道结构中折叠。
影响因子：
6.8
作者：
Montal,M
通讯作者：
Montal,M