VIPP1 Involved in Chloroplast Membrane Integrity Has GTPase Activity in Vitro
VIPP1 Involved in Chloroplast Membrane Integrity Has GTPase Activity in Vitro
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VIPP1 参与叶绿体膜完整性，在体外具有 GTP 酶活性
DOI：
10.1104/pp.18.00145
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发表时间：
2018
期刊：
影响因子：
7.4
通讯作者：
Sakamoto Wataru
中科院分区：
文献类型：
--
作者：
Ohnishi Norikazu;Zhang Lingang;Sakamoto Wataru
PLASTID1 中的囊泡诱导蛋白 (VIPP1) 在产氧光合生物中是保守的，并且似乎与细菌 PspA 蛋白不同。 VIPP1 定位于叶绿体包膜和类囊体膜，在那里形成高分子量的同聚体。尽管VIPP1的多种作用已被推断，包括类囊体膜形成、包膜维持、膜融合和光合活性调节，但其在叶绿体膜质量控制中的确切作用仍不清楚。 VIPP1 通过其氨基末端结构域形成寡聚体，并以 Mg2+ 依赖性方式触发膜融合。我们之前证明拟南芥（Arabidopsis thaliana）VIPP1也表现出响应体内渗透压和热应激的动态复合体分解。这些结果表明 VIPP1 介导叶绿体中的膜融合/重塑。考虑到调节细胞内膜融合/重塑事件的蛋白质机器通常需要 GTP 结合和/或水解的能力，我们质疑 VIPP1 是否具有类似的特性。我们使用大肠杆菌表达的纯化 VIPP1-His 融合蛋白进行了体外测定。 VIPP1-His 显示 GTP 水解活性，该活性被不可水解的 GTP 类似物 GTPγS 竞争性抑制，并且依赖于 GTP 结合。特别有趣的是，来自 E. 的祖先 PspA。大肠杆菌也具有GTP水解活性。尽管 VIPP1 不包含规范的 G 结构域，但发现氨基末端 α 螺旋对于 GTP 结合和 GTP 水解以及寡聚物的形成都很重要。总的来说，我们的结果表明 VIPP1/PspA 的特性与 GTPases 相似。
VESICLE-INDUCING PROTEIN IN PLASTID1 (VIPP1) is conserved among oxygenic photosynthetic organisms and appears to have diverged from the bacterial PspA protein. VIPP1 localizes to the chloroplast envelope and thylakoid membrane, where it forms homooligomers of high molecular mass. Although multiple roles of VIPP1 have been inferred, including thylakoid membrane formation, envelope maintenance, membrane fusion, and regulation of photosynthetic activity, its precise role in chloroplast membrane quality control remains unknown. VIPP1 forms an oligomer through its amino-terminal domain and triggers membrane fusion in an Mg2+-dependent manner. We previously demonstrated that Arabidopsis (Arabidopsis thaliana) VIPP1 also exhibits dynamic complex disassembly in response to osmotic and heat stresses in vivo. These results suggest that VIPP1 mediates membrane fusion/remodeling in chloroplasts. Considering that protein machines that regulate intracellular membrane fusion/remodeling events often require a capacity for GTP binding and/or hydrolysis, we questioned whether VIPP1 has similar properties. We conducted an in vitro assay using a purified VIPP1-His fusion protein expressed inEscherichia colicells. VIPP1-His showed GTP hydrolysis activity that was inhibited competitively by an unhydrolyzable GTP analog, GTPγS, and that depends on GTP binding. It is particularly interesting that the ancestral PspA fromE. colialso possesses GTP hydrolysis activity. Although VIPP1 does not contain a canonical G domain, the amino-terminal α-helix was found to be important for both GTP binding and GTP hydrolysis as well as for oligomer formation. Collectively, our results reveal that the properties of VIPP1/PspA are similar to those of GTPases.