Calcium regulation of chloroplast protein translocation is mediated by calmodulin binding to Tic32
Calcium regulation of chloroplast protein translocation is mediated by calmodulin binding to Tic32
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DOI：
10.1073/pnas.0607150103
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发表时间：
2006-10-24
影响因子：
11.1
通讯作者：
Vothknecht, Ute C.
中科院分区：
文献类型：
--
作者：
Chigri, Fatima;Hoermann, Friederike;Vothknecht, Ute C.
核编码蛋白进入叶绿体的过程在包膜两侧受到严格控制。调节回路包括氧化还原控制和钙调节，钙调蛋白可能是后者的调节剂。使用钙调蛋白-琼脂糖亲和层析，我们可以确定内膜易位子成分 Tic32 是该膜的主要钙调蛋白结合蛋白。钙调蛋白结合测定证实了异源表达以及天然 Tic32 的相互作用。这种相互作用是钙依赖性的，并由靠近豌豆 Tic32 C 近端的 Leu-296 和 Leu-314 之间的钙调蛋白结合域介导。此外，我们可以将 Tic32 确定为真正的 NADPH 依赖性脱氢酶。 NADPH 但不影响 NADH 或 NADP(+) 影响 Tic110 与 Tic32 以及 Tic62 的相互作用。同时，Tic32的脱氢酶活性受到钙调蛋白的影响。特别是，NADPH 和钙调蛋白与 Tic32 的结合似乎是相互排斥的。这些结果表明叶绿体蛋白输入的氧化还原调节和钙调节在 Tic 易位子处进行，并且两者都可以由 Tic32 介导。
The import of nuclear-encoded proteins into chloroplasts is tightly controlled on both sides of the envelope membranes. Regulatory circuits include redox-control as well as calcium-regulation, with calmodulin being the likely mediator of the latter. Using affinity-chromatography on calmodulin-agarose, we could identify the inner envelope translocon component Tic32 as the predominant calmodulin-binding protein of this membrane. Calmodulin-binding assays corroborate the interaction for heterologously expressed as well as native Tic32. The interaction is calcium-dependent and is mediated by a calmodulin-binding domain between Leu-296 and Leu-314 close to the C-proximal end of the pea Tic32. We furthermore could establish Tic32 as a bona fide NADPH-dependent dehydrogenase. NADPH but not NADH or NADP(+) affects the interaction of Tic110 with Tic32 as well as Tic62. At the same time, dehydrogenase activity of Tic32 is affected by calmodulin. In particular, binding of NADPH and calmodulin to Tic32 appear to be mutually exclusive. These results suggest that redox modulation and calcium regulation of chloroplast protein import convene at the Tic translocon and that both could be mediated by Tic32.