Cooperative Protein Folding by Two Protein Thiol Disulfide Oxidoreductases and ERO1 in Soybean
Cooperative Protein Folding by Two Protein Thiol Disulfide Oxidoreductases and ERO1 in Soybean
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DOI：
10.1104/pp.15.01781
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发表时间：
2016-02-01
期刊：
影响因子：
7.4
通讯作者：
Urade, Reiko
中科院分区：
文献类型：
--
作者：
Matsusaki, Motonori;Okuda, Aya;Urade, Reiko
真核细胞内质网 (ER) 中产生的大多数蛋白质通过二硫键形成（氧化折叠）进行折叠。氧化折叠由蛋白质二硫键异构酶（PDI）和PDI相关的内质网蛋白硫醇二硫键氧化还原酶（ER氧化还原酶）催化。在酵母和哺乳动物中，ER 氧化还原素-1 (Ero1) 提供相当于 PDI 活性中心的氧化作用。在这项研究中，我们表达了重组大豆 Ero1 (GmERO1a)，发现 GmERO1a 氧化多种大豆 ER 氧化还原酶，而哺乳动物 Ero1 对 PDI 具有高度特异性。其中一种 ER 氧化还原酶 GmPDIM 在体内和体外与 GmPDIL-2 相关，但不能被 GmERO1a 氧化。因此，我们在体外研究了 GmPDIM、GmERO1a 和 GmPDIL-2 可能的协同氧化折叠，发现 GmPDIL-2 协同加速氧化重折叠。在此过程中，GmERO1a优先氧化GmPDIM的a、a'、b结构域中a'结构域的活性中心。引入GmPDIM的a'结构域活性中心的二硫键显示转移到GmPDIM的a结构域的活性中心，并且GmPDIM的a结构域直接氧化GmPDIL-2的a或a'结构域的活性中心。因此，我们提出，氧化当量从一种内质网氧化还原酶到另一种内质网氧化还原酶的传递可能在植物中多种内质网氧化还原酶的协同氧化折叠中发挥重要作用。
Most proteins produced in the endoplasmic reticulum (ER) of eukaryotic cells fold via disulfide formation (oxidative folding). Oxidative folding is catalyzed by protein disulfide isomerase (PDI) and PDI-related ER protein thiol disulfide oxidoreductases (ER oxidoreductases). In yeast and mammals, ER oxidoreductin-1s (Ero1s) supply oxidizing equivalent to the active centers of PDI. In this study, we expressed recombinant soybean Ero1 (GmERO1a) and found that GmERO1a oxidized multiple soybean ER oxidoreductases, in contrast to mammalian Ero1s having a high specificity for PDI. One of these ER oxidoreductases, GmPDIM, associated in vivo and in vitro with GmPDIL-2, was unable to be oxidized by GmERO1a. We therefore pursued the possible cooperative oxidative folding by GmPDIM, GmERO1a, and GmPDIL-2 in vitro and found that GmPDIL-2 synergistically accelerated oxidative refolding. In this process, GmERO1a preferentially oxidized the active center in the a' domain among the a, a', and b domains of GmPDIM. A disulfide bond introduced into the active center of the a' domain of GmPDIM was shown to be transferred to the active center of the a domain of GmPDIM and the a domain of GmPDIM directly oxidized the active centers of both the a or a' domain of GmPDIL-2. Therefore, we propose that the relay of an oxidizing equivalent from one ER oxidoreductase to another may play an essential role in cooperative oxidative folding by multiple ER oxidoreductases in plants.