Protein secondary structure determines the temporal relationship between folding and disulfide formation
Protein secondary structure determines the temporal relationship between folding and disulfide formation
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DOI：
10.1074/jbc.ra119.011983
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发表时间：
2020-02-21
影响因子：
4.8
通讯作者：
Bulleid, Neil J.
中科院分区：
文献类型：
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作者：
Robinson, Philip J.;Kanemura, Shingo;Bulleid, Neil J.
相对于蛋白质的折叠阶段，蛋白质中的二硫键是如何以及何时形成的，这是细胞生物学中的一个基本问题。有两个模型描述了这种关系：折叠前体模型和准随机模型，前者在二硫化物形成之前形成新生结构，后者在折叠之前形成。在这里，我们研究了三种结构不同的底物-2-微球蛋白、催乳素和ADAM10的去整合素域(ADAM10)的氧化折叠，以了解这些机制如何应用于细胞环境。我们使用了一个真核无细胞翻译系统，在这个系统中，我们可以识别停滞的翻译中间产物中的二硫异构体，以表征与内质网转位和非天然二硫的存在相关的二硫化物形成的时间。我们的结果表明，在缺乏二级结构的结构域中，二硫键在构象折叠之前形成，这是一个容易形成非天然二硫键的过程，而在具有特定二级结构的蛋白质中，天然二硫键的形成发生在部分折叠之后。这些发现表明，新生的蛋白质结构促进了共翻译折叠过程中正确的二硫键形成。
How and when disulfide bonds form in proteins relative to the stage of their folding is a fundamental question in cell biology. Two models describe this relationship: the folded precursor model, in which a nascent structure forms before disulfides do, and the quasi-stochastic model, where disulfides form prior to folding. Here we investigated oxidative folding of three structurally diverse substrates, ?2-microglobulin, prolactin, and the disintegrin domain of ADAM metallopeptidase domain 10 (ADAM10), to understand how these mechanisms apply in a cellular context. We used a eukaryotic cell-free translation system in which we could identify disulfide isomers in stalled translation intermediates to characterize the timing of disulfide formation relative to translocation into the endoplasmic reticulum and the presence of non-native disulfides. Our results indicate that in a domain lacking secondary structure, disulfides form before conformational folding through a process prone to nonnative disulfide formation, whereas in proteins with defined secondary structure, native disulfide formation occurs after partial folding. These findings reveal that the nascent protein structure promotes correct disulfide formation during cotranslational folding.