While the field of computational protein design has witnessed amazing progression in recent years, folding properties still constitute a significant barrier towards designing new and larger proteins. In order to assess and improve folding properties of designed proteins, we have developed a genetics-based folding assay and selection system based on the essential enzyme, orotate phosphoribosyl transferase from Escherichia coli. This system allows for both screening of candidate designs with good folding properties and genetic selection of improved designs. Thus, we identified single amino acid substitutions in two failed designs that rescued poorly folding and unstable proteins. Furthermore, when these substitutions were transferred into a well-structured design featuring a complex folding profile, the resulting protein exhibited native-like cooperative folding with significantly improved stability. In protein design, a single amino acid can make the difference between folding and misfolding, and this approach provides a useful new platform to identify and improve candidate designs.

中文翻译：

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改善计算设计的蛋白质的折叠特性。

尽管近年来蛋白质计算设计领域取得了惊人的进步，但折叠特性仍然构成设计新的更大蛋白质的重要​​障碍。为了评估和改善设计蛋白质的折叠特性，我们开发了一种基于遗传学的折叠测定和选择系统，该系统基于必需酶，来自大肠杆菌的乳清酸磷酸核糖基转移酶。该系统既可以筛选具有良好折叠特性的候选设计，又可以进行改良设计的遗传选择。因此，我们在两个失败的设计中鉴定出单个氨基酸取代，这些设计挽救了折叠不良和不稳定的蛋白质。此外，当这些替代品转移到结构复杂，折叠轮廓复杂的设计中时，所得蛋白质显示出类似天然的协同折叠，并显着提高了稳定性。在蛋白质设计中，单个氨基酸可以在折叠和错误折叠之间发挥作用，这种方法为识别和改进候选设计提供了一个有用的新平台。