Targeting metastable coiled-coil domains by computational design
Targeting metastable coiled-coil domains by computational design
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DOI：
10.1021/ja802447e
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发表时间：
2008-09-10
影响因子：
15
通讯作者：
Alber, Tom
中科院分区：
文献类型：
--
作者：
Barth, Patrick;Schoeffler, Allyn;Alber, Tom
大约30%的真核生物基因组被预测编码部分未折叠蛋白。许多这些非结构化域接触多个合作伙伴在短暂的相互作用，细胞内稳态的关键。了解这些瞬时结合事件的功能意义是当前的挑战，可以解决与设计的肽抑制剂。然而，目前大多数蛋白质设计方法仅针对结构明确的稳定结构。为了解决这一限制，我们实施了一种计算设计策略，该策略在用于结合特异性的固定骨架序列搜索和所设计界面的结构优化之间交替。我们应用这种方法来创建特定的肽抑制剂的C-末端亚稳态卷曲螺旋结构域的基本酵母Septin Cdc 12 p。通过圆二色性和平衡超电泳证明了所设计序列的特异性结合。我们的研究结果验证了计算方法来设计特定的肽配体的蛋白质结构域缺乏内在的结构稳定性，并设置在体内Cdc 12 p卷曲螺旋功能的功能分析阶段。
Approximately 30% of eukaryotic genomes are predicted to encode partially unfolded proteins. Many of these unstructured domains contact multiple partners in short-lived interactions critical for cellular homeostasis. Understanding the functional implications of these transient binding events is a current challenge that could be addressed with designed peptide inhibitors. Most current protein design methodologies, however, target only structurally well-defined, stable structures. To address this limitation, we implemented a computational design strategy that alternates between a fixed backbone sequence search for binding specificity and structural optimization of the designed interfaces. We applied this method to create specific peptide inhibitors of the C-terminal metastable coiled-coil domain of the essential yeast septin Cdc12p. Specific binding of the designed sequences was demonstrated by circular dichroism and equilibrium ultracentrifugation. Our results validate computational methods to design specific peptide ligands to protein domains lacking intrinsic structural stability and set the stage for functional analysis of Cdc12p coiled coil function in vivo.