Chu Xiakun, Muñoz Victor
IMDEA Nanosciences, Faraday 9, Campus de Cantoblanco, Madrid, 28049, Spain.
Phys Chem Chem Phys. 2017 Nov 1;19(42):28527-28539. doi: 10.1039/c7cp04380e.
Transcription factors are thought to efficiently search for their target DNA site via a combination of conventional 3D diffusion and 1D diffusion along the DNA molecule mediated by non-specific electrostatic interactions. This process requires the DNA-binding protein to quickly exchange between a search competent and a target recognition mode, but little is known as to how these two binding modes are encoded in the conformational properties of the protein. Here, we investigate this issue on the engrailed homeodomain (EngHD), a DNA-binding domain that folds ultrafast and exhibits a complex conformational behavior consistent with the downhill folding scenario. We explore the interplay between folding and DNA recognition using a coarse-grained computational model that allows us to manipulate the folding properties of the protein and monitor its non-specific and specific binding to DNA. We find that conformational disorder increases the search efficiency of EngHD by promoting a fast gliding search mode in addition to sliding. When gliding, EngHD remains loosely bound to DNA moving linearly along its length. A partially disordered EngHD also binds more dynamically to the target site, reducing the half-life of the specific complex via a spring-loaded mechanism. These findings apply to all conditions leading to partial disorder. However, we also find that at physiologically relevant temperatures EngHD is well folded and can only obtain the conformational flexibility required to accelerate 1D diffusion when it folds/unfolds within the downhill scenario (crossing a marginal free energy barrier). In addition, the conformational flexibility of native downhill EngHD enables its fast reconfiguration to lock into the specific binding site upon arrival, thereby affording finer control of the on- and off-rates of the specific complex. Our results provide key mechanistic insights into how DNA-binding domains optimize specific DNA recognition through the control of their conformational dynamics and folding mechanism.
转录因子被认为是通过传统的三维扩散和由非特异性静电相互作用介导的沿DNA分子的一维扩散相结合的方式，有效地搜索其目标DNA位点。这个过程要求DNA结合蛋白在搜索能力模式和目标识别模式之间快速切换，但对于这两种结合模式如何在蛋白质的构象特性中编码却知之甚少。在这里，我们以engrailed同源结构域（EngHD）为研究对象来探讨这个问题，EngHD是一种DNA结合结构域，其折叠速度超快，并且呈现出与下坡折叠情形相一致的复杂构象行为。我们使用一种粗粒度计算模型来探索折叠与DNA识别之间的相互作用，该模型使我们能够操控蛋白质的折叠特性，并监测其与DNA的非特异性和特异性结合。我们发现，构象无序除了促进滑动外，还通过促进快速滑动搜索模式来提高EngHD的搜索效率。在滑动时，EngHD与DNA保持松散结合，沿其长度线性移动。部分无序的EngHD与目标位点的结合也更具动态性，通过弹簧加载机制缩短了特异性复合物的半衰期。这些发现适用于所有导致部分无序的情况。然而，我们还发现，在生理相关温度下，EngHD折叠良好，只有当它在下坡情形下折叠/展开（跨越一个边缘自由能垒）时，才能获得加速一维扩散所需的构象灵活性。此外，天然下坡EngHD的构象灵活性使其在到达时能够快速重新配置，锁定到特异性结合位点，从而对特异性复合物的结合和解离速率进行更精细的控制。我们的结果为DNA结合结构域如何通过控制其构象动力学和折叠机制来优化特异性DNA识别提供了关键的机制性见解。
