The Role of Noncognate Sites in the 1D Search Mechanism of EcoRI
The Role of Noncognate Sites in the 1D Search Mechanism of EcoRI
复制标题
DOI：
10.1016/j.bpj.2019.04.035
复制
发表时间：
2019-06-18
影响因子：
3.4
通讯作者：
Price, Allen C.
中科院分区：
文献类型：
--
作者：
Piatt, Sadie C.;Loparo, Joseph J.;Price, Allen C.
一维搜索是DNA目标识别的重要步骤。理论研究表明，一维扩散的序列依赖性可以帮助解决快速搜索和高目标亲和力的竞争需求，这种冲突被称为速度选择悖论。该分辨率要求扩散能景观与潜在的特定结合能相关。在这项工作中，我们报告了量子点标记EcoRI的一维搜索观测结果。我们的数据支持这样一种观点，即蛋白质通过旋转耦合滑动在波纹状能量景观上搜索DNA。我们观察到，虽然EcoRI主要在低盐浓度下沿着DNA滑动，但在高浓度下，它的扩散是滑动和跳跃的结合。我们还观察到基因组星形位点的长时间停顿，这些位点与目标序列只有一个核苷酸的差异。为了使这些观察结果与先前的生化和结构数据相一致，我们提出了一种搜索模型，在该模型中，蛋白质在快速搜索期间滑过序列无关的能量景观，但迅速与“半特异性”结合模式相互转化，其中半位点被探测。这种半位点相互作用稳定了过渡到完全特定的结合模式，然后可以导致目标识别。
A one-dimensional (1D) search is an essential step in DNA target recognition. Theoretical studies have suggested that the sequence dependence of 1D diffusion can help resolve the competing demands of a fast search and high target affinity, a conflict known as the speed-selectivity paradox. The resolution requires that the diffusion energy landscape is correlated with the underlying specific binding energies. In this work, we report observations of a 1D search by quantum dot-labeled EcoRI. Our data supports the view that proteins search DNA via rotation-coupled sliding over a corrugated energy landscape. We observed that whereas EcoRI primarily slides along DNA at low salt concentrations, at higher concentrations, its diffusion is a combination of sliding and hopping. We also observed long-lived pauses at genomic star sites, which differ by a single nucleotide from the target sequence. To reconcile these observations with prior biochemical and structural data, we propose a model of search in which the protein slides over a sequence-independent energy landscape during fast search but rapidly interconverts with a "hemispecific" binding mode in which a half site is probed. This half site interaction stabilizes the transition to a fully specific mode of binding, which can then lead to target recognition.