Kavli Institute of Nanoscience, Delft University of Technology, Delft, The Netherlands.
Mol Cell. 2012 Jun 8;46(5):616-24. doi: 10.1016/j.molcel.2012.03.029. Epub 2012 May 3.
In E. coli homologous recombination, a filament of RecA protein formed on DNA searches and pairs a homologous sequence within a second DNA molecule with remarkable speed and fidelity. Here, we directly probe the strength of the two-molecule interactions involved in homology search and recognition using dual-molecule manipulation, combining magnetic and optical tweezers. We find that the filament's secondary DNA-binding site interacts with a single strand of the incoming double-stranded DNA during homology sampling. Recognition requires opening of the helix and is strongly promoted by unwinding torsional stress. Recognition is achieved upon binding of both strands of the incoming dsDNA to each of two ssDNA-binding sites in the filament. The data indicate a physical picture for homology recognition in which the fidelity of the search process is governed by the distance between the DNA-binding sites.
在大肠杆菌同源重组中，RecA 蛋白丝在 DNA 上形成，以惊人的速度和保真度搜索并将同源序列与第二个 DNA 分子中的同源序列配对。在这里，我们使用双分子操作，结合磁和光镊，直接探测同源搜索和识别中涉及的两种分子相互作用的强度。我们发现，丝的第二个 DNA 结合位点在同源采样过程中与进入的双链 DNA 的单链相互作用。识别需要打开螺旋，并且受到解旋扭转应力的强烈促进。当进入的双链 DNA 的两条链都与丝中的两个 ssDNA 结合位点结合时，就可以实现识别。这些数据为同源识别提供了一个物理图景，其中搜索过程的保真度由 DNA 结合位点之间的距离决定。
