Department of Physics, Harvard University, Cambridge, MA 02138, USA.
Proc Natl Acad Sci U S A. 2009 Nov 24;106(47):19824-9. doi: 10.1073/pnas.0911214106. Epub 2009 Nov 10.
Using a parallel single molecule magnetic tweezers assay we demonstrate homologous pairing of two double-stranded (ds) DNA molecules in the absence of proteins, divalent metal ions, crowding agents, or free DNA ends. Pairing is accurate and rapid under physiological conditions of temperature and monovalent salt, even at DNA molecule concentrations orders of magnitude below those found in vivo, and in the presence of a large excess of nonspecific competitor DNA. Crowding agents further increase the reaction rate. Pairing is readily detected between regions of homology of 5 kb or more. Detected pairs are stable against thermal forces and shear forces up to 10 pN. These results strongly suggest that direct recognition of homology between chemically intact B-DNA molecules should be possible in vivo. The robustness of the observed signal raises the possibility that pairing might even be the "default" option, limited to desired situations by specific features. Protein-independent homologous pairing of intact dsDNA has been predicted theoretically, but further studies are needed to determine whether existing theories fit sequence length, temperature, and salt dependencies described here.
使用平行单分子磁镊检测方法，我们证明了在没有蛋白质、二价金属离子、拥挤试剂或游离 DNA 末端的情况下，两条双链 (ds) DNA 分子的同源配对。在生理温度和单价盐条件下，即使在 DNA 分子浓度远低于体内浓度的情况下，以及在存在大量非特异性竞争 DNA 的情况下，配对也是准确和快速的。拥挤试剂进一步提高了反应速率。在同源性为 5kb 或更长的区域之间，很容易检测到配对。检测到的配对可以抵抗高达 10pN 的热和剪切力。这些结果强烈表明，在体内应该有可能直接识别化学完整的 B-DNA 分子之间的同源性。观察到的信号的稳健性提出了这样一种可能性，即配对甚至可能是“默认”选项，仅受特定特征限制的所需情况。完整 dsDNA 的无蛋白同源配对在理论上已被预测，但需要进一步研究以确定现有理论是否符合此处描述的序列长度、温度和盐依赖性。
