Laboratory of Single Molecule Biophysics, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, United States.
Department of Biochemistry, ELTE-MTA "Momentum" Motor Enzymology Research Group, Eötvös Loránd University, Budapest, Hungary.
Elife. 2019 Aug 29;8:e45909. doi: 10.7554/eLife.45909.
RecQ helicases promote genomic stability through their unique ability to suppress illegitimate recombination and resolve recombination intermediates. These DNA structure-specific activities of RecQ helicases are mediated by the helicase-and-RNAseD like C-terminal (HRDC) domain, via unknown mechanisms. Here, employing single-molecule magnetic tweezers and rapid kinetic approaches we establish that the HRDC domain stabilizes intrinsic, sequence-dependent, pauses of the core helicase (lacking the HRDC) in a DNA geometry-dependent manner. We elucidate the core unwinding mechanism in which the unwinding rate depends on the stability of the duplex DNA leading to transient sequence-dependent pauses. We further demonstrate a non-linear amplification of these transient pauses by the controlled binding of the HRDC domain. The resulting DNA sequence- and geometry-dependent pausing may underlie a homology sensing mechanism that allows rapid disruption of unstable (illegitimate) and stabilization of stable (legitimate) DNA strand invasions, which suggests an intrinsic mechanism of recombination quality control by RecQ helicases.
RecQ 解旋酶通过其抑制非同源重组和解决重组中间体的独特能力来促进基因组稳定性。RecQ 解旋酶的这些 DNA 结构特异性活性是通过未知机制由解旋酶和 RNA 酶 D 样 C 末端（HRDC）结构域介导的。在这里，我们采用单分子磁镊和快速动力学方法，建立了 HRDC 结构域以 DNA 几何结构依赖的方式稳定核心解旋酶（缺乏 HRDC）的固有、序列依赖性停顿的方式。我们阐明了核心解旋机制，其中解旋速率取决于导致瞬时序列依赖性停顿的双链 DNA 的稳定性。我们进一步通过 HRDC 结构域的受控结合证明了这些瞬时停顿的非线性放大。由此产生的 DNA 序列和几何结构依赖性停顿可能是同源性感应机制的基础，该机制允许快速破坏不稳定（非同源）和稳定（同源）DNA 链入侵，这表明 RecQ 解旋酶通过内在机制进行重组质量控制。
