Nanomechanical measurements of the sequence-dependent folding landscapes of single nucleic acid hairpins
Nanomechanical measurements of the sequence-dependent folding landscapes of single nucleic acid hairpins
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DOI：
10.1073/pnas.0511048103
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发表时间：
2006-04-18
影响因子：
11.1
通讯作者：
Block, SM
中科院分区：
文献类型：
--
作者：
Woodside, MT;Behnke-Parks, WM;Block, SM
核酸发夹结构为探索二级结构的形成提供了一个强有力的模型系统。我们报告了一个系统的研究动力学和热力学的折叠过渡的个别DNA发夹不同的茎长，环长，茎GC含量。折叠是在高分辨率的光学陷阱中使用具有快速响应时间的独特的力钳装置机械诱导的。我们测量了20个不同的发夹序列，其具有6-30 bp长的准随机茎序列，3-30 nt长的多聚胸苷环，以及0%至100%范围内的茎GC含量。对于所有的发夹研究，折叠和展开的特点是一个单一的过渡。根据这些速率的力依赖性，我们确定了能垒的位置和高度，发现双链体形成的过渡态涉及环旁边1-2 bp的形成。通过测量展开能量跨越一个数量级，过渡率覆盖六个数量级，发夹开口距离与亚纳米精度，我们的研究结果定义的能量景观折叠的基本特征。我们发现定量协议在整个测量范围内的混合景观模型，结合热力学最近邻自由能和纳米机械DNA拉伸能。
Nucleic acid hairpins provide a powerful model system for probing the formation of secondary structure. We report a systematic study of the kinetics and thermodynamics of the folding transition for individual DNA hairpins of varying stem length, loop length, and stem GC content. Folding was induced mechanically in a high-resolution optical trap using a unique force clamp arrangement with fast response times. We measured 20 different hairpin sequences with quasi-random stem sequences that were 6-30 bp long, polythymidine loops that were 3-30 nt long, and stem GC content that ranged from 0% to 100%. For all hairpins studied, folding and unfolding were characterized by a single transition. From the force dependence of these rates, we determined the position and height of the energy barrier, finding that the transition state for duplex formation involves the formation of 1-2 bp next to the loop. By measuring unfolding energies spanning one order of magnitude, transition rates covering six orders of magnitude, and hairpin opening distances with subnanometer precision, our results define the essential features of the energy landscape for folding. We find quantitative agreement over the entire range of measurements with a hybrid landscape model that combines thermodynamic nearest-neighbor free energies and nano-mechanical DNA stretching energies.