Department of Chemical Engineering, Virginia Tech, Blacksburg, VA 24060, USA. Department of Inorganic and Analytic Chemistry, University of Geneva, 1205 Geneva, Switzerland.
Nanotechnology. 2016 Jun 24;27(25):255701. doi: 10.1088/0957-4484/27/25/255701. Epub 2016 May 13.
Tethering is used to deliver specific biological and industrial functions. For example, single-stranded DNA (ssDNA) is tethered to polymerases and long sequences of double-stranded DNA (dsDNA) during replication, and to solids in DNA microarrays. However, tethering ssDNA to a large object limits not only the available ssDNA conformations, but also the range of time-scales over which the mechanical responses of ssDNA are important. In this work we examine the effect of tethering by measurement of the mechanical response of ssDNA that is tethered at each end to two separate atomic force microscope cantilevers in aqueous solution. Thermal motion of the cantilevers drives the ends of the ssDNA chain at frequencies near 2 kHz. The presence of a tethered molecule makes a large difference to the asymmetric cross-correlation of two cantilevers, which enables resolution of the mechanical properties in our experiments. By analysis of the correlated motion of the cantilevers we extract the friction and stiffness of the ssDNA. We find that the measured friction is much larger than the friction that is usually associated with the unencumbered motion of ssDNA. We also find that the measured relaxation time, ∼30 μs, is much greater than prior measurements of the free-molecule relaxation time. We attribute the difference to the loss of conformational possibilities as a result of constraining the ends of the ssDNA.
tethering 用于传递特定的生物和工业功能。例如，在复制过程中，单链 DNA（ssDNA）被束缚在聚合酶和长序列的双链 DNA（dsDNA）上，在 DNA 微阵列中被束缚在固体上。然而，将 ssDNA 束缚在一个大物体上不仅限制了可用的 ssDNA 构象，还限制了 ssDNA 力学响应的时间尺度范围。在这项工作中，我们通过测量在水溶液中两端分别束缚在两个独立原子力显微镜悬臂上的 ssDNA 的力学响应来研究束缚的影响。悬臂的热运动以接近 2 kHz 的频率驱动 ssDNA 链的末端。束缚分子的存在对两个悬臂的非对称互相关有很大的影响，这使得我们的实验能够分辨出力学性质。通过对悬臂的相关运动进行分析，我们提取了 ssDNA 的摩擦和刚度。我们发现，测量到的摩擦比通常与 ssDNA 无阻碍运动相关的摩擦大得多。我们还发现，测量到的弛豫时间约为 30 μs，比先前自由分子弛豫时间的测量值大得多。我们将这种差异归因于 ssDNA 末端受到约束导致构象可能性的丧失。
