Department of Chemistry and Chemical Engineering, California Institute of Technology , Pasadena, California 91125, United States.
Langmuir. 2013 Dec 31;29(52):16141-9. doi: 10.1021/la403262v. Epub 2013 Dec 11.
A method of DNA monolayer formation has been developed using copper-free click chemistry that yields enhanced surface homogeneity and enables variation in the amount of DNA assembled; extremely low-density DNA monolayers, with as little as 5% of the monolayer being DNA, have been formed. These DNA-modified electrodes (DMEs) were characterized visually, with AFM, and electrochemically, and were found to facilitate DNA-mediated reduction of a distally bound redox probe. These low-density monolayers were found to be more homogeneous than traditional thiol-modified DNA monolayers, with greater helix accessibility through an increased surface area-to-volume ratio. Protein binding efficiency of the transcriptional activator TATA-binding protein (TBP) was also investigated on these surfaces and compared to that on DNA monolayers formed with standard thiol-modified DNA. Our low-density monolayers were found to be extremely sensitive to TBP binding, with a signal decrease in excess of 75% for 150 nM protein. This protein was detectable at 4 nM, on the order of its dissociation constant, with our low-density monolayers. The improved DNA helix accessibility and sensitivity of our low-density DNA monolayers to TBP binding reflects the general utility of this method of DNA monolayer formation for DNA-based electrochemical sensor development.
已经开发出一种使用无铜点击化学的 DNA 单层形成方法，该方法可提高表面均一性，并能够改变组装的 DNA 量；形成了极低密度的 DNA 单层，其中单层的 DNA 含量低至 5%。这些 DNA 修饰电极 (DME) 通过视觉、AFM 和电化学进行了表征，发现它们有利于 DNA 介导的远程结合氧化还原探针的还原。与传统的硫醇修饰 DNA 单层相比，这些低密度单层更均匀，由于表面积与体积比增加，螺旋的可及性更高。还研究了转录激活因子 TATA 结合蛋白 (TBP) 在这些表面上的蛋白结合效率，并与使用标准硫醇修饰 DNA 形成的 DNA 单层进行了比较。我们的低密度单层对 TBP 结合非常敏感，超过 150 nM 蛋白时信号下降超过 75%。在我们的低密度单层上，这种蛋白质可检测到 4 nM，接近其离解常数。我们的低密度 DNA 单层对 TBP 结合的 DNA 螺旋可及性和灵敏度的提高反映了这种 DNA 单层形成方法在基于 DNA 的电化学传感器开发中的普遍适用性。
