Dual Aptamer-Functionalized 3D Plasmonic Metamolecule for Thrombin Sensing

Abstract

DNA nanotechnology offers the possibility to rationally design structures with emergent properties by precisely controlling their geometry and functionality. Here, we demonstrate a DNA-based plasmonic metamolecule that is capable of sensing human thrombin proteins. The chiral reconfigurability of a DNA origami structure carrying two gold nanorods was used to provide optical read-out of thrombin binding through changes in the displayed plasmonic circular dichroism. In our experiments, each arm of the structure was modified with one of two different thrombin-binding aptamers—thrombin-binding aptamer (TBA) and HD22—in such a way that a thrombin molecule could be sandwiched by the aptamers to lock the metamolecule in a state of defined chirality. Our structure exhibited a Kd of 1.4 nM, which was an order of magnitude lower than those of the individual aptamers. The increased sensitivity arose from the avidity gained by the cooperative binding of the two aptamers, which was also reflected by a Hill coefficient of 1.3 ± 0.3. As we further exploited the strong plasmonic circular dichroism (CD) signals of the metamolecule, our method allowed one-step, high sensitivity optical detection of human thrombin proteins in solution.

Challenges and Perspectives of DNA Nanostructures in Biomedicine

“…[73] Thegeneral detection concepts introduced above can also be adapted for the detection of proteins and other biomarkers.T his usually requires the introduction of target-specific aptamers into the DN. [74] Detection of the bound analyte can then be achieved using AFM, [70,75] electrochemistry, [76] fluorimetry, [60] and CD spectroscopy, [77,78] and successful target detection was recently demonstrated even in whole blood. [76] Nevertheless,most of these works can be considered proof-ofprinciple studies that employed well-characterized aptamers with ah igh affinity toward some model targets such as thrombin, [75,78] ATP, [60,76] or adenosine.…”

“…[74] Detection of the bound analyte can then be achieved using AFM, [70,75] electrochemistry, [76] fluorimetry, [60] and CD spectroscopy, [77,78] and successful target detection was recently demonstrated even in whole blood. [76] Nevertheless,most of these works can be considered proof-ofprinciple studies that employed well-characterized aptamers with ah igh affinity toward some model targets such as thrombin, [75,78] ATP, [60,76] or adenosine. [77] Successful aptamerbased detection of an infection-related target was demonstrated by Godonoga et al,w ho incorporated aptamers against the malaria-protein biomarker PfLDH in 2D DO substrates (Figure 3c).…”

DNA Origami as Emerging Technology for the Engineering of Fluorescent and Plasmonic-Based Biosensors

“…These interactions can be translated into optical signals by plasmonic or by fluorescence excitation [50,51]. Optical DNA origami sensors are developing into sensitive, rapid, accurate, flexible, and multi-target sensing tools with a low limit of detection (LOD) [18,19,21,[52][53][54][55]. They are used in biology, environment monitoring, and the food safety industry, and have a great potential for applications in biomedicine to detect and monitor diseases and pathogens [56].…”

“…However, in terms of sensing and qualitatively detection of targets, dynamic acting plasmonic DNA nanostructures are more promising. Different dynamic designs were proposed, including tetrahedron metamolecule [135], a DNA-guided plasmonic helix [136], or the two-arms metamolecule [19,20,52,137,138]. Nevertheless, to the best of our knowledge, only the two-armed metamolecule was used for sensing (Figure 6).…”

Herausforderungen und Perspektiven von DNA‐Nanostrukturen in der Biomedizin

“…Dies erfordert normalerweise die Einbringung von Target-spezifischen Aptameren in die DN. [74] Die Detektion des gebundenen Analyten kann dann wieder mittels AFM, [70,75] Elektrochemie, [76] Fluorimetrie [60] und CD-Spektroskopie [77,78] erfolgen. Erfolgreiche Target-Detektion wurde kürzlich sogar in Vollblut demonstriert.…”