Modular DNA origami-based electrochemical detection of DNA and proteins
Byoung-jin Jeon, Matteo M. Guareschi, Jaimie M. Stewart, Emily Wu, Ashwin Gopinath, Netzahualcóyotl Arroyo-Currás, Philippe Dauphin-Ducharme, Kevin W. Plaxco, P… 展开作者
Byoung-jin Jeon, Matteo M. Guareschi, Jaimie M. Stewart, Emily Wu, Ashwin Gopinath, Netzahualcóyotl Arroyo-Currás, Philippe Dauphin-Ducharme, Kevin W. Plaxco, Philip S. Lukeman, Paul W. K. Rothemund
备注：
14 pages in main, 6 figures; 16 pages in supplementary information, 8 figures, 6 tables
研究开发了一种基于 DNA origami 的模块化电化学生物传感器，通过替换分析物结合域即可适配多种分子，利用统一的 100 纳米尺度构象变化实现核酸和蛋白质的单步定量检测，且具备优异的检测限与可再生性。
摘要：
生物标志物的多样性和异质性使得开发用于分析物单步定量的通用方法变得困难。对于单个生物标志物，通过检测分析物结合时亲和结合剂发生的构象变化来进行检测的电化学方法已展现出前景。然而，由于构象变化必须在纳米尺度的工作距离内运作，因此必须为每种分析物开发一种具有独特构象变化的全新传感器。在此，我们展示了一种由 DNA origami 构建的模块化电化学生物传感器，仅需替换其分析物结合域即可轻松适应多种分子。所有传感器变体均不依赖单个氧化还原报告基团的独特纳米尺度运动，而是依赖于相同的 100 纳米尺度构象变化，该变化使数十个报告基团足够靠近金电极表面，从而可通过方波伏安法（一种标准电化学技术）测量信号。为验证传感器的机制，我们使用单链 DNA 作为分析物，并优化了氧化还原报告基团的数量和各种连接子长度。通过简单地向适当的 DNA 连接子添加生物素或抗 PDGF 适配体，实现了传感器对链霉亲和素和 PDGF-BB 分析物的适应。经过几何优化的链霉亲和素传感器表现出的信号增益和检测限明显优于可比的无试剂电化学传感器。使用后，相同的传感器可在温和条件下再生：在经历四轮 DNA 链置换和重新杂交后，性能基本得以保持。通过利用 DNA 纳米结构的模块化特性，我们的工作为任意核酸和蛋白质的单步定量提供了一条直接的途径。
英文摘要：
The diversity and heterogeneity of biomarkers has made the development of general methods for single-step quantification of analytes difficult. For individual biomarkers, electrochemical methods that detect a conformational change in an affinity binder upon analyte binding have shown promise. However, because the conformational change must operate within a nanometer-scale working distance, an entirely new sensor, with a unique conformational change, must be developed for each analyte. Here, we demonstrate a modular electrochemical biosensor, built from DNA origami, which is easily adapted to diverse molecules by merely replacing its analyte binding domains. Instead of relying on a unique nanometer-scale movement of a single redox reporter, all sensor variants rely on the same 100-nanometer scale conformational change, which brings dozens of reporters close enough to a gold electrode surface that a signal can be measured via square wave voltammetry, a standard electrochemical technique. To validate our sensor's mechanism, we used single-stranded DNA as an analyte, and optimized the number of redox reporters and various linker lengths. Adaptation of the sensor to streptavidin and PDGF-BB analytes was achieved by simply adding biotin or anti-PDGF aptamers to appropriate DNA linkers. Geometrically-optimized streptavidin sensors exhibited signal gain and limit of detection markedly better than comparable reagentless electrochemical sensors. After use, the same sensors could be regenerated under mild conditions: performance was largely maintained over four cycles of DNA strand displacement and rehybridization. By leveraging the modularity of DNA nanostructures, our work provides a straightforward route to the single-step quantification of arbitrary nucleic acids and proteins.
DNA based nanostructures built on a long single stranded DNA scaffold, known as DNA origamis, offer the possibility to organize various molecules at the nanometer scale in one pot experiments. The folding of the scaffold is guaranteed by the presence of short, single stranded DNA sequences (staples), that hold together separate regions of the scaffold. In this paper, we modelize the annealing-melting properties of these DNA constructions. The model captures important features such as the hysteresis between melting and annealing, as well as the dependence upon the topology of the scaffold. We s…
We use the oxDNA coarse-grained model to provide a detailed characterization of the fundamental structural properties of DNA origamis, focussing on archetypal 2D and 3D origamis. The model reproduces well the characteristic pattern of helix bending in a 2D origami, showing that it stems from the intrinsic tendency of anti-parallel four-way junctions to splay apart, a tendency that is enhanced both by less screened electrostatic interactions and by increased thermal motion. We also compare to the structure of a 3D origami whose structure has been determined by cryo-electron microscopy. The oxDN…
The silicification of DNA origami structures increases their mechanical and thermal stability and provides chemical protection. So far, it is unclear how silicification affects the internal structure of the DNA origami and whether the whole DNA framework is embedded or if silica just forms an outer shell. By using in situ small-angle X-ray scattering (SAXS), we show that the net-cationic silica precursor TMAPS induces substantial condensation of the DNA origami, which is further enhanced by the addition of TEOS at early reaction times to an almost 10 % size reduction. We identify the SAXS Poro…
Biological materials are self-assembled with near-atomic precision in living cells, whereas synthetic 3D structures generally lack such precision and controllability. Recently, DNA nanotechnology, especially DNA origami technology, has been useful in the bottom-up fabrication of well-defined nanostructures ranging from tens of nanometres to sub-micrometres. In this Primer, we summarize the methodologies of DNA origami technology, including origami design, synthesis, functionalization and characterization. We highlight applications of origami structures in nanofabrication, nanophotonics and nan…
The specificity and simplicity of the Watson-Crick base pair interactions make DNA one of the most versatile construction materials for creating nanoscale structures and devices. Among several DNA-based approaches, the DNA origami technique excels in programmable self-assembly of complex, arbitrary shaped structures with dimensions of hundreds of nanometers. Importantly, DNA origami can be used as templates for assembly of functional nanoscale components into three-dimensional structures with high precision and controlled stoichiometry. This is often beyond the reach of other nanofabrication t…
Mechanically interlocked molecules have marked a breakthrough in the field of topological chemistry and boosted the vigorous development of molecular machinery. As an archetypal example of the interlocked molecules, catenanes comprise macrocycles that are threaded through one another like links in a chain. Inspired by the transition metal-templated approach of catenanes synthesis, the hierarchical assembly of DNA origami catenanes templated by gold nanoparticles is demonstrated in this work. DNA origami catenanes, which contain two, three or four interlocked rings are successfully created. In …
Intercalation of drug molecules into synthetic DNA nanostructures formed through self-assembled origami has been postulated as a valuable future method for targeted drug delivery. This is due to the excellent biocompatibility of synthetic DNA nanostructures, and high potential for flexible programmability including facile drug release into or near to target cells. Such favourable properties may enable high initial loading and efficient release for a predictable number of drug molecules per nanostructure carrier, important for efficient delivery of safe and effective drug doses to minimise non-…
We demonstrate hierarchical assembly of plasmonic toroidal metamolecules, which exhibit tailored optical activity in the visible spectral range. Each metamolecule consists of four identical origami-templated helical building blocks. Such toroidal metamolecules show stronger chiroptical response than monomers and dimers of the helical building blocks. Enantiomers of the plasmonic structures yield opposite circular dichroism spectra. The experimental results agree well with the theoretical simulations. We also demonstrate that given the circular symmetry of the structures, distinct chiroptical r…
Sliding is one of the fundamental mechanical movements in machinery. In macroscopic systems, double-rack pinion machines employ gears to slide two linear tracks along opposite directions. In microscopic systems, kinesin-5 proteins crosslink and slide apart antiparallel microtubules, promoting spindle bipolarity and elongation during mitosis. Here we demonstrate an artificial nanoscopic analog, in which gold nanocrystals can mediate coordinated sliding of two antiparallel DNA origami filaments powered by DNA fuels. Stepwise and reversible sliding along opposite directions is in situ monitored a…
In nano-optics, a formidable challenge remains in precise transport of a single optical nano-object along a programmed and routed path toward a predefined destination. Molecular motors in living cells that can walk directionally along microtubules have been the inspiration for realizing artificial molecular walkers. Here we demonstrate an active plasmonic system, in which a plasmonic nanorod can execute directional, progressive and reverse nanoscale walking on two or three-dimensional DNA origami.Such a walker comprises an anisotropic gold nanorod as its body and discrete DNA strands as its fe…
DNA nanotechnology allows for the realization of complex nanoarchitectures in which the spatial arrangements of different constituents and most functions can be enabled by DNA. When optically active components are integrated in such systems, the resulting nanoarchitectures not only provide great insights into the self-assembly of nanoscale elements in a systematic way but also impart tailored optical functionality to DNA origami. In this Letter, we demonstrate DNA-assembled multilayer nanosystems, which can carry out coordinated and reversible sliding motion powered by DNA fuels. Gold nanopart…
DNA origami is a modular platform for the combination of molecular and colloidal components to create optical, electronic, and biological devices. Integration of such nanoscale devices with microfabricated connectors and circuits is challenging: large numbers of freely diffusing devices must be fixed at desired locations with desired alignment. We present a DNA origami molecule whose energy landscape on lithographic binding sites has a unique maximum. This property enables device alignment within 3.2$^{\circ}$ on SiO$_2$. Orientation is absolute (all degrees of freedom are specified) and arbit…
Manipulation of temperature can be used to actuate DNA origami nano-hinges containing gold nanoparticles. We develop a physical model of this system that uses partition function analysis of the interaction between the nano-hinge and nanoparticle to predict the probability that the nano-hinge is open at a given temperature. The model agrees well with experimental data and predicts experimental conditions that allow the actuation temperature of the nano-hinge to be tuned over a range of temperatures from $30$${}^{\circ}\mathrm{C}$ to $45$${}^{\circ}\mathrm{C}$. Additionally, the model reveals su…
We introduce an extended version of oxDNA, a coarse-grained model of DNA designed to capture the thermodynamic, structural and mechanical properties of single- and double-stranded DNA. By including explicit major and minor grooves, and by slightly modifying the coaxial stacking and backbone-backbone interactions, we improve the ability of the model to treat large (kilobase-pair) structures such as DNA origami which are sensitive to these geometric features. Further, we extend the model, which was previously parameterised to just one salt concentration ([Na$^+$]=0.5M), so that it can be used fo…
