On the biophysics and kinetics of toehold-mediated DNA strand displacement
TL;DR: This work study strand displacement at multiple levels of detail, using an intuitive model of a random walk on a 1D energy landscape, a secondary structure kinetics model with single base-pair steps and a coarse-grained molecular model that incorporates 3D geometric and steric effects to provide a biophysical explanation of strand displacement kinetics.
Abstract: Dynamic DNA nanotechnology often uses toehold-mediated strand displacement for controlling reaction kinetics. Although the dependence of strand displacement kinetics on toehold length has been experimentally characterized and phenomenologically modeled, detailed biophysical understanding has remained elusive. Here, we study strand displacement at multiple levels of detail, using an intuitive model of a random walk on a 1D energy landscape, a secondary structure kinetics model with single base-pair steps and a coarse-grained molecular model that incorporates 3D geometric and steric effects. Further, we experimentally investigate the thermodynamics of three-way branch migration. Two factors explain the dependence of strand displacement kinetics on toehold length: (i) the physical process by which a single step of branch migration occurs is significantly slower than the fraying of a single base pair and (ii) initiating branch migration incurs a thermodynamic penalty, not captured by state-of-the-art nearest neighbor models of DNA, due to the additional overhang it engenders at the junction. Our findings are consistent with previously measured or inferred rates for hybridization, fraying and branch migration, and they provide a biophysical explanation of strand displacement kinetics. Our work paves the way for accurate modeling of strand displacement cascades, which would facilitate the simulation and construction of more complex molecular systems.
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TL;DR: This review provides a comprehensive survey of recent developments in DNA origami structure, design, assembly, and directed self-assembly, as well as its broad applications.
Abstract: DNA has become one of the most extensively used molecular building blocks for engineering self-assembling materials. DNA origami is a technique that uses hundreds of short DNA oligonucleotides, called staple strands, to fold a long single-stranded DNA, which is called a scaffold strand, into various designer nanoscale architectures. DNA origami has dramatically improved the complexity and scalability of DNA nanostructures. Due to its high degree of customization and spatial addressability, DNA origami provides a versatile platform with which to engineer nanoscale structures and devices that can sense, compute, and actuate. These capabilities open up opportunities for a broad range of applications in chemistry, biology, physics, material science, and computer science that have often required programmed spatial control of molecules and atoms in three-dimensional (3D) space. This review provides a comprehensive survey of recent developments in DNA origami structure, design, assembly, and directed self-assemb...
755 citations
TL;DR: This work discusses how QDs are steadily revolutionizing the development of new biosensors along with a myriad of other photonically active nanomaterial-based bioconjugates, and how this field has grown over a relatively short time span.
Abstract: Luminescent semiconductor quantum dots (QDs) are one of the more popular nanomaterials currently utilized within biological applications. However, what is not widely appreciated is their growing role as versatile energy transfer (ET) donors and acceptors within a similar biological context. The progress made on integrating QDs and ET in biological configurations and applications is reviewed in detail here. The goal is to provide the reader with (1) an appreciation for what QDs are capable of in this context, (2) how this field has grown over a relatively short time span, and, in particular, (3) how QDs are steadily revolutionizing the development of new biosensors along with a myriad of other photonically active nanomaterial-based bioconjugates. An initial discussion of QD materials along with key concepts surrounding their preparation and bioconjugation is provided given the defining role these aspects play in the QDs ability to succeed in subsequent ET applications. The discussion is then divided around...
542 citations
TL;DR: It is demonstrated that discrete three-dimensional DNA components can specifically self-assemble in solution on the basis of shape-complementarity and without base pairing, and may be finely controlled by global parameters such as cation concentration or temperature and by an allosteric mechanism based on strand-displacement reactions.
Abstract: We demonstrate that discrete three-dimensional (3D) DNA components can specifically self-assemble in solution on the basis of shape-complementarity and without base pairing. Using this principle, we produced homo- and heteromultimeric objects, including micrometer-scale one- and two-stranded filaments and lattices, as well as reconfigurable devices, including an actuator, a switchable gear, an unfoldable nanobook, and a nanorobot. These multidomain assemblies were stabilized via short-ranged nucleobase stacking bonds that compete against electrostatic repulsion between the components' interfaces. Using imaging by electron microscopy, ensemble and single-molecule fluorescence resonance energy transfer spectroscopy, and electrophoretic mobility analysis, we show that the balance between attractive and repulsive interactions, and thus the conformation of the assemblies, may be finely controlled by global parameters such as cation concentration or temperature and by an allosteric mechanism based on strand-displacement reactions.
537 citations
TL;DR: The enormous progress of dynamic DNA nanotechnology over the past years seems poised for practical application, with toehold-mediated strand displacement reactions used with ever increasing sophistication.
Abstract: Dynamic DNA nanotechnology, a subfield of DNA nanotechnology, is concerned with the study and application of nucleic acid strand-displacement reactions. Strand-displacement reactions generally proceed by three-way or four-way branch migration and initially were investigated for their relevance to genetic recombination. Through the use of toeholds, which are single-stranded segments of DNA to which an invader strand can bind to initiate branch migration, the rate with which strand displacement reactions proceed can be varied by more than 6 orders of magnitude. In addition, the use of toeholds enables the construction of enzyme-free DNA reaction networks exhibiting complex dynamical behavior. A demonstration of this was provided in the year 2000, in which strand displacement reactions were employed to drive a DNA-based nanomachine (Yurke, B.; et al. Nature 2000, 406, 605–608). Since then, toehold-mediated strand displacement reactions have been used with ever increasing sophistication and the field of dynam...
413 citations
TL;DR: A DNA robot is demonstrated that performs a nanomechanical task substantially more sophisticated than previous work and modularity could allow diverse new functions performed by robots using the same set of building blocks.
Abstract: INTRODUCTION Since the 1980s, the design and synthesis of molecular machines has been identified as a grand challenge for molecular engineering. Robots are an important type of molecular machine that automatically carry out complex nanomechanical tasks. DNA molecules are excellent materials for building molecular robots, because their geometric, thermodynamic, and kinetic properties are well understood and highly programmable. So far, the development of DNA robots has been limited to simple functions. Most DNA robots were designed to perform a single function: walking in a controlled direction. A few demonstrations included a second function combined with walking (for example, picking up nanoparticles or choosing a path at a junction). However, these relatively more complex functions were also more difficult to control, and the complexity of the tasks was limited to what the robot can perform within 3 to 12 steps. In addition, each robot design was tailored for a specific task, complicating efforts to develop new robots that perform new tasks by combining functions and mechanisms. RATIONALE The design and synthesis of molecular robots presents two critical challenges, those of modularity and algorithm simplicity, which have been transformative in other areas of molecular engineering. For example, simple and modular building blocks have been used for scaling up molecular information processing with DNA circuits. As in DNA circuits, simple building blocks for DNA robots could enable more complex nanomechanical tasks, whereas modularity could allow diverse new functions performed by robots using the same set of building blocks. RESULTS We demonstrate a DNA robot that performs a nanomechanical task substantially more sophisticated than previous work. We developed a simple algorithm and three modular building blocks for a DNA robot that performs autonomous cargo sorting. The robot explores a two-dimensional testing ground on the surface of DNA origami, picks up multiple cargos of two types that are initially at unordered locations, and delivers each type to a specified destination until all cargo molecules are sorted into two distinct piles. The robot is designed to perform a random walk without any energy supply. Exploiting this feature, a single robot can repeatedly sort multiple cargos. Localization on DNA origami allows for distinct cargo-sorting tasks to take place simultaneously in one test tube or for multiple robots to collectively perform the same task. On average, our robot performed approximately 300 steps while sorting the cargos. The number of steps is one to two magnitudes larger than the previously demonstrated DNA robots performing additional tasks while walking. Using exactly the same robot design, the system could be generalized to multiple types of cargos with arbitrary initial distributions, and to many instances of distinct tasks in parallel, whereas each task can be assigned a distinct number of robots depending on the difficulty of the task. CONCLUSION Using aptamers, antibodies, or direct conjugation, small chemicals, metal nanoparticles, and proteins could be transported as cargo molecules so that the cargo-sorting DNA robots could have potential applications in autonomous chemical synthesis, in manufacturing responsive molecular devices, and in programmable therapeutics. The building blocks developed in this work could also be used for diverse functions other than cargo sorting. For example, inspired by ant foraging, adding a new building block for leaving pheromone-like signals on a path, DNA robots could be programmed to find the shortest path and efficiently transport cargo molecules. With simple communication between the robots, they could perform even more sophisticated tasks. With more effort in developing modular and collective molecular robots, and with simple and systematic approaches, molecular robots could eventually be easily programmed like macroscopic robots, but working in microscopic environments.
397 citations
References
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TL;DR: In this article, a modified Monte Carlo integration over configuration space is used to investigate the properties of a two-dimensional rigid-sphere system with a set of interacting individual molecules, and the results are compared to free volume equations of state and a four-term virial coefficient expansion.
Abstract: A general method, suitable for fast computing machines, for investigating such properties as equations of state for substances consisting of interacting individual molecules is described. The method consists of a modified Monte Carlo integration over configuration space. Results for the two‐dimensional rigid‐sphere system have been obtained on the Los Alamos MANIAC and are presented here. These results are compared to the free volume equation of state and to a four‐term virial coefficient expansion.
35,161 citations
"On the biophysics and kinetics of t..." refers methods in this paper
...This model is a continuous-time variant of the Metropolis (40) scheme for calculating thermodynamic averages....
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...We also considered the Kawasaki (3) method for assigning unimolecular transition rates in the IEL: kij = kuni × e−(∆Gbox(j)−∆Gbox(i))/2RT (8) kji = kuni × e−(∆Gbox(i)−∆Gbox(j))/2RT (9) As with the Metropolis rate model, this choice guarantees detailed balance and thus that the system eventually converges to a Boltzmann distribution: Prob(i) = 1 Q e− ∆Gbox(i) RT where Q = ∑ i e− ∆Gbox(i) RT ....
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...In the analysis below, the Metropolis method is assumed throughout....
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...All complexes were purified by nondenaturing 0 5 10 15 0 2 4 6 8 Toehold length lo g 1 0( k e ff) Dangles = Some Dangles = None Dangles = All Matching toehold Experiment Experiment A B 0 5 10 15 0 2 4 6 8 Toehold length Metropolis Kawasaki Figure S6: Multistrand simulations at 25 ◦C with different choices: (A) (i) in treating free energy contributions due to dangles (10) (options “Some”(default), “None” and “All” in the NUPACK (11) energy model (12)) and (ii) with substrate overhangs only as long as the toehold on the invader - i.e. matching length bottom toeholds on the substrate, rather than the full 15 base overhang used by Zhang and Winfree (1) (B) different ways of assigning absolute transition rates for unimolecular steps while satisfying detailed balance....
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...For each dangles option, kbi and kuni were separately calibrated to the same data (hybridization, zippering) used for calibrating the Metropolis method, as described in Schaeffer (9)....
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Book•
01 Jan 1995TL;DR: Detailed notes on Bayesian Computation Basics of Markov Chain Simulation, Regression Models, and Asymptotic Theorems are provided.
Abstract: FUNDAMENTALS OF BAYESIAN INFERENCE Probability and Inference Single-Parameter Models Introduction to Multiparameter Models Asymptotics and Connections to Non-Bayesian Approaches Hierarchical Models FUNDAMENTALS OF BAYESIAN DATA ANALYSIS Model Checking Evaluating, Comparing, and Expanding Models Modeling Accounting for Data Collection Decision Analysis ADVANCED COMPUTATION Introduction to Bayesian Computation Basics of Markov Chain Simulation Computationally Efficient Markov Chain Simulation Modal and Distributional Approximations REGRESSION MODELS Introduction to Regression Models Hierarchical Linear Models Generalized Linear Models Models for Robust Inference Models for Missing Data NONLINEAR AND NONPARAMETRIC MODELS Parametric Nonlinear Models Basic Function Models Gaussian Process Models Finite Mixture Models Dirichlet Process Models APPENDICES A: Standard Probability Distributions B: Outline of Proofs of Asymptotic Theorems C: Computation in R and Stan Bibliographic Notes and Exercises appear at the end of each chapter.
16,079 citations
TL;DR: The objective of this web server is to provide easy access to RNA and DNA folding and hybridization software to the scientific community at large by making use of universally available web GUIs (Graphical User Interfaces).
Abstract: The abbreviated name,‘mfold web server’,describes a number of closely related software applications available on the World Wide Web (WWW) for the prediction of the secondary structure of single stranded nucleic acids. The objective of this web server is to provide easy access to RNA and DNA folding and hybridization software to the scientific community at large. By making use of universally available web GUIs (Graphical User Interfaces),the server circumvents the problem of portability of this software. Detailed output,in the form of structure plots with or without reliability information,single strand frequency plots and ‘energy dot plots’, are available for the folding of single sequences. A variety of ‘bulk’ servers give less information,but in a shorter time and for up to hundreds of sequences at once. The portal for the mfold web server is http://www.bioinfo.rpi.edu/applications/ mfold. This URL will be referred to as ‘MFOLDROOT’.
12,535 citations
11,456 citations