Membrane activity of a DNA-based ion channel depends on the stability of its double-stranded structure.

Structural DNA nanotechnology has emerged as a promising method for designing spontaneously-inserting and fully-controllable synthetic ion channels. However, both insertion efficiency and stability of existing DNA-based ion channels leave much room for improvement. Here, we demonstrate an approach to overcoming the unfavorable DNA-lipid interactions that hinder the formation of a stable transmembrane pore. Our all-atom MD simulations and experiments show that the insertion-driving cholesterol modifications, when introduced at an end of a DNA strand, are likely to cause fraying of the terminal base pairs as the DNA nanostructure adopts its energy-minimum configuration in the membrane. We also find that fraying of base pairs distorts nicked DNA constructs when embedded in a lipid bilayer. Here, we show that DNA nanostructures that do not have discontinuities (nicks) in their DNA backbones form considerably more stable DNA-induced conductive pores and insert into lipid membranes with a higher efficiency than the equivalent nicked constructs. Moreover, lack of nicks allows to design and maintain membrane-spanning helices in a tilted orientation within lipid bilayer. Thus, reducing the conformational degrees of freedom of the DNA nanostructures enables better control over their function as synthetic ion channels.

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Choice of fluorophore affects dynamic DNA nanostructures

10.1101/2020.12.12.422444 ◽

2020 ◽

Author(s):

Kevin Jahnke ◽

Helmut Grubmüller ◽

Maxim Igaev ◽

Kerstin Göpfrich

Keyword(s):

Energy Landscape ◽

Dna Nanotechnology ◽

Reaction Diffusion ◽

Md Simulations ◽

Dna Origami ◽

Dna Nanostructures ◽

Diffusion Modeling ◽

Ph Responsive ◽

Dna Nanostructure ◽

Potential Impact

The ability to dynamically remodel DNA origami structures or functional nanodevices is highly desired in the field of DNA nanotechnology. Concomitantly, the use of fluorophores to track and validate the dynamics of such DNA-based architectures is commonplace and often unavoidable. It is therefore crucial to be aware of the side effects of popular fluorophores, which are often exchanged without considering the potential impact on the system. Here, we show that the choice of fluorophore can strongly affect the reconfiguration of DNA nanostructures. To this end, we encapsulate a triple-stranded DNA (tsDNA) into water-in-oil compartments and functionalize their periphery with a single-stranded DNA handle (ssDNA). Thus, the tsDNA can bind and unbind from the periphery by reversible opening of the triplex and subsequent strand displacement. Using a combination of experiments, molecular dynamics (MD) simulations, and reaction-diffusion modeling, we demonstrate for twelve different fluorophore combinations that it is possible to alter or even inhibit the DNA nanostructure formation - without changing the DNA sequence. Besides its immediate importance for the design of pH-responsive switches and fluorophore labelling, our work presents a strategy to precisely tune the energy landscape of dynamic DNA nanodevices.

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Theoretical and computational modeling of peptide-lipid bilayer interaction studied by dynamic force spectroscopy

10.32469/10355/76254 ◽

2019 ◽

Author(s):

◽

Milica Utjesanovic

Keyword(s):

Computational Modeling ◽

Lipid Bilayer ◽

Lipid Bilayers ◽

Lipid Membranes ◽

Lipid Membrane ◽

Conformational Dynamics ◽

Quantitative Description ◽

Md Simulations ◽

Dynamic Force ◽

Detachment Rate

This thesis consists of three interrelated theoretical and computational modeling projects that investigate different aspects of peptide-lipid membrane interactions. (1) A general theoretical approach is formulated for the quantitative description of the detachment force distribution, P(F), and the corresponding force dependent detachment rate, k(F), of a peptide from a lipid bilayer, by assuming that peptide detachment from lipid membranes occurs stochastically along a few dominant diffusive pathways. Besides providing a consistent interpretation of the experimental data, the new method also predicts that k(F) exhibits catch-bond behavior (when, counter intuitively, the detachment rate decreases with increasing force). (2) The proposed multiple detachment pathways method is tested and validated for a particular peptide (SecA2-11) interacting with both zwitterionic POPC lipid and polar E. Coli membranes. Furthermore, molecular dynamics (MD) simulations are used to explored the conformational dynamics of SecA2-11 during its interaction with both POPC and anionic POPG lipid bilayers. (3) Finally, MD simulations are used to explore the conformational dynamics and energetics of the peptide melittin (MWT) and its diastereomer (MD4) interacting with POPC and POPG lipid bilayers. The obtained results provide further insight into the role of secondary structure in peptide-lipid bilayer interactions.

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Determinants of ligand-functionalized DNA nanostructure-cell interactions

10.1101/2021.02.24.432702 ◽

2021 ◽

Author(s):

Glenn A.O. Cremers ◽

Bas J.H.M. Rosier ◽

Ab Meijs ◽

Nicholas B. Tito ◽

Sander M.J. van Duijnhoven ◽

...

Keyword(s):

Three Dimensional ◽

Dna Nanotechnology ◽

Absolute Number ◽

Design Parameters ◽

Dna Nanostructures ◽

Surface Receptors ◽

Protein Ligands ◽

Dna Nanostructure ◽

Functionalized Nanomaterials ◽

Ligand Orientation

AbstractSynthesis of ligand-functionalized nanomaterials with control over size, shape and ligand orientation, facilitates the design of tailored nanomedicines for therapeutic purposes. DNA nanotechnology has emerged as a powerful tool to rationally construct two- and three-dimensional nanostructures, enabling site-specific incorporation of protein ligands with control over stoichiometry and orientation. To efficiently target cell surface receptors, exploration of the parameters that modulate cellular accessibility of these nanostructures is essential. In this study we systematically investigate tunable design parameters of antibody-functionalized DNA nanostructures binding to therapeutically relevant receptors. We show that, although the native affinity of antibody-functionalized DNA nanostructures remains unaltered, the absolute number of bound surface receptors is lower compared to soluble antibodies and is mainly governed by nanostructure size and DNA handle location. The obtained results provide key insights in the ability of ligand-functionalized DNA nanostructures to bind surface receptors and yields design rules for optimal cellular targeting.

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DNA nanotechnology-empowered nanoscopic imaging of biomolecules

Chemical Society Reviews ◽

10.1039/d0cs01281e ◽

2021 ◽

Author(s):

Fan Li ◽

Jiang Li ◽

Baijun Dong ◽

Fei Wang ◽

Chunhai Fan ◽

...

Keyword(s):

Dna Nanotechnology ◽

Dna Nanostructures ◽

Dna Nanostructure ◽

Imaging Probes

DNA nanotechnology has led to the rise of DNA nanostructures, which possess programmable shapes and are capable of organizing different functional molecules and materials. A variety of DNA nanostructure-based imaging probes have been developed.

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Synthetic Ion Channels via Self-Assembly: A Route for Embedding Porous Polyoxometalate Nanocapsules in Lipid Bilayer Membranes

Nano Letters ◽

10.1021/nl802366k ◽

2008 ◽

Vol 8 (11) ◽

pp. 3916-3921 ◽

Cited By ~ 37

Author(s):

Rogan Carr ◽

Ira A. Weinstock ◽

Asipu Sivaprasadarao ◽

Achim Müller ◽

Aleksei Aksimentiev

Keyword(s):

Ion Channels ◽

Lipid Bilayer ◽

Self Assembly ◽

Lipid Bilayer Membranes ◽

Bilayer Membranes ◽

Synthetic Ion Channels

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Adenita: interactive 3D modelling and visualization of DNA nanostructures

Nucleic Acids Research ◽

10.1093/nar/gkaa593 ◽

2020 ◽

Vol 48 (15) ◽

pp. 8269-8275 ◽

Cited By ~ 3

Author(s):

Elisa de Llano ◽

Haichao Miao ◽

Yasaman Ahmadi ◽

Amanda J Wilson ◽

Morgan Beeby ◽

...

Keyword(s):

Dna Nanotechnology ◽

Software Tool ◽

Building Blocks ◽

Dna Origami ◽

Dna Nanostructures ◽

User Interactions ◽

Dna Nanostructure ◽

Dna Tiles ◽

The Creation ◽

Interactive 3D

Abstract DNA nanotechnology is a rapidly advancing field, which increasingly attracts interest in many different disciplines, such as medicine, biotechnology, physics and biocomputing. The increasing complexity of novel applications requires significant computational support for the design, modelling and analysis of DNA nanostructures. However, current in silico design tools have not been developed in view of these new applications and their requirements. Here, we present Adenita, a novel software tool for the modelling of DNA nanostructures in a user-friendly environment. A data model supporting different DNA nanostructure concepts (multilayer DNA origami, wireframe DNA origami, DNA tiles etc.) has been developed allowing the creation of new and the import of existing DNA nanostructures. In addition, the nanostructures can be modified and analysed on-the-fly using an intuitive toolset. The possibility to combine and re-use existing nanostructures as building blocks for the creation of new superstructures, the integration of alternative molecules (e.g. proteins, aptamers) during the design process, and the export option for oxDNA simulations are outstanding features of Adenita, which spearheads a new generation of DNA nanostructure modelling software. We showcase Adenita by re-using a large nanorod to create a new nanostructure through user interactions that employ different editors to modify the original nanorod.

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Molecular Dynamics Simulation of the Interaction Between Benzanthrone Dye and Model Lipid Membranes

East European Journal of Physics ◽

10.26565/2312-4334-2020-3-17 ◽

2020 ◽

Keyword(s):

Molecular Dynamics ◽

Molecular Dynamics Simulation ◽

Lipid Bilayer ◽

Lipid Bilayers ◽

Lipid Membranes ◽

Fluorescent Dyes ◽

Md Simulations ◽

Dynamics Simulation ◽

Membrane Composition ◽

Model Lipid Membranes

The benzanthrone fluorescent dyes are known as environmentally-sensitive reporters for exploring the physicochemical properties and structural alterations of lipid membranes. In the present work the 100-ns molecular dynamics simulation (MD) was used to characterize the bilayer location and the nature of interactions between the benzanthrone fluorescent dye ABM and the model lipid membranes composed of the zwitterionic lipid phosphatidylcholine (PC) and its mixtures with the anionic lipid phosphatidylglycerol (PG20) and sterol cholesterol (Chol30). The MD simulations were performed in the CHARMM36m force field using the GROMACS package. The ABM molecule, which was initially placed at a distance of 30 Å from the midplane of the lipid bilayer, after 10 ns of simulation was found to be completely incorporated into the membrane interior and remained within the lipid bilayer for the rest of the simulation time. The analysis of the MD simulation results showed that the lipid bilayer location of the benzanthrone dye ABM depends on the membrane composition, with the distance from bilayer center being gradually shifted from 0.78 nm in the neat PC bilayer to 0.95 nm and 1.5 nm in the PG- and Chol-containing membranes, respectively. In addition, the partitioning of the ABM into the neat PC bilayer was followed by the probe translocation from the outer membrane leaflet to the inner one. A separate series of MD simulations was aimed at examining the ABM influence on the lipid bilayer structure. It was found that ABM partitioning into the lipid bilayers of various composition has no significant effect on the orientation of the fatty acid chains and leads only to a small increase of the deuterium order parameter for the carbon atoms 5-to-8 in the sn-2 acyl chains of the neat PC membranes. In addition, the interaction of the ABM with the model lipid membranes caused the slight decrease of the surface area per lipid pointing to the slight increase of the packing density of lipid molecules in the presence of ABM. The results obtained provide a basis for deeper understanding of the membrane interactions of benzanthrone dyes and may be useful for the design of the novel fluorescent probes for membrane studies.

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Ultrasound exposure enhances cellular uptake of nanoscale cargos

10.1101/2021.09.11.459898 ◽

2021 ◽

Author(s):

Kahkashan Bansal ◽

Anjali Rajwar ◽

Himanshu Shekhar ◽

Dhiraj Bhatia

Keyword(s):

Drug Delivery ◽

Cellular Uptake ◽

Propidium Iodide ◽

Dna Nanotechnology ◽

Chemotherapeutic Agents ◽

Dna Nanostructures ◽

Ultrasound Exposure ◽

Dna Nanostructure ◽

Cellular Components ◽

Further Development

DNA nanotechnology utilizes DNA as a structural molecule to design palette of nanostructures with different shapes and sizes. DNA nanocages have demonstrated significant potential for drug delivery. Therefore, enhancing the delivery of DNA nanocages into cells can improve their efficacy as drug delivery agents. Numerous studies have reported the effects of ultrasound for enhancing drug delivery across biological barriers. The mechanical bioeffects caused by cell-ultrasound interaction can cause sonoporation, leading to enhanced uptake of drugs, nanoparticles, and chemotherapeutic agents through membranes. Whether ultrasound exposure can enhance the delivery of DNA nanocages has not been explored, which is the focus of this study. Specifically, we investigated the effects of ultrasound on the cellular uptake of propidium Iodide, fluorescent dextrans, and DNA nanostructures). We provide evidence of modulation of pore formation in the cell membrane by ultrasound by studying the intracellular uptake of the impermeable dye, propidium iodide. Treatment of cells with low amplitudes of ultrasound enhanced the uptake of different sizes of dextrans and DNA based nanodevices. These findings could serve as the foundation for further development ultrasound-enabled DNA nanostructure delivery and for specific understanding of underlying biological mechanisms of interaction between ultrasound parameters and cellular components; the knowledge that can be further explored for potential biological and biomedical applications.

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In vivo cloning of artificial DNA nanostructures

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.0805416105 ◽

2008 ◽

Vol 105 (46) ◽

pp. 17626-17631 ◽

Cited By ~ 81

Author(s):

Chenxiang Lin ◽

Sherri Rinker ◽

Xing Wang ◽

Yan Liu ◽

Nadrian C. Seeman ◽

...

Keyword(s):

Dna Nanotechnology ◽

Dna Nanostructures ◽

Scientific Enterprise ◽

Dna Nanostructure ◽

Copy Numbers ◽

Artificial Dna ◽

Difficult Challenge ◽

Endonuclease Vii

Mimicking nature is both a key goal and a difficult challenge for the scientific enterprise. DNA, well known as the genetic-information carrier in nature, can be replicated efficiently in living cells. Today, despite the dramatic evolution of DNA nanotechnology, a versatile method that replicates artificial DNA nanostructures with complex secondary structures remains an appealing target. Previous success in replicating DNA nanostructures enzymatically in vitro suggests that a possible solution could be cloning these nanostructures by using viruses. Here, we report a system where a single-stranded DNA nanostructure (Holliday junction or paranemic cross-over DNA) is inserted into a phagemid, transformed into XL1-Blue cells and amplified in vivo in the presence of helper phages. High copy numbers of cloned nanostructures can be obtained readily by using standard molecular biology techniques. Correct replication is verified by a number of assays including nondenaturing PAGE, Ferguson analysis, endonuclease VII digestion, and hydroxyl radical autofootprinting. The simplicity, efficiency, and fidelity of nature are fully reflected in this system. UV-induced psoralen cross-linking is used to probe the secondary structure of the inserted junction in infected cells. Our data suggest the possible formation of the immobile four-arm junction in vivo.

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Controlling aggregation of cholesterol-modified DNA nanostructures

Nucleic Acids Research ◽

10.1093/nar/gkz914 ◽

2019 ◽

Vol 47 (21) ◽

pp. 11441-11451 ◽

Cited By ~ 7

Author(s):

Alexander Ohmann ◽

Kerstin Göpfrich ◽

Himanshu Joshi ◽

Rebecca F Thompson ◽

Diana Sobota ◽

...

Keyword(s):

Molecular Dynamics ◽

Molecular Dynamics Simulations ◽

Lipid Membranes ◽

Structural Control ◽

Polyacrylamide Gel Electrophoresis ◽

Dna Nanotechnology ◽

Dna Nanostructures ◽

Hydrophobic Moiety ◽

Modified Dna ◽

Dynamics Simulations

Abstract DNA nanotechnology allows for the design of programmable DNA-built nanodevices which controllably interact with biological membranes and even mimic the function of natural membrane proteins. Hydrophobic modifications, covalently linked to the DNA, are essential for targeted interfacing of DNA nanostructures with lipid membranes. However, these hydrophobic tags typically induce undesired aggregation eliminating structural control, the primary advantage of DNA nanotechnology. Here, we study the aggregation of cholesterol-modified DNA nanostructures using a combined approach of non-denaturing polyacrylamide gel electrophoresis, dynamic light scattering, confocal microscopy and atomistic molecular dynamics simulations. We show that the aggregation of cholesterol-tagged ssDNA is sequence-dependent, while for assembled DNA constructs, the number and position of the cholesterol tags are the dominating factors. Molecular dynamics simulations of cholesterol-modified ssDNA reveal that the nucleotides wrap around the hydrophobic moiety, shielding it from the environment. Utilizing this behavior, we demonstrate experimentally that the aggregation of cholesterol-modified DNA nanostructures can be controlled by the length of ssDNA overhangs positioned adjacent to the cholesterol. Our easy-to-implement method for tuning cholesterol-mediated aggregation allows for increased control and a closer structure–function relationship of membrane-interfacing DNA constructs — a fundamental prerequisite for employing DNA nanodevices in research and biomedicine.

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