Detecting Single‐Molecule Dynamics on Lipid Membranes with Quenchers‐in‐a‐Liposome FRET

Ma, Dawei; Xu, Chong; Hou, Wen-Qing; Zhao, Chunyu; Ma, Jianbing; Huang, Xing-Yuan; Jia, Qi; Ma, Lu; Diao, Jiajie; Liu, Cong; Li, Ming; Lü, Ying

doi:10.1002/ange.201813888

Cited by 9 publications

(3 citation statements)

References 44 publications

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“…For small DNA motifs, dsDNA bound more efficiently to liposomes than ssDNA, and the addition of a 6-nt ssDNA overhang had minimal effect. Membrane-bound ssDNA has been observed to lie close to the surface of lipid bilayer membranes in fluorescence studies ( 79 ), while dsDNA remains in a stable position protruding normal to the membrane surface ( 80 ). Thus, the orientation difference of dsDNA compared to ssDNA may result in improved binding.…”

Section: Discussionmentioning

confidence: 99%

Binding of DNA origami to lipids: maximizing yield and switching via strand displacement

Singh

Darley

Ridone

et al. 2021

Nucleic Acids Research

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Liposomes are widely used as synthetic analogues of cell membranes and for drug delivery. Lipid-binding DNA nanostructures can modify the shape, porosity and reactivity of liposomes, mediated by cholesterol modifications. DNA nanostructures can also be designed to switch conformations by DNA strand displacement. However, the optimal conditions to facilitate stable, high-yield DNA–lipid binding while allowing controlled switching by strand displacement are not known. Here, we characterized the effect of cholesterol arrangement, DNA structure, buffer and lipid composition on DNA–lipid binding and strand displacement. We observed that binding was inhibited below pH 4, and above 200 mM NaCl or 40 mM MgCl2, was independent of lipid type, and increased with membrane cholesterol content. For simple motifs, binding yield was slightly higher for double-stranded DNA than single-stranded DNA. For larger DNA origami tiles, four to eight cholesterol modifications were optimal, while edge positions and longer spacers increased yield of lipid binding. Strand displacement achieved controlled removal of DNA tiles from membranes, but was inhibited by overhang domains, which are used to prevent cholesterol aggregation. These findings provide design guidelines for integrating strand displacement switching with lipid-binding DNA nanostructures. This paves the way for achieving dynamic control of membrane morphology, enabling broader applications in nanomedicine and biophysics.

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Section: Discussionmentioning

confidence: 99%

Binding of DNA origami to lipids: maximizing yield and switching via strand displacement

Singh

Darley

Ridone

et al. 2021

Nucleic Acids Research

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“…Based on the above results, the mapping technique enables real-time tracking of a single liposome, which is significant for understanding the transport process of liposomes as nanocarriers. − To study the surface chemistry dependence of single liposome thermodynamics, we mapped the real-time localization on substrates with different modifications and electrical charges. First, we measured the average diameter of liposomes as 70 nm by DLS, and the zeta potential measurement shows that the liposome surface carries positive charges (Supporting Information, Figure S11).…”

Section: Resultsmentioning

confidence: 99%

Imaging the Heterogeneous Localization of a Single Molecule

et al. 2021

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Single-molecule localization allows determining the underlying biological and biochemical processes and promotes the development of super-resolution imaging techniques. Here, we present an optical technique of tracking the motion of a single nanoparticle linked to a substrate via a biomolecule tether to reveal the localization of single biomolecules and the transient states of single nanoparticle switching between specific binding pairs. The affinities, steric hindrance, and conformational variation of a singlemolecule binding pair uncover the dynamic details and intrinsic mechanism of binding processes with high specificity and accuracy (a few nanometers). The application of tracking motions of single soft liposomes on different modified surfaces was further demonstrated, which revealed the characteristic behavior related to surface chemistry. Our results show that the trajectory of nanoscale liposomes loaded with small-drug molecules is linked to the compositional inhomogeneity, which provides a route for thorough comprehension of the fundamental biotechnological process.

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“…13 We recently developed an in vitro fluorescence method to measure membrane protein dynamics in artificial lipid bilayers. 14 The method enabled us to extract information about the transmembrane motions of biomolecules while disregarding their in-plane motions. However, complex feedback networks and/or molecular crowding in cells may alter the functions and dynamics of biomolecules, potentially posing obstacles to the physiological relevance of information gained from in vitro systems.…”

mentioning

confidence: 99%

Subnanometer-Precision Measurements of Transmembrane Motions of Biomolecules in Plasma Membranes Using Quenchers in Extracellular Environment

Hou

et al. 2020

Nano Lett.

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It is a big challenge to measure position changes of biomolecules in the direction normal to the plasma membranes of living cells. We developed a one donor-multiple quenchers Fӧrster resonance energy transfer method by using non-fluorescent quenchers in the extracellular environment. It senses subnanometer position changes of a fluorophore-labeled biomolecule in the plasma membrane. The method was validated by monitoring flip-flops of individual lipid molecules incorporated in plasma membranes. We studies membrane perforation by a host defense peptide from the extracellular side and found that the pore-forming peptide is dynamic, switching among different insertion depths. The method is especially useful in studying interactions of membrane proteins with the inner surfaces of plasma membranes. Our method will find wide applications in systematic analysis of fundamental cellular processes at plasma membranes.

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Detecting Single‐Molecule Dynamics on Lipid Membranes with Quenchers‐in‐a‐Liposome FRET

Cited by 9 publications

References 44 publications

Binding of DNA origami to lipids: maximizing yield and switching via strand displacement

Binding of DNA origami to lipids: maximizing yield and switching via strand displacement

Imaging the Heterogeneous Localization of a Single Molecule

Subnanometer-Precision Measurements of Transmembrane Motions of Biomolecules in Plasma Membranes Using Quenchers in Extracellular Environment

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