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Reconstitution of Ultrawide DNA Origami Pores in Liposomes for Transmembrane Transport of Macromolecules

[Image: see text] Molecular traffic across lipid membranes is a vital process in cell biology that involves specialized biological pores with a great variety of pore diameters, from fractions of a nanometer to >30 nm. Creating artificial membrane pores covering similar size and complexity will ai...

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Autores principales: Fragasso, Alessio, De Franceschi, Nicola, Stömmer, Pierre, van der Sluis, Eli O., Dietz, Hendrik, Dekker, Cees
Formato: Online Artículo Texto
Lenguaje:English
Publicado: American Chemical Society 2021
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8388114/
https://www.ncbi.nlm.nih.gov/pubmed/34170119
http://dx.doi.org/10.1021/acsnano.1c01669
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author Fragasso, Alessio
De Franceschi, Nicola
Stömmer, Pierre
van der Sluis, Eli O.
Dietz, Hendrik
Dekker, Cees
author_facet Fragasso, Alessio
De Franceschi, Nicola
Stömmer, Pierre
van der Sluis, Eli O.
Dietz, Hendrik
Dekker, Cees
author_sort Fragasso, Alessio
collection PubMed
description [Image: see text] Molecular traffic across lipid membranes is a vital process in cell biology that involves specialized biological pores with a great variety of pore diameters, from fractions of a nanometer to >30 nm. Creating artificial membrane pores covering similar size and complexity will aid the understanding of transmembrane molecular transport in cells, while artificial pores are also a necessary ingredient for synthetic cells. Here, we report the construction of DNA origami nanopores that have an inner diameter as large as 30 nm. We developed methods to successfully insert these ultrawide pores into the lipid membrane of giant unilamellar vesicles (GUVs) by administering the pores concomitantly with vesicle formation in an inverted-emulsion cDICE technique. The reconstituted pores permit the transmembrane diffusion of large macromolecules, such as folded proteins, which demonstrates the formation of large membrane-spanning open pores. The pores are size selective, as dextran molecules with a diameter up to 28 nm can traverse the pores, whereas larger dextran molecules are blocked. By FRAP measurements and modeling of the GFP influx rate, we find that up to hundreds of pores can be functionally reconstituted into a single GUV. Our technique bears great potential for applications across different fields from biomimetics, to synthetic biology, to drug delivery.
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spelling pubmed-83881142021-08-31 Reconstitution of Ultrawide DNA Origami Pores in Liposomes for Transmembrane Transport of Macromolecules Fragasso, Alessio De Franceschi, Nicola Stömmer, Pierre van der Sluis, Eli O. Dietz, Hendrik Dekker, Cees ACS Nano [Image: see text] Molecular traffic across lipid membranes is a vital process in cell biology that involves specialized biological pores with a great variety of pore diameters, from fractions of a nanometer to >30 nm. Creating artificial membrane pores covering similar size and complexity will aid the understanding of transmembrane molecular transport in cells, while artificial pores are also a necessary ingredient for synthetic cells. Here, we report the construction of DNA origami nanopores that have an inner diameter as large as 30 nm. We developed methods to successfully insert these ultrawide pores into the lipid membrane of giant unilamellar vesicles (GUVs) by administering the pores concomitantly with vesicle formation in an inverted-emulsion cDICE technique. The reconstituted pores permit the transmembrane diffusion of large macromolecules, such as folded proteins, which demonstrates the formation of large membrane-spanning open pores. The pores are size selective, as dextran molecules with a diameter up to 28 nm can traverse the pores, whereas larger dextran molecules are blocked. By FRAP measurements and modeling of the GFP influx rate, we find that up to hundreds of pores can be functionally reconstituted into a single GUV. Our technique bears great potential for applications across different fields from biomimetics, to synthetic biology, to drug delivery. American Chemical Society 2021-06-25 2021-08-24 /pmc/articles/PMC8388114/ /pubmed/34170119 http://dx.doi.org/10.1021/acsnano.1c01669 Text en © 2021 The Authors. Published by American Chemical Society https://creativecommons.org/licenses/by-nc-nd/4.0/Permits non-commercial access and re-use, provided that author attribution and integrity are maintained; but does not permit creation of adaptations or other derivative works (https://creativecommons.org/licenses/by-nc-nd/4.0/).
spellingShingle Fragasso, Alessio
De Franceschi, Nicola
Stömmer, Pierre
van der Sluis, Eli O.
Dietz, Hendrik
Dekker, Cees
Reconstitution of Ultrawide DNA Origami Pores in Liposomes for Transmembrane Transport of Macromolecules
title Reconstitution of Ultrawide DNA Origami Pores in Liposomes for Transmembrane Transport of Macromolecules
title_full Reconstitution of Ultrawide DNA Origami Pores in Liposomes for Transmembrane Transport of Macromolecules
title_fullStr Reconstitution of Ultrawide DNA Origami Pores in Liposomes for Transmembrane Transport of Macromolecules
title_full_unstemmed Reconstitution of Ultrawide DNA Origami Pores in Liposomes for Transmembrane Transport of Macromolecules
title_short Reconstitution of Ultrawide DNA Origami Pores in Liposomes for Transmembrane Transport of Macromolecules
title_sort reconstitution of ultrawide dna origami pores in liposomes for transmembrane transport of macromolecules
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8388114/
https://www.ncbi.nlm.nih.gov/pubmed/34170119
http://dx.doi.org/10.1021/acsnano.1c01669
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