ArticleJournal of visualized experiments : JoVE2015
Reconstitution of a transmembrane protein, the voltage-gated ion channel, KvAP, into giant unilamellar vesicles for microscopy and patch clamp studies.
Article in Journal of visualized experiments : JoVE, 2015. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 27 papers.
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Who cites it
27 citing papers in PubMed.
- Bottom-up reconstitution of model membrane systems: a mini-review.Biochemical Society transactions · 2026Review
- Reconstitution of Active Plant HBio-protocol · 2026Article
- Giant unilamellar vesicles as a model system for studying ion transport.Biophysical reviews · 2025Review
- Functional reconstitution of plant plasma membrane HScientific reports · 2025Article
- Calcium ions promote migrasome formation via Synaptotagmin-1.The Journal of cell biology · 2024Article
- Protein-Rich Rafts in Hybrid Polymer/Lipid Giant Unilamellar Vesicles.Biomacromolecules · 2024Article
- Formation of Cell-Sized Liposomes Incorporating a β-Barrel-Structured Porin through Rehydration of a Phospholipid-Membrane Protein Dried Film.ACS omega · 2024Article
- Secondary Ion Mass Spectrometry of Single Giant Unilamellar Vesicles Reveals Compositional Variability.Journal of the American Chemical Society · 2023Article
- Review
- Thermoplasmonic Vesicle Fusion Reveals Membrane Phase Segregation of Influenza Spike Proteins.Nano letters · 2023Article
- Minimal Out-of-Equilibrium Metabolism for Synthetic Cells: A Membrane Perspective.ACS synthetic biology · 2023Review
- A simple method to make, trap and deform a vesicle in a gel.Scientific reports · 2023Article
- Experimental platform for the functional investigation of membrane proteins in giant unilamellar vesicles.Soft matter · 2022Article
- A Fluorescence-based Assay for Measuring Phospholipid Scramblase Activity in Giant Unilamellar Vesicles.Bio-protocol · 2022Article
- Article
- Towards a synthetic cell cycle.Nature communications · 2021Review
- Endoplasmic reticulum phospholipid scramblase activity revealed after protein reconstitution into giant unilamellar vesicles containing a photostable lipid reporter.Scientific reports · 2021Article
- Fusion Pore Formation Observed during SNARE-Mediated Vesicle Fusion with Pore-Spanning Membranes.Biophysical journal · 2020Article
- Physicochemical considerations for bottom-up synthetic biology.Emerging topics in life sciences · 2019Article
- Combining patch-clamping and fluorescence microscopy for quantitative reconstitution of cellular membrane processes with Giant Suspended Bilayers.Scientific reports · 2019Article
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Authors and funding
4 authors.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Giant Unilamellar Vesicles (GUVs) are a popular biomimetic system for studying membrane associated phenomena. However, commonly used protocols to grow GUVs must be modified in order to form GUVs containing functional transmembrane proteins. This article describes two dehydration-rehydration methods - electroformation and gel-assisted swelling - to form GUVs containing the voltage-gated potassium channel, KvAP. In both methods, a solution of protein-containing small unilamellar vesicles is partially dehydrated to form a stack of membranes, which is then allowed to swell in a rehydration buffer. For the electroformation method, the film is deposited on platinum electrodes so that an AC field can be applied during film rehydration. In contrast, the gel-assisted swelling method uses an agarose gel substrate to enhance film rehydration. Both methods can produce GUVs in low (e.g., 5 mM) and physiological (e.g., 100 mM) salt concentrations. The resulting GUVs are characterized via fluorescence microscopy, and the function of reconstituted channels measured using the inside-out patch-clamp configuration. While swelling in the presence of an alternating electric field (electroformation) gives a high yield of defect-free GUVs, the gel-assisted swelling method produces a more homogeneous protein distribution and requires no special equipment.
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