1. DNA-Based Optical Quantification of Ion Transport across Giant Vesicles
- Author
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Fletcher, Marcus, Zhu, Jinbo, Rubio-Sánchez, Roger, Sandler, Sarah E, Nahas, Kareem Al, Michele, Lorenzo Di, Keyser, Ulrich F, Tivony, Ran, Fletcher, Marcus [0000-0003-3036-1168], Zhu, Jinbo [0000-0003-4935-825X], Rubio-Sánchez, Roger [0000-0001-5574-5809], Sandler, Sarah E [0000-0001-9689-8684], Nahas, Kareem Al [0000-0003-4568-5894], Michele, Lorenzo Di [0000-0002-1458-9747], Keyser, Ulrich F [0000-0003-3188-5414], Tivony, Ran [0000-0003-0331-9538], and Apollo - University of Cambridge Repository
- Subjects
Ions ,Octanols ,Ion Transport ,G-quadruplex ,Lipid Bilayers ,giant unilamellar vesicles ,microfluidics ,General Engineering ,Gramicidin ,General Physics and Astronomy ,DNA ,ion sensor ,Ion Channels ,Potassium ,General Materials Science ,Protons ,Unilamellar Liposomes - Abstract
Accurate measurements of ion permeability through cellular membranes remains challenging due to the lack of suitable ion-selective probes. Here we use giant unilamellar vesicles (GUVs) as membrane models for the direct visualization of mass translocation at the single-vesicle level. Ion transport is indicated with a fluorescently adjustable DNA-based sensor that accurately detects sub-millimolar variations in K+ concentration. In combination with microfluidics, we employed our DNA-based K+ sensor for extraction of the permeation coefficient of potassium ions. We measured K+ permeability coefficients at least 1 order of magnitude larger than previously reported values from bulk experiments and show that permeation rates across the lipid bilayer increase in the presence of octanol. In addition, an analysis of the K+ flux in different concentration gradients allows us to estimate the complementary H+ flux that dissipates the charge imbalance across the GUV membrane. Subsequently, we show that our sensor can quantify the K+ transport across prototypical cation-selective ion channels, gramicidin A and OmpF, revealing their relative H+/K+ selectivity. Our results show that gramicidin A is much more selective to protons than OmpF with a H+/K+ permeability ratio of ∼104.
- Published
- 2022