Abstract
Ion channels are membrane proteins that play important roles in a wide range of fundamental cellular processes. Studying membrane proteins at a molecular level becomes challenging in complex cellular environments. Instead, many studies focus on the isolation and reconstitution of the membrane proteins into model lipid membranes. Such simpler, in vitro, systems offer the advantage of control over the membrane and protein composition and the lipid environment. Rhodopsin and rhodopsin-like ion channels are widely studied due to their light-interacting properties and are a natural candidate for investigation with fluorescence methods. Here we review techniques for synthesizing liposomes and for reconstituting membrane proteins into lipid bilayers. We then summarize fluorescence assays which can be used to verify the functionality of reconstituted membrane proteins in synthetic liposomes.
Highlights
Membranes 2021, 11, 857. https://Ion channels provide a pathway for the movement of ions into and out of cells and organelles in all living organisms [1]
Other types of liposomes outside of unilamellar include multilamellar vesicles (MLVs) which are formed of multiple concentric bilayers, and multivesicular liposomes (MVLs) which are formed of multiple non-concentric bilayers or interconnected monolayers within a larger membrane vesicle (Figure 1C,D) [28]
The flow of the protons can be monitored by pH-sensitive fluorescent dyes as a function of time, and we describe three broad categories of pH-sensitive dyes: (1) hydrophobic membrane-permeable fluorescent pH-indicators, that can be added in the bulk solution of proteoliposomes, (2) water-soluble, membrane-impermeable pH-sensitive dyes, that can be encapsulated in proteoliposomes, and (3) lipids with a pH-sensitive head group, that can be included in the lipid mixture (Figure 4B)
Summary
Ion channels provide a pathway for the movement of ions into and out of cells and organelles in all living organisms [1] They are involved in many important cellular processes including membrane shaping and stabilization, immune response, and muscle contraction [1,2,3]. In vitro studies have been used to disclose various features of reconstituted ion channels in synthetic lipid bilayers [13,14]. We summarize recent approaches for membrane protein reconstitution into lipid bilayers and the limitations of each method. This results the formationof a group facing outwards andand their tails facing inwards This results inin the formation of hydrophobic a hydrophobic environment encased withing lipid membrane leaflet.
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