An Introduction to Passive Ion Transport across Model Lipid Membranes for Undergraduate Students: Proton Permeation Measurements in Liposomes

Abstract

In an upper-division biochemistry laboratory experiment, students use a variety of experimental techniques and apply several biochemical and biophysical concepts to measure and interpret proton permeation rates across model membranes. Model membranes in the form of liposomes are prepared from lipids by the extrusion method. The liposomes encapsulate the membrane-impermeable fluorescent pH-indicator pyranine, which facilitates pH detection in their aqueous interior. Rates of proton permeation across the lipid membrane in response to acidification of the external volume are measured by fluorescence spectroscopy. Data analysis introduces students to flux calculations and illustrates the concept that concentration differences are the driving force of diffusion processes such as passive ion transport across membranes. Students are given the task of formulating and then testing a hypothesis regarding the effects of bilayer thickness, liposome radius, magnitude of pH difference across the bilayer, and the presence of potential uncouplers on experimentally observable permeation rates.