Tic20 insertion into liposomes and channel formation. (A) Flotation experiments of Tic20-proteoliposomes and Tic20 without vesicles in a sucrose gradient. Samples containing 1.6 M sucrose were loaded at the bottom of a sucrose step gradient and centrifuged to equilibrium (100,000 g, 19 h, 4°C). Fractions were analysed by silver-staining. (B) Flotation experiments of Tic20-proteoliposomes (similar to (A)) incubated under the indicated buffer conditions for 30 min at 4°C before centrifugation. (C) Swelling assay of liposomes, Tic20-proteoliposomes and NtTic110-proteoliposomes containing 20 mM Tris-HCl (pH 8.0), 100 mM NaCl. Change in optical density was measured at 500 nm (OD500 nm) of 1 ml solutions every minute. Arrow indicates the addition of 300 mM KCl. Presented results are the average of at least five repetitions; standard deviations were within 1.5-3%.