van der Heide et al. studied activation of the ABC transporter OpuA, which allows bacteria to accumulate glycine betaine in response to increased osmotic conditions, in reconstituted liposome preparations under varying osmotic conditions. Activation of the transporter by an osmotic gradient was dependent on the percentage of anionic lipids in the reconstituted liposomes, and in liposomes without any anionic lipids, the transporter failed to be activated. Lipids can form multiple structures depending on their biochemical properties, and maximal activation of OpuA depended on the presence of lipids that did not form bilayers. However, OpuA activity was relatively insensitive to changes in the length of the acyl chains, which would alter the membrane thickness of the liposomes. Inclusion of ions in the lumen of the vesicles to create a situation of high ionic strength stimulated OpuA activity even in the absence of an osmotic gradient. Thus, the authors propose that increased osmotic conditions alter the cytoplasmic ionic strength and alter the interaction between the transporter and the anionic phospholipid head groups, leading to transporter activation.