Multidrug resistance transporter EmrE is only moderately sensitive to lipid composition.

Integral membrane proteins (IMPs) are fully embedded in the lipid bilayer and are often affected by their interactions with the surrounding lipids. Variation in the lipid environment can result in changes to hydrophobic matching, membrane lateral pressure, and electrostatic potentials, which can disrupt IMP folding and function. Here, we investigate the lipid-dependent behavior of the small multidrug resistance transporter EmrE. EmrE is an IMP native to E. coli and functions to expel toxic compounds from the bacterial cytoplasm. We know from previous studies that EmrE exhibits large variations in binding affinity, alternating access rate (the key step for transport of substrate across the membrane) and net transport rate depending on the transported substrate. Prior literature suggests that EmrE function may also be lipid dependent. In this study, we use NMR and isothermal titration calorimetry to study the impact of lipids on EmrE structure, substrate binding, and alternating access rate. We selected a set of lipids that vary in acyl chain saturation, acyl chain length, headgroup size, and charge. Our results show that EmrE retains highly similar structure, binding affinity, and alternating access rates across all lipid environments tested. Changes in binding enthalpy and entropy are observed in different lipids, but enthalpy-entropy compensation results in much smaller impacts on overall substrate affinity. Thus, despite its small size, key features of EmrE important for transport activity are only modestly perturbed by the lipid environment when substrate is present, with small-molecule substrates having a much greater impact on key steps in the EmrE transport cycle. Impacts of lipid on the transporter in the absence of small-molecule substrates may also affect EmrE transport activity, but are not examined here.