Impact of anionic lipids on the energy landscape of conformational transition in anion exchanger 1 (AE1)

Anion Exchanger 1 (AE1) is an elevator-type transporter that plays a key role in acid-base homeostasis of erythrocytes. Here, we report three high-resolution cryo-electron microscopy (cryo-EM) structures of distinct states of AE1: two inward-facing (IF1 and IF2) and one outward-facing (OF). Uptake assay revealed the modulatory effect of phosphatidylinositol 4,5-bisphosphate (PIP₂) lipids on AE1. Molecular dynamics simulations are conducted on these structures to determine the anion binding sites in AE1. We then use advanced enhanced sampling to study the OF⇌IF transition in AE1 in three systems: apo, HCO₃^--bound, and an AE1 system in which cryo-EM-determined PIP₂ lipids had been removed. The transition pathways were then used to calculate the free energy of the OF⇌IF transition in AE1 under different conditions. The results show how substrate reduces the transition barrier against transport. Furthermore, they capture the inhibitory effect of PIP₂ lipids and provide a molecular mechanism for this inhibitory effect.