Molecular dynamics simulations of lipid-protein interactions in SLC4 proteins

The SLC4 family of secondary bicarbonate transporters is responsible for the transport of HCO₃^-, CO₃^2-, Cl^-, Na⁺, K⁺, NH₃, and H⁺, which are necessary for regulation of pH and ion homeostasis. They are widely expressed in numerous tissues throughout the body and function in different cell types with different membrane properties. Potential lipid roles in SLC4 function have been reported in experimental studies, focusing mostly on two members of the family: AE1 (Cl^-/HCO₃^- exchanger) and NBCe1 (Na⁺-CO₃^2-cotransporter). Previous computational studies of the outward-facing state of AE1 with model lipid membranes revealed enhanced protein-lipid interactions between cholesterol (CHOL) and phosphatidylinositol bisphosphate (PIP2). However, the protein-lipid interactions in other members of the family and other conformation states are still poorly understood and this precludes the detailed studies of a potential regulatory role for lipids in the SLC4 family. In this work, we performed coarse-grained and atomistic molecular dynamics simulations on three members of the SLC4 family with different transport modes: AE1, NBCe1, and NDCBE (an Na⁺-CO₃^2-/Cl^- exchanger), in model HEK293 membranes consisting of CHOL, PIP2, phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, and sphingomyelin. The recently resolved inward-facing state of AE1 was also included in the simulations. Lipid-protein contact analysis of the simulated trajectories was performed with the ProLint server, which provides a multitude of visualization tools for illustration of areas of enhanced lipid-protein contact and identification of putative lipid binding sites within the protein matrix. We observed enrichment of CHOL and PIP2 around all proteins with subtle differences in their distribution depending on the protein type and conformation state. Putative binding sites were identified for CHOL, PIP2, phosphatidylcholine, and sphingomyelin in the three studied proteins, and their potential roles in the SLC4 transport function, conformational transition, and protein dimerization are discussed.