Multidrug resistance-associated proteins are ABC C-family exporters. They are crucial in pharmacology as they transport various substrates across membranes. However, the role of the degenerate nucleotide-binding site (NBS) remains unclear likewise the interplay with the surrounding lipid environment. Here, we propose a dynamic and structural overview of MRP1 from ca. 110 μs molecular dynamics simulations. ATP binding to NBS1 is likely maintained along several transport cycles. Asymmetric NBD behaviour is ensured by lower signal transduction from NBD1 to the rest of the protein owing to the absence of ball-and-socket conformation between NBD1 and coupling helices. Even though surrounding lipids play an active role in the allosteric communication between the substrate-binding pocket and NBDs, our results suggest that lipid composition has a limited impact, mostly by affecting transport kinetics. We believe that our work can be extended to other degenerate NBS ABC proteins and provide hints for deciphering mechanistic differences among ABC transporters.

The human SLC22A6/OAT1 plays an important role in the elimination of a broad range of endogenous substances and xenobiotics thus attracting attention from the pharmacological community. Furthermore, OAT1 is also involved in key physiological events such as the remote inter-organ communication. Despite its significance, the knowledge about hOAT1 structure and the transport mechanism at the atomic level remains fragmented owing to the lack of resolved structures. By means of protein-threading modeling refined by μs-scaled Molecular Dynamics simulations, the present study provides the first robust model of hOAT1 in outward-facing conformation. Taking advantage of the AlphaFold 2 predicted structure of hOAT1 in inward-facing conformation, we here provide the essential structural and functional features comparing both states. The intracellular motifs conserved among Major Facilitator Superfamily members create a so-called “charge-relay system” that works as molecular switches modulating the conformation. The principal element of the event points at interactions of charged residues that appear crucial for the transporter dynamics and function. Moreover, hOAT1 model was embedded in different lipid bilayer membranes highlighting the crucial structural dependence on lipid-protein interactions. MD simulations supported the pivotal role of phosphatidylethanolamine components to the protein conformation stability. The present model is made available to decipher the impact of any observed polymorphism and mutation on drug transport as well as to understand substrate binding modes.

The human SLC22A6/OAT1 plays an important role in the disposition of a broad range of endogenous substances and xenobiotics. This is particularly important from the pharmacological point of view since OAT1 is involved in drug elimination events. Furthermore, OAT1 is also involved in key physiological events such as the remote inter-organ communication. Despite its significance, the knowledge about OAT1 structure and the transport mechanism at the atomic level remains fragmented owing to the lack of resolved structures. By means of protein-threading modeling refined by μs-scaled Molecular Dynamics simulations, the present study provides the first robust model of hOAT1 in outward-facing conformation. Taking advantage of the AlphaFold 2 predicted structure of hOAT1 in inward-facing conformation, we here provide the essential structural and functional features comparing both states. The intracellular motifs conserved among Major Facilitator Superfamily members create a so-called "charge-relay system" that works as molecular switches modulating the conformation. The principal element of the event points at interactions charged residues that appear crucial for the transporter dynamics and function. Besides, hOAT1 model was embedded in different lipid bilayer membranes highlighting the crucial structural dependence on lipid-protein. MD simulations supported the pivotal role of phosphatidylethanolamine (PE) components on the protein conformation stability. The present model is made available to decipher the impact of any observed polymorphism and mutation on drug transport as well as to understand substrate binding modes.