a–c, Conformation space of the cryo-EM structures from this study and previously published ABC tranporter structures is shown based on three parameters: the intracellular gate angle (IC angle), the extracellular gate angle (EC angle), and the distance between two nucleotide-binding domains (NBD distance). The electron microscopy structures obtained in this study are shown as blue circles, and previously published structures are shown as orange triangles. The clustering of structures in three major conformations (IF, OFopen and OFoccluded) is highlighted by dashed rectangles in a and b. The cryo-EM structures obtained in this study cover all main conformations. The new URasym conformations show the closest resemblance to OFoccluded. The IC angle is defined as the angle between two vectors: vector 1 between the centre of mass of the whole extracellular part of the TMD region and the centre of mass of the intracellular parts of TM1TmrA, TM2TmrA, TM3TmrA, TM6TmrA, TM4TmrB and TM5TmrB; and vector 2 between the centre of mass of the whole extracellular part of the TMD region and the centre of mass of the intracellular parts of TM1TmrB, TM2TmrB, TM3TmrB, TM6TmrB, TM4TmrA and TM5TmrA. The EC angle is defined as the angle between two vectors: vector 3 between the centres of mass of the NBDs and the extracellular parts TM1TrmA, TM2TrmA, TM3TrmB, TM4TrmB, TM5TrmB and TM6TrmB; and vector 4 between the centre of mass of the NBDs and of the extracellular parts TM1TmrB, TM2TmrB, TM3TmrA, TM4TmrA, TM5TmrA and TM6TmrA. The NBD distance is between the centres of mass of two NBDs (N-terminal loops and C-terminal helices were excluded). IF, IF conformation; O, OFopen conformation; C, OFoccluded conformation; Vi, TmrAB ATP-Vi ; EQ, TmrAEQB. PDB IDs: PCAT, 4RY2 and 4S0F for the IF and OFoccluded conformations, respectively51; McjD, 4PL016; Pgp, 6C0V52; MsbA, 5TTP53; PglK, 5C7354; MRP, 6BHU55; Sav, 2ONJ18; TmrAB, 5MKK12; TM287 (TM287/288), 4Q4A56. d, Time-dependent IC angle in independent molecular dynamics simulations of IFwide and IFnarrow, with ATP/ATP, ADP/ATP and ADP/ADP in the canonical/non-canonical sites, respectively. Horizontal lines indicate the IC angles of the IFnarrow and IFwide structures.