Catalytic activity of MsbA reconstituted in nanodisc particles is modulated by remote interactions with the bilayer

ATP-binding cassette (ABC) transporters couple hydrolysis of ATP with vectorial transport across the cell membrane. We have reconstituted ABC transporter MsbA in nanodiscs of various sizes and lipid compositions to test whether ATPase activity is modulated by the properties of the bilayer. ATP hydrolysis rates, Michaelis–Menten parameters, and dissociation constants of substrate analog ATP-γ-S demonstrated that physicochemical properties of the bilayer modulated binding and ATPase activity. This is remarkable when considering that the catalytic unit is located ∼50 Å from the transmembrane region. Our results validated the use of nanodiscs as an effective tool to reconstitute MsbA in an active catalytic state, and highlighted the close relationship between otherwise distant transmembrane and ATPase modules.

► MsbA is a cell transporter with ATPase and transmembrane domains separated by ∼50 Å. ► MsbA was successfully reconstituted in nanodiscs in a functional state. ► Catalytic domain is sensitive to physical properties of bilayer. ► ATPase and transmembrane domains are physically distant, but functionally linked.