Membrane Lateral Pressure Regulates Dipolar Relaxation Dynamics at the Active Site of an ATPase

Copper is an essential cofactor for redox enzymes but toxic when it accumulates in the cell. Copper homeostasis relies on transmembrane Cu+ transporting PIB-type ATPases. Molecular Dynamics simulations of the Cu-ATPase CopA from Legionella pneumophila[1] (LpCopA) suggest a water-accessible Cu+-binding site at the conserved CPC motif in the E2.Pi intermediate, where Cu+ occlusion is expected from homology with the Ca2+-ATPase, SERCAII[2,3]. However, the role of the additional physical constraints imposed by the biological membranes on LpCopA is not known. We have labeled the transmembrane cysteines of the copper-binding site with the polarity-sensitive fluorophore 6-bromoacetyl-2-dimethylaminonaphthalene (BADAN). Time-resolved dipolar relaxation studies of the dye show that membrane lateral pressure affects the hydration and dipole mobility of the ion binding site with high topological discrimination. The data show that the lipidic phase can contribute to the energetics of the ion transport cycle by providing lateral forces that affect hydration and dehydration events.

References
[1] P. Gourdon, P. et al. 2011, Nature 475, 59–64
[2] M. Andersson et al. 2014, Nature Struct.&Mol.Biol. 21, 43-48
[3] J. Møller et al. 2010, Biophys.Q. Rev. 43, 501–566