Figure 6: MD simulations of extracellular Na⁺ interactions in Na⁺/substrate-free, outward-oriented states.

LeuT structure with Leu25 at S1 substrate-binding site (the crystal structure, (a) and with Leu25 rotated out of the substrate-binding site (modelled, b). The cyan mesh represents an isosurface corresponding to at least 1% Na⁺ occupancy of the volume during the simulations on the extracellular side. Green circles indicate the approximate positions of the Na1 and Na2 sites in a–c, r.m.s.d of Leu25 (calculated using the Cα, Cβ and Cγ atoms) for four independent MD simulations shown in different colours. The r.m.s.d is measured with respect to the outward-facing, Na⁺- and substrate-free return state of LeuT with Leu25 positioned at the S1 site (top) and with respect to the outward-facing, Na⁺- and substrate-bound state of LeuT with Leu25 positioned away from the S1 site (PDB entry code 2A65).