Conformational transitions of the sodium-dependent sugar transporter, vSGLT
Sodium-dependent transporters couple the flow of Na ions down their electrochemical potential gradient to the uphill transport of various ligands. Many of these transporters share a common core structure composed of a five-helix inverted repeat and deliver their cargo utilizing an alternating-access mechanism. A detailed characterization of inward-facing conformations of the Na-dependent sugar transporter from (vSGLT) has previously been reported, but structural details on additional conformations and on how Na and ligand influence the equilibrium between other states remains unknown. Here, double electron-electron resonance spectroscopy, structural modeling, and molecular dynamics are utilized to deduce ligand-dependent equilibria shifts of vSGLT in micelles. In the absence and presence of saturating amounts of Na, vSGLT favors an inward-facing conformation. Upon binding both Na and sugar, the equilibrium shifts toward either an outward-facing or occluded conformation. While Na alone does not stabilize the outward-facing state, gating charge calculations together with a kinetic model of transport suggest that the resting negative membrane potential of the cell, absent in detergent-solubilized samples, may stabilize vSGLT in an outward-open conformation where it is poised for binding external sugars. In total, these findings provide insights into ligand-induced conformational selection and delineate the transport cycle of vSGLT.