The mechanism of sodium and substrate release from the binding pocket of vSGLT

Sodium-coupled transport In membrane proteins with a 'LeuT-fold', it is not clear how ion- and substrate-transport are coupled. Watanabe et al . present a comprehensive study of the structure and biochemical properties of the sodium/galactose transporter (vSGLT) from Vibrio parahaemolyticus , and a new crystal structure of the 'inward-open' conformation. These experiments show that sodium exit causes a reorientation of transmembrane helix 1, opening an inner gate required for substrate exit, while also triggering minor rigid body movements in two sets of transmembrane helical bundles. This cascade of conformational changes is responsible for the proper timing of ion and substrate release. Here, a comprehensive study of the sodium/galactose transporter (vSGLT) is presented, consisting of molecular dynamics simulations, biochemical characterization and a new crystal structure of the 'inward-open' conformation. These experiments show that sodium exit causes a reorientation of transmembrane helix 1, opening an inner gate required for substrate exit, while also triggering minor rigid-body movements in two sets of transmembrane helical bundles. This cascade of conformational changes is responsible for the proper timing of ion and substrate release. Membrane co-transport proteins that use a five-helix inverted repeat motif have recently emerged as one of the largest structural classes of secondary active transporters 1 , 2 . However, despite many structural advances there is no clear evidence of how ion and substrate transport are coupled. Here we report a comprehensive study of the sodium/galactose transporter from Vibrio parahaemolyticus (vSGLT), consisting of molecular dynamics simulations, biochemical characterization and a new crystal structure of the inward-open conformation at a resolution of 2.7 Å. Our data show that sodium exit causes a reorientation of transmembrane helix 1 that opens an inner gate required for substrate exit, and also triggers minor rigid-body movements in two sets of transmembrane helical bundles. This cascade of events, initiated by sodium release, ensures proper timing of ion and substrate release. Once set in motion, these molecular changes weaken substrate binding to the transporter and allow galactose readily to enter the intracellular space. Additionally, we identify an allosteric pathway between the sodium-binding sites, the unwound portion of transmembrane helix 1 and the substrate-binding site that is essential in the couplin