2.A.22.4.2

The amino acid (leucine):2 Na⁺ symporter, LeuT_Aa (Yamashita et al., 2005). LeuT possesses two ion binding sites, NA1 and NA2, both highly specific for Na⁺ but with differing mechanisms of binding (Noskov and Roux, 2008). X-ray structures have been determined for LeuT in substrate-free outward-open and apo inward-open states (Krishnamurthy and Gouaux, 2012).  Extracytoplasmic substrate binding at an allosteric site controls activity (Zhao et al. 2011).  It has been proposed that the 5 TMS repeat derived from a DedA domain (9.B.27; Khafizov et al. 2010).  Mechanistic aspect of Na⁺ binding have been studied (Perez and Ziegler 2013).  Structural studies of mutant LeuT proteins suggest how antidepressants bind to biogenic amine transporters (Wang et al. 2013).  The detailed mechanism was studied by Zhao and Noskov, 2013.  Uptake involves movement of the substrate amino acid from the outward facing binding site, S1, to the inward facing binding site, S2, coupled with confrmational changes in the protein (Cheng and Bahar 2013).  The complete substrate translocation pathway has been proposed (Cheng and Bahar 2014). The inward facing conformation of LeuT has been solved (Grouleff et al. 2015).  Substrate-induced unlocking of the inner gatemay determinethe catalytic efficiency of the transporter (Billesbølle et al. 2015). Of the two Na⁺ binding sites, occupation of Na2 stabilizes outward-facing conformations presumably through a direct interaction between Na⁺ and transmembrane helices 1 and 8 whereas Na⁺ binding at Na1 influences conformational change through a network of intermediary interactions (Tavoulari et al. 2015). TMS1A movements revealed a substantially different inward-open conformation in lipid bilayer from that inferred from the crystal structure, especiallly with respect to the inner vestibule (Sohail et al. 2016). Partial unwinding of transmembrane helices 1, 5, 6 and7 drives LeuT from a substrate-bound, outward-facing occluded conformation toward an inward-facing open state (Merkle et al. 2018). A conserved tyrosine residue in the substrate binding site is required for substrate binding to convert LeuT to inward-open states by establishing an interaction between the two transporter domains (Zhang et al. 2018). The X-ray structure of LeuT in an inward-facing occluded conformation has revealed the mechanism of substrate release (Gotfryd et al. 2020). This involves a major tilting of the cytoplasmic end of TMS5, which, together with release of the N-terminus but without coupled movement of TM1) opens a wide cavity towards the second Na⁺ binding site. The X-ray structure of LeuT in an inward-facing occluded conformation has been solved, revealing the mechanism of substrate release (Gotfryd et al. 2020).  In nine transporters having the LeuT fold, the bundle (first two TMSs of each 5 TMS repeat) rotates relative to the hash (third and fourth TMSs). Motions of the arms (fifth TMS) to close or open the intracellular and outer vestibules are common, as is a TMS1a swing, with notable variations in the opening-closing motions of the outer vestibule. These analyses suggest that LeuT-fold transporters layer distinct motions on a common bundle-hash rock (Licht et al. 2024).