The importance of being aromatic: π interactions in sodium symporters

In the LeuT family of sodium solute symporters, 13-17% of the residues in transmembrane domains are aromatic. The unique properties of aromatic amino acids allow them to play specialized roles in proteins, but their function in membrane transporters is underappreciated. Here we analyze the π bonding pattern in the LeuT (5TMIR) family and then describe the role of a triad of aromatic residues in sodium-dependent sugar cotransporters (SGLTs). In SLC5 symporters, three aromatic residues in TM6 (SGLT1 W289, Y290, and W291) are conserved in only those transporting sugars and inositols. We used biophysical analysis of mutants to discover their functional roles, which we have interpreted in terms of CH-π, π-π, and cation-π bonding. We discovered that (1) glucose binding involves CH-π stacking with Y290, (2) π T-stacking interactions between Y290 and W291 and H-bonding between Y290 and N78 (TM1) are essential to form the sodium and sugar binding sites, (3) the Na⁺:sugar stoichiometry is determined by these residues, and (4) W289 may be important in stabilizing the structure through H-bonding to TM3. We also find that the WYW triad plays a role in Na⁺ coordination at the Na1 site, possibly through cation-π interactions. Surprisingly, this Na ⁺ is not necessarily coupled to glucose translocation. Our analysis of π interactions in other LeuT proteins suggests that they also contribute to the structure and function in this whole family of transporters.