Transmembrane Segment XI of the Na⁺/H⁺ Antiporter of S. pombe is a Critical Part of the Ion Translocation Pore

The Na⁺/H⁺ exchanger of the plasma membrane of S. pombe (SpNHE1) removes intracellular sodium in exchange for an extracellular proton. We examined the structure and functional role of amino acids 360-393 of putative transmembrane (TM) segment XI of SpNHE1. Structural analysis suggested that it had a helical propensity over amino acids 360-368, an extended region from 369-378 and was helical over amino acids 379-386. TM XI was sensitive to side chain alterations. Mutation of eight amino acids to alanine resulted in loss of one or both of LiCl or NaCl tolerance when re-introduced into SpNHE1 deficient S. pombe. Mutation of seven other amino acids had minor effects. Analysis of structure and functional mutations suggested that Glu³⁶¹ may be involved in cation coordination on the cytoplasmic face of the protein with a negative charge in this position being important. His³⁶⁷, Ile³⁷¹ and Gly³⁷² were important in function. Ile³⁷¹ may have important hydrophobic interactions with other residues and Gly³⁷² may be important in maintaining an extended conformation. Several residues from Val³⁷⁷ to Leu³⁸⁴ are important in function possibly involved in hydrophobic interactions with other amino acids. We suggest that TM XI forms part of the ion translocation core of this Na⁺/H⁺ exchanger.