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Crystal structure of Ca²⁺/H⁺ antiporter protein YfkE reveals the mechanisms of Ca²⁺ efflux and its pH regulation
- Wu, Mousheng, Tong, Shuilong, Waltersperger, Sandro, Diederichs, Kay, Wang, Meitian, Zheng, Lei
- Proceedings of the National Academy of Sciences of the United States of America 2013 v.110 no.28 pp. 11367-11372
- Bacillus subtilis, Methanococcus, amino acid substitution, antiporters, calcium, cell membranes, crosslinking, crystal structure, cytoplasm, energy, evolutionary adaptation, glutamic acid, homeostasis, hydrophilicity, hydrophobicity, pH, protein subunits, sodium
- Ca ²⁺ efflux by Ca ²⁺ cation antiporter (CaCA) proteins is important for maintenance of Ca ²⁺ homeostasis across the cell membrane. Recently, the monomeric structure of the prokaryotic Na ⁺/Ca ²⁺ exchanger (NCX) antiporter NCX_Mj protein from Methanococcus jannaschii shows an outward-facing conformation suggesting a hypothesis of alternating substrate access for Ca ²⁺ efflux. To demonstrate conformational changes essential for the CaCA mechanism, we present the crystal structure of the Ca ²⁺/H ⁺ antiporter protein YfkE from Bacillus subtilis at 3.1-Å resolution. YfkE forms a homotrimer, confirmed by disulfide crosslinking. The protonated state of YfkE exhibits an inward-facing conformation with a large hydrophilic cavity opening to the cytoplasm in each protomer and ending in the middle of the membrane at the Ca ²⁺-binding site. A hydrophobic “seal” closes its periplasmic exit. Four conserved α-repeat helices assemble in an X-like conformation to form a Ca ²⁺/H ⁺ exchange pathway. In the Ca ²⁺-binding site, two essential glutamate residues exhibit different conformations compared with their counterparts in NCX_Mj, whereas several amino acid substitutions occlude the Na ⁺-binding sites. The structural differences between the inward-facing YfkE and the outward-facing NCX_Mj suggest that the conformational transition is triggered by the rotation of the kink angles of transmembrane helices 2 and 7 and is mediated by large conformational changes in their adjacent transmembrane helices 1 and 6. Our structural and mutational analyses not only establish structural bases for mechanisms of Ca ²⁺/H ⁺ exchange and its pH regulation but also shed light on the evolutionary adaptation to different energy modes in the CaCA protein family.