TY - JOUR
T1 - Exploring the Li+ transporting mutant of NCX_Mj for assigning ion binding sites of mitochondrial NCLX
AU - Giladi, Moshe
AU - Mitra, Sunayana
AU - Simhaev, Luba
AU - Hiller, Reuben
AU - Refaeli, Bosmat
AU - Strauss, Tali
AU - Baiz, Carlos R.
AU - Khananshvili, Daniel
N1 - Publisher Copyright:
© 2022
PY - 2022/11
Y1 - 2022/11
N2 - The plasma membrane (NCX) and mitochondrial (NCLX) Na+/Ca2+ exchangers are structurally related proteins, although they operate under strictly different ionic conditions and membrane potentials. In contrast with NCX, NCLX can transport either Li+ or Na+ in exchange for Ca2+. Whereas the crystal structure of the archaeal NCX (NCX_Mj) describes the binding sites for alternative binding of 3Na+ or 1Ca2+, these features remain elusive for NCLX due to the lack of structural information. To elucidate the ion-binding features of mitochondrial NCLX, we analyzed here the Li+-transporting NCLX_Mj mutant, produced by replacing the ion-coordinating residues in the archaeal NCX (NCX_Mj) to match the ion-coordinating residues of human NCLX. The NCLX_Mj-mediated Na+/Ca2+ or Li+/Ca2+ exchange rates are insensitive to varying voltage, consistent with an electroneutral ion exchange. Molecular dynamics (MD) simulations revealed that NCLX_Mj contains two novel Li+ binding sites with four ion-coordinating residues, derived from the three Na+ binding sites of NCX_Mj. The ion-coordination modes, observed in the MD simulations, were further supported by two-dimensional infrared (2D IR) spectroscopy and by testing the mutational effects on the ion fluxes. Collectively, our results revealed a structural basis for Li+ binding and electroneutral transport (2Na+/Li+:1Ca2+) by NCLX_Mj, meaning that the NCLX-mediated electroneutral transport may predefine mitochondrial Ca2+ and Na+ signaling to modulate cellular functions.
AB - The plasma membrane (NCX) and mitochondrial (NCLX) Na+/Ca2+ exchangers are structurally related proteins, although they operate under strictly different ionic conditions and membrane potentials. In contrast with NCX, NCLX can transport either Li+ or Na+ in exchange for Ca2+. Whereas the crystal structure of the archaeal NCX (NCX_Mj) describes the binding sites for alternative binding of 3Na+ or 1Ca2+, these features remain elusive for NCLX due to the lack of structural information. To elucidate the ion-binding features of mitochondrial NCLX, we analyzed here the Li+-transporting NCLX_Mj mutant, produced by replacing the ion-coordinating residues in the archaeal NCX (NCX_Mj) to match the ion-coordinating residues of human NCLX. The NCLX_Mj-mediated Na+/Ca2+ or Li+/Ca2+ exchange rates are insensitive to varying voltage, consistent with an electroneutral ion exchange. Molecular dynamics (MD) simulations revealed that NCLX_Mj contains two novel Li+ binding sites with four ion-coordinating residues, derived from the three Na+ binding sites of NCX_Mj. The ion-coordination modes, observed in the MD simulations, were further supported by two-dimensional infrared (2D IR) spectroscopy and by testing the mutational effects on the ion fluxes. Collectively, our results revealed a structural basis for Li+ binding and electroneutral transport (2Na+/Li+:1Ca2+) by NCLX_Mj, meaning that the NCLX-mediated electroneutral transport may predefine mitochondrial Ca2+ and Na+ signaling to modulate cellular functions.
KW - 2D IR
KW - Ion selectivity
KW - Ion-binding sites
KW - NCLX
KW - NCX
KW - Sodium-calcium exchanger
UR - http://www.scopus.com/inward/record.url?scp=85137853433&partnerID=8YFLogxK
U2 - 10.1016/j.ceca.2022.102651
DO - 10.1016/j.ceca.2022.102651
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C2 - 36116246
AN - SCOPUS:85137853433
SN - 0143-4160
VL - 107
JO - Cell Calcium
JF - Cell Calcium
M1 - 102651
ER -