title: Identification of residues that control Li+versus Na+dependent Ca2+exchange at the transport site of the mitochondrial NCLX.
abstract: The Na+/Ca2+/Li+exchanger (NCLX) is a member of the Na+/Ca2+exchanger family. NCLX is unique in its capacity to transport both Na+and Li+, unlike other members, which are Na+selective. The major aim of this study was twofold, i.e., to identify NCLX residues that confer Li+or Na+selective Ca2+transport and map their putative location on NCLX cation transport site.
  
  
  
  We combined molecular modeling to map transport site of NCLX with euryarchaeal H+/Ca2+exchanger, CAX_Af, and fluorescence analysis to monitor Li+versus Na+dependent mitochondrial Ca2+efflux of transport site mutants of NCLX in permeabilized cells.
  
  
  
  Mutation of Asn149, Pro152, Asp153, Gly176, Asn467, Ser468, Gly494 and Asn498 partially or strongly abolished mitochondrial Ca2+exchange activity in intact cells. In permeabilized cells, N149A, P152A, D153A, N467Q, S468T and G494S demonstrated normal Li+/Ca2+exchange activity but a reduced Na+/Ca2+exchange activity. On the other hand, D471A showed dramatically reduced Li+/Ca2+exchange, but Na+/Ca2+exchange activity was unaffected. Finally, simultaneous mutation of four putative Ca2+binding residues was required to completely abolish both Na+/Ca2+and Li+/Ca2+exchange activities.
  
  
  
  We identified distinct Na+and Li+selective residues in the NCLX transport site. We propose that functional segregation in Li+and Na+sites reflects the functional properties of NCLX required for Ca2+exchange under the unique membrane potential and ion gradient across the inner mitochondrial membrane.
  
  
  
  The results of this study provide functional insights into the unique Li+and Na+selectivity of the mitochondrial exchanger. This article is part of a Special Issue entitled: ECS Meeting edited by Claus Heizmann, Joachim Krebs and Jacques Haiech.
authors.affiliation: Department of Physiology and Cell Biology, Faculty of Health Sciences, Ben-Gurion University of the Negev, 84105 Beer Sheva, Israel.
authors.forename: Soumitra
authors.initials: S
authors.lastname: Roy
authors.affiliation: Department of Physiology and Cell Biology, Faculty of Health Sciences, Ben-Gurion University of the Negev, 84105 Beer Sheva, Israel.
authors.forename: Kuntal
authors.initials: K
authors.lastname: Dey
authors.affiliation: Department of Physiology and Cell Biology, Faculty of Health Sciences, Ben-Gurion University of the Negev, 84105 Beer Sheva, Israel.
authors.forename: Michal
authors.initials: M
authors.lastname: Hershfinkel
authors.affiliation: Department of Clinical Biochemistry and Pharmacology, Faculty of Health Sciences, Ben Gurion University of the Negev, 84105 Beer Sheva, Israel. Electronic address: ohanaeh@bgu.ac.il.
authors.forename: Ehud
authors.initials: E
authors.lastname: Ohana
authors.affiliation: Department of Physiology and Cell Biology, Faculty of Health Sciences, Ben-Gurion University of the Negev, 84105 Beer Sheva, Israel. Electronic address: israel.sekler@gmail.com.
authors.forename: Israel
authors.initials: I
authors.lastname: Sekler
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D001692:Biological Transport; D002118:Calcium / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D008094:Lithium / Q000378:metabolism*; D008928:Mitochondria / Q000378:metabolism*; D024101:Mitochondrial Proteins; D009154:Mutation; D017386:Sequence Homology, Amino Acid; D012964:Sodium / Q000378:metabolism*; D019831:Sodium-Calcium Exchanger / Q000737:chemistry / Q000378:metabolism*
chemical_list: D024101:Mitochondrial Proteins; C483046:SLC8B1 protein, human; D019831:Sodium-Calcium Exchanger; D008094:Lithium; D012964:Sodium; D002118:Calcium
keywords: 3D modelling; Li(+)/Ca(2+) exchange; NCLX; Na(+)/Ca(2+) exchange; Permeabilized cells