Asymmetric Preorganization of Inverted Pair Residues in the Sodium-Calcium Exchanger

Sci Rep. 2016 Feb 15:6:20753. doi: 10.1038/srep20753.


In analogy with many other proteins, Na(+)/Ca(2+) exchangers (NCX) adapt an inverted twofold symmetry of repeated structural elements, while exhibiting a functional asymmetry by stabilizing an outward-facing conformation. Here, structure-based mutant analyses of the Methanococcus jannaschii Na(+)/Ca(2+) exchanger (NCX_Mj) were performed in conjunction with HDX-MS (hydrogen/deuterium exchange mass spectrometry) to identify the structure-dynamic determinants of functional asymmetry. HDX-MS identified hallmark differences in backbone dynamics at ion-coordinating residues of apo-NCX_Mj, whereas Na(+)or Ca(2+) binding to the respective sites induced relatively small, but specific, changes in backbone dynamics. Mutant analysis identified ion-coordinating residues affecting the catalytic capacity (kcat/Km), but not the stability of the outward-facing conformation. In contrast, distinct "noncatalytic" residues (adjacent to the ion-coordinating residues) control the stability of the outward-facing conformation, but not the catalytic capacity. The helix-breaking signature sequences (GTSLPE) on the α1 and α2 repeats (at the ion-binding core) differ in their folding/unfolding dynamics, while providing asymmetric contributions to transport activities. The present data strongly support the idea that asymmetric preorganization of the ligand-free ion-pocket predefines catalytic reorganization of ion-bound residues, where secondary interactions with adjacent residues couple the alternating access. These findings provide a structure-dynamic basis for ion-coupled alternating access in NCX and similar proteins.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Archaeal Proteins / chemistry*
  • Archaeal Proteins / genetics
  • Archaeal Proteins / metabolism
  • Calcium / chemistry*
  • Calcium / metabolism
  • Catalytic Domain
  • Deuterium Exchange Measurement
  • Escherichia coli / genetics
  • Escherichia coli / metabolism
  • Gene Expression
  • Hydrogen-Ion Concentration
  • Ion Transport
  • Mass Spectrometry
  • Methanocaldococcus / chemistry*
  • Methanocaldococcus / genetics
  • Methanocaldococcus / metabolism
  • Models, Molecular
  • Protein Binding
  • Protein Folding
  • Protein Interaction Domains and Motifs
  • Protein Structure, Secondary
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Sodium / chemistry*
  • Sodium / metabolism
  • Sodium-Calcium Exchanger / chemistry*
  • Sodium-Calcium Exchanger / genetics
  • Sodium-Calcium Exchanger / metabolism
  • Structure-Activity Relationship


  • Archaeal Proteins
  • Recombinant Proteins
  • Sodium-Calcium Exchanger
  • Sodium
  • Calcium