Dynamic features of allosteric Ca2+ sensor in tissue-specific NCX variants

Cell Calcium. 2012 Jun;51(6):478-85. doi: 10.1016/j.ceca.2012.04.007. Epub 2012 May 7.

Abstract

The Na(+)-Ca(2+) exchanger (NCX) mediated Ca(2+) fluxes are essential for handling Ca(2+) homeostasis in many cell-types. Eukaryotic NCX variants contain regulatory CBD1 and CBD2 domains, whereas in distinct variants the Ca(2+) binding to Ca3-Ca4 sites of CBD1 results either in sustained activation, inhibition or no effect. CBD2 contains an alternatively spliced segment, which is expressed in a tissue-specific manner although its impact on allosteric regulation remains unclear. Recent studies revealed that the Ca(2+) binding to Ca3-Ca4 sites results in interdomain tethering of CBDs, which rigidifies CBDs movements with accompanied slow dissociation of "occluded" Ca(2+). Here we investigate the effects of CBD2 variants on Ca(2+) occlusion in the two-domain construct (CBD12). Mutational studies revealed that both sites (Ca3 and Ca4) contribute to Ca(2+) occlusion, whereas after dissociation of the first Ca(2+) ion the second Ca(2+) ion becomes occluded. This mechanism is common for the brain, kidney and cardiac splice variants of CBD12, although the occluded Ca(2+) exhibits 20-50-fold difference in off-rates among the tested variants. Therefore, the spliced exons on CBD2 affect the rate-limiting step of the occluded Ca(2+) dissociation at the primary regulatory sensor to shape dynamic features of allosteric regulation in NCX variants.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Allosteric Regulation
  • Alternative Splicing
  • Amino Acid Sequence
  • Animals
  • Binding Sites
  • Brain Chemistry
  • Calcium / chemistry*
  • Calcium Signaling
  • Cloning, Molecular
  • Dogs
  • Drosophila Proteins / chemistry
  • Drosophila Proteins / genetics
  • Drosophila melanogaster / genetics
  • Escherichia coli / chemistry
  • Escherichia coli / genetics
  • Exons
  • Kidney / chemistry
  • Molecular Sequence Data
  • Mutation
  • Point Mutation
  • Protein Binding
  • Protein Conformation
  • Protein Isoforms / chemistry
  • Protein Structure, Tertiary
  • Sodium-Calcium Exchanger / chemistry*

Substances

  • Drosophila Proteins
  • Protein Isoforms
  • Sodium-Calcium Exchanger
  • Calcium