The single pore residue Asp542 determines Ca2+ permeation and Mg2+ block of the epithelial Ca2+ channel

J Biol Chem. 2001 Jan 12;276(2):1020-5. doi: 10.1074/jbc.M006184200.


The epithelial Ca(2+) channel (ECaC), which was recently cloned from rabbit kidney, exhibits distinctive properties that support a facilitating role in transcellular Ca(2+) (re)absorption. ECaC is structurally related to the family of six transmembrane-spanning ion channels with a pore-forming region between S5 and S6. Using point mutants of the conserved negatively charged amino acids present in the putative pore, we have identified a single aspartate residue that determines Ca(2+) permeation of ECaC and modulation by extracellular Mg(2+). Mutation of the aspartate residue, D542A, abolishes Ca(2+) permeation and Ca(2+)-dependent current decay as well as block by extracellular Mg(2+), whereas monovalent cations still permeate the mutant channel. Variation of the side chain length in mutations D542N, D542E, and D542M attenuated Ca(2+) permeability and Ca(2+)-dependent current decay. Block of monovalent currents through ECaC by Mg(2+) was decreased. Exchanging the aspartate residue for a positively charged amino acid, D542K, resulted in a nonfunctional channel. Mutations of two neighboring negatively charged residues, i.e. Glu(535) and Asp(550), had only minor effects on Ca(2+) permeation properties.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Amino Acid Substitution
  • Animals
  • Aspartic Acid*
  • Calcium / metabolism*
  • Calcium Channels / chemistry*
  • Calcium Channels / drug effects
  • Calcium Channels / physiology*
  • Humans
  • Kidney / metabolism
  • Kinetics
  • Magnesium / pharmacology*
  • Membrane Potentials / drug effects
  • Membrane Potentials / physiology
  • Mice
  • Models, Molecular
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Protein Structure, Secondary
  • Rabbits
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / drug effects
  • Recombinant Proteins / metabolism
  • Sequence Alignment
  • Sequence Homology, Amino Acid
  • TRPV Cation Channels
  • Transfection


  • Calcium Channels
  • ECaC protein, Oryctolagus cuniculus
  • Recombinant Proteins
  • TRPV Cation Channels
  • TRPV5 protein, human
  • Trpv5 protein, mouse
  • Aspartic Acid
  • Magnesium
  • Calcium