Mutation of Aspartate 555 of the Sodium/Bicarbonate Transporter SLC4A4/NBCe1 Induces Chloride Transport

J Biol Chem. 2009 Jun 5;284(23):15970-9. doi: 10.1074/jbc.M109.001438. Epub 2009 Mar 31.


To understand the mechanism for ion transport through the sodium/bicarbonate transporter SLC4A4 (NBCe1), we examined amino acid residues, within transmembrane domains, that are conserved among electrogenic Na/HCO(3) transporters but are substituted with residues at the corresponding site of all electroneutral Na/HCO(3) transporters. Point mutants were constructed and expressed in Xenopus oocytes to assess function using two-electrode voltage clamp. Among the mutants, D555E (charge-conserved substitution of the aspartate at position 555 with a glutamate) produced decreasing HCO(3)(-) currents at more positive membrane voltages. Immunohistochemistry showed D555E protein expression in oocyte membranes. D555E induced Na/HCO(3)-dependent pH recovery from a CO(2)-induced acidification. Current-voltage relationships revealed that D555E produced an outwardly rectifying current in the nominally CO(2)/HCO(3)(-)-free solution that was abolished by Cl(-) removal from the bath. In the presence of CO(2)/HCO(3)(-), however, the outward current produced by D555E decreased only slightly after Cl(-) removal. Starting from a Cl(-)-free condition, D555E produced dose-dependent outward currents in response to a series of chloride additions. The D555E-mediated chloride current decreased by 70% in the presence of CO(2)/HCO(3)(-). The substitution of Asp(555) with an asparagine also produced a Cl(-) current. Anion selectivity experiments revealed that D555E was broadly permissive to other anions including NO(3)(-). Fluorescence measurements of chloride transport were done with human embryonic kidney HEK 293 cells expressing NBCe1 and D555E. A marked increase in chloride transport was detected in cells expressing D555E. We conclude that Asp(555) plays a role in HCO(3)(-) selectivity.

MeSH terms

  • Amino Acid Substitution
  • Animals
  • Aspartic Acid / genetics*
  • Biological Transport
  • Cell Line
  • Chlorides / metabolism*
  • Female
  • Humans
  • Hydrogen-Ion Concentration
  • Kidney / physiology
  • Kinetics
  • Models, Molecular
  • Mutagenesis, Site-Directed*
  • Oocytes / physiology*
  • Patch-Clamp Techniques
  • Plasmids
  • Protein Conformation
  • Sodium-Bicarbonate Symporters / chemistry
  • Sodium-Bicarbonate Symporters / genetics*
  • Transfection
  • Xenopus


  • Chlorides
  • SLC4A4 protein, human
  • Sodium-Bicarbonate Symporters
  • Aspartic Acid