Single-channel function of recombinant type 2 inositol 1,4, 5-trisphosphate receptor

Biophys J. 2000 Sep;79(3):1388-99. doi: 10.1016/S0006-3495(00)76391-0.


A full-length rat type 2 inositol 1,4,5-trisphosphate (InsP(3)) receptor cDNA construct was generated and expressed in COS-1 cells. Targeting of the full-length recombinant type 2 receptor protein to the endoplasmic reticulum was confirmed by immunocytochemistry using isoform specific affinity-purified antibodies and InsP(3)R-green fluorescent protein chimeras. The receptor protein was solubilized and incorporated into proteoliposomes for functional characterization. Single-channel recordings from proteoliposomes fused into planar lipid bilayers revealed that the recombinant protein formed InsP(3)- and Ca(2+)-sensitive ion channels. The unitary conductance ( approximately 250 pS; 220/20 mM Cs(+) as charge carrier), gating, InsP(3), and Ca(2+) sensitivities were similar to those previously described for the native type 2 InsP(3)R channel. However, the maximum open probability of the recombinant channel was slightly lower than that of its native counterpart. These data show that our full-length rat type 2 InsP(3)R cDNA construct encodes a protein that forms an ion channel with functional attributes like those of the native type 2 InsP(3)R channel. The possibility of measuring the function of single recombinant type 2 InsP(3)R is a significant step toward the use of molecular tools to define the determinants of isoform-specific InsP(3)R function and regulation.

Publication types

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

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Binding, Competitive
  • COS Cells
  • Calcium / pharmacology
  • Calcium / physiology
  • Calcium Channels / chemistry
  • Calcium Channels / physiology*
  • Inositol 1,4,5-Trisphosphate / metabolism*
  • Inositol 1,4,5-Trisphosphate Receptors
  • Ion Channel Gating / physiology
  • Kinetics
  • Lipid Bilayers
  • Membrane Potentials / physiology
  • Microsomes / metabolism
  • Molecular Sequence Data
  • Proteolipids
  • Rats
  • Receptors, Cytoplasmic and Nuclear / chemistry
  • Receptors, Cytoplasmic and Nuclear / physiology*
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / metabolism
  • Transfection


  • Calcium Channels
  • Inositol 1,4,5-Trisphosphate Receptors
  • Lipid Bilayers
  • Proteolipids
  • Receptors, Cytoplasmic and Nuclear
  • Recombinant Proteins
  • proteoliposomes
  • Inositol 1,4,5-Trisphosphate
  • Calcium