Identification of a region within the Na,K-ATPase alpha subunit that contributes to differential ouabain sensitivity

Mol Cell Biol. 1989 Sep;9(9):3744-9. doi: 10.1128/mcb.9.9.3744-3749.1989.

Abstract

To analyze determinants within the Na,K-ATPase alpha subunit that contribute to differential ouabain sensitivity, we constructed and expressed a panel of chimeric cDNA molecules between ouabain-resistant and ouabain-sensitive alpha subunit cDNAs. When introduced into ouabain-sensitive monkey CV-1 cells, ouabain-resistant rat alpha 1 subunit cDNA and chimeras in which the 5' end of ouabain-sensitive human alpha 1 or rat alpha 2 subunit cDNA was replaced by the 5' end of rat alpha 1 subunit cDNA conferred resistance to 100 microM ouabain. Monkey cells transfected with the reciprocal chimeras were unable to survive selection in 1 microM ouabain. Rat alpha 2 subunit cDNA and a chimera in which the 5' end of rat alpha 1 subunit cDNA was replaced by the 5' end of rat alpha 2 subunit cDNA conferred resistance to 0.5 microM ouabain. These results suggest that determinants of ouabain resistance reside within the amino-terminal portions of the rat alpha 1 and alpha 2 subunits. Expression of chimeric alpha subunit cDNAs should prove useful for elucidating the structural basis of Na,K-ATPase function.

Publication types

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

MeSH terms

  • Animals
  • Binding Sites
  • Chimera
  • DNA / genetics
  • Humans
  • Ouabain / pharmacology*
  • Protein Conformation
  • Rats
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Sodium-Potassium-Exchanging ATPase / antagonists & inhibitors*
  • Sodium-Potassium-Exchanging ATPase / genetics
  • Sodium-Potassium-Exchanging ATPase / metabolism

Substances

  • Recombinant Proteins
  • Ouabain
  • DNA
  • Sodium-Potassium-Exchanging ATPase