Na+-K+-ATPase α-subunit containing Q905-V930 of gastric H+-K+- ATPase α preferentially assembles with H+-K+-ATPase β

Shyang Guang Wang, Kurt A. Eakle, Robert Levenson, Robert A. Farley

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16 Scopus citations

Abstract

Amino acids N886-A911 of the rat Na+-K+-ATPase α3-subunit were replaced by the corresponding region (Q905-V930) of the rat gastric H+-K+- ATPase α-subunit. The chimera (NGH26) was expressed in yeast with the rat Na+-K+-ATPase β1-subunit (rβ1), the rat H+-K+-ATPase β-subunit (HKβ), the chimeric β-subunit NHβ1 (containing the carboxy-terminal ectodomain of HKβ), or the chimeric β-subunit HNβ1 (containing the carboxy-terminal ectodomain of rβ1). Increased resistance to trypsin digestion indicated that NGH26 preferentially assembled with HKβ and NHβ1 rather than with rβ1 or HNβ1. Ouabain binding also indicated that more functional complexes were assembled when NGH26 was expressed with HKβ or NHβ1. These results suggest that the sequence Q905-V930 interacts with the HKβ-subunit on the extracellular side of the cell membrane. The NGH26+HKβ complex is less stable than α3+HKβ when heated and also has a lower binding affinity for ouabain [dissociation constant (K(d)) = 63 nM] compared with α3+rβ1 or α3+HKβ (K(d)= 5-10 nM). In contrast, the NGH26+NHβ1 complex is thermally as stable as α3+rβ1 complexes, and its ouabain binding affinity (K(d) = 10 nM) is the same as the wild type. These results indicate that the amino acids Q905-V930 of the rat gastric H+-K+-ATPase α- subunit preferentially associate with the extracellular domain of H+-K+- ATPase β-subunit to form functional pump complexes and that the cytoplasmic and/or transmembrane region of the β-subunit influences the stability of the αβ complexes.

Original languageEnglish (US)
Pages (from-to)C923-C930
JournalAmerican Journal of Physiology - Cell Physiology
Volume272
Issue number3 41-3
StatePublished - Mar 1 1997

All Science Journal Classification (ASJC) codes

  • Physiology
  • Cell Biology

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