The role of the third extracellular loop of the Na+,K+-ATPase alpha subunit in a luminal gating mechanism.

Details

Serval ID
serval:BIB_0CAB75E1B292
Type
Article: article from journal or magazin.
Collection
Publications
Institution
Title
The role of the third extracellular loop of the Na+,K+-ATPase alpha subunit in a luminal gating mechanism.
Journal
Journal of Physiology
Author(s)
Capendeguy O., Horisberger J.D.
ISSN
0022-3751
Publication state
Published
Issued date
05/2005
Peer-reviewed
Oui
Volume
565
Number
Pt 1
Pages
207-218
Language
english
Abstract
Na+,K+-ATPase is responsible for maintaining the cross-membrane Na+ and K+ gradients of animal cells. This P-type ATPase works via a complex transport cycle, during which it binds and occludes three intracellular Na+ ions and then two extracellular K+ ions, which it then releases on the other side of the membrane. The cation pathway through the protein, and the structures responsible for occluding cations inside the protein, have not yet been definitely identified. We used cysteine mutagenesis to explore the accessibility and the role of five conserved residues in the short third extracellular loop, between the fifth and the sixth transmembrane helices. The P801C and L802C mutants were not affected by extracellular sulfhydryl reagents. The presence of cysteine residues at three positions (G803C, T804C and V805C) conferred sensitivity to omeprazole, a known inhibitor of the gastric proton pump, and to [2-(trimethylammonium)-ethyl]methanethiosulphonate bromide (MTSET). The effects of omeprazole and MTSET were modulated by the presence of extracellular K+, indicating that the accessibility of these positions depended on the conformational state of the protein. MTSET binding to cysteine at position 803 partially inhibited the Na+,K+-pump function by decreasing its affinity towards extracellular K+, suggesting a restriction of the access of extracellular K+ ions to their binding sites. In contrast, MTSET binding to cysteine at position 805 partially inhibited the Na+,K+-pump function by reducing its maximum turnover rate, probably by slowing a rate-limiting conformational change. These residues occupy positions that are critical for either the cation pathway or the conformational modifications.
Keywords
Animals, Bufo marinus/genetics, Bufo marinus/metabolism, Cell Membrane/chemistry, Cell Membrane/physiology, Cells, Cultured, Cysteine/chemistry, Cysteine/metabolism, Ion Channel Gating/physiology, Membrane Potentials/physiology, Mutagenesis, Site-Directed, Oocytes/physiology, Protein Conformation, Protein Subunits, Recombinant Proteins/chemistry, Recombinant Proteins/metabolism, Sodium-Potassium-Exchanging ATPase/chemistry, Sodium-Potassium-Exchanging ATPase/genetics, Structure-Activity Relationship, Xenopus laevis
Pubmed
Web of science
Open Access
Yes
Create date
24/01/2008 12:38
Last modification date
20/08/2019 12:34
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