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The ion pathway through the opened Na+,K+-ATPase pump

2008; Nature Portfolio; Volume: 456; Issue: 7220 Linguagem: Inglês

10.1038/nature07350

1476-4687

Ayako Takeuchi, Nicolás Reyes, Pablo Artigas, David C. Gadsby,

Marine Toxins and Detection Methods

P-type ATPases are molecular machines that use energy derived from ATP hydrolysis to pump ions across membranes. Recent work on the Na+, K+ ATPase has identified a pathway for ions from the extracellular side of the protein to its ion binding sites. Here, Takeuchi et al. employ the marine toxin palytoxin to 'hold open' the gates of the pump, allowing direct measurements of ion flow. They define the entire route for ions from one side of the membrane to the other. P-type ATPases are molecular machines that use energy derived from ATP hydrolysis to pump ions across membranes. Recent work on the Na+, K+ ATPase has identified a pathway for ions from the extracellular side of the protein to its ion binding sites. This paper employs the marine toxin palytoxin to 'hold open' the gates of the pump, allowing direct measurements of ion flow and defines the entire route for ions from one side of the membrane to the other. P-type ATPases pump ions across membranes, generating steep electrochemical gradients that are essential for the function of all cells. Access to the ion-binding sites within the pumps alternates between the two sides of the membrane1 to avoid the dissipation of the gradients that would occur during simultaneous access. In Na+,K+-ATPase pumps treated with the marine agent palytoxin, this strict alternation is disrupted and binding sites are sometimes simultaneously accessible from both sides of the membrane, transforming the pumps into ion channels (see, for example, refs 2, 3). Current recordings in these channels can monitor accessibility of introduced cysteine residues to water-soluble sulphydryl-specific reagents4. We found previously5 that Na+,K+ pump-channels open to the extracellular surface through a deep and wide vestibule that emanates from a narrower pathway between transmembrane helices 4 and 6 (TM4 and TM6). Here we report that cysteine scans from TM1 to TM6 reveal a single unbroken cation pathway that traverses palytoxin-bound Na+,K+ pump-channels from one side of the membrane to the other. This pathway comprises residues from TM1, TM2, TM4 and TM6, passes through ion-binding site II, and is probably conserved in structurally and evolutionarily related P-type pumps, such as sarcoplasmic- and endoplasmic-reticulum Ca2+-ATPases and H+,K+-ATPases.