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

Area of Science:

• Membrane protein structure and function
• Ion transport mechanisms
• Cellular electrophysiology

Background:

• P-type ATPases are crucial for maintaining cellular electrochemical gradients.
• Normally, ion-binding sites in these pumps alternate access between membrane sides to prevent gradient dissipation.
• Palytoxin disrupts this alternation in Na(+),K(+)-ATPase, converting pumps into ion channels.

Purpose of the Study:

• To investigate the structural basis of ion permeation in palytoxin-bound Na(+),K(+)-ATPase pump-channels.
• To identify the specific amino acid residues forming the ion pathway.
• To explore the conservation of this pathway in related P-type pumps.

Main Methods:

• Cysteine scanning mutagenesis across transmembrane helices (TM1-TM6) of Na(+),K(+)-ATPase.
• Utilizing water-soluble sulfhydryl-specific reagents to probe accessibility within the pump-channel.
• Electrophysiological recordings to monitor ion flow and reagent accessibility.

Main Results:

• A single, unbroken cation pathway was identified spanning the membrane in palytoxin-bound pump-channels.
• This pathway involves residues from TM1, TM2, TM4, and TM6, including ion-binding site II.
• The pathway is accessible from both sides of the membrane, consistent with channel activity.

Conclusions:

• Palytoxin binding creates a continuous ion permeation pathway through Na(+),K(+)-ATPase.
• This pathway is likely conserved in other P-type ATPases, such as Ca(2+)-ATPases and H(+),K(+)-ATPases.
• Understanding this pathway offers insights into P-type ATPase structure and ion channel function.