Structural identification of cation binding pockets in the plasma membrane proton pump

Area of Science:

• Biochemistry
• Structural Biology
• Membrane Protein Research

Background:

• P-type plasma membrane H(+)-ATPases are vital ion pumps.
• Their activity is regulated by protons (H+) and cations like potassium (K+) and calcium (Ca2+).
• Understanding cation binding is key to elucidating pump regulation and function.

Purpose of the Study:

• To determine the structural basis of cation binding to P-type H(+)-ATPases.
• To identify specific cation binding sites using X-ray crystallography.
• To explore the potential of holmium ions (Ho3+) as probes for proton binding sites.

Main Methods:

• X-ray crystallography was employed to visualize cation binding.
• Rubidium (Rb+) as a K+ congener and terbium (Tb3+) and holmium (Ho3+) as Ca2+ congeners were used.
• Structural analysis focused on identifying ion coordination and location within the enzyme.

Main Results:

• The binding site for Rb+ was located in the phosphorylation domain, coordinated by an aspartate residue.
• A single Tb3+ ion was found near the nucleotide-binding site.
• Ho3+ ions occupied two distinct sites: one between nucleotide-binding and phosphorylation domains, and another in the transmembrane domain.

Conclusions:

• The identified cation binding sites likely play roles in enzyme regulation and proton transport.
• Ho3+ ions represent a novel chemical tool for identifying proton binding sites in H(+)-ATPases.
• Structural insights advance our understanding of P-type ATPase mechanisms.