Interactions between K+ and ATP binding to the (Na+ + K+)-dependent ATPase

Area of Science:

• Biochemistry
• Molecular Biology
• Enzymology

Background:

• The (Na+ + K+)-dependent ATPase, also known as ATP phosphohydrolase (EC 3.6.1.3), is a crucial enzyme in cellular ion transport.
• Understanding the regulatory mechanisms of Na+/K+-ATPase activity is vital for comprehending cellular homeostasis and function.
• Previous studies suggest complex interactions between enzyme substrates and ions, but the precise nature of these interactions remains to be fully elucidated.

Purpose of the Study:

• To quantitatively model the antagonistic effects between potassium (K+) and Mg2+-ATP on Na+/K+-ATPase affinity.
• To identify the specific class of K+ binding sites involved in these interactions.
• To elucidate the role of alpha-sites in modulating enzyme activity in response to K+ and Mg2+-ATP concentrations.

Main Methods:

• Investigated the kinetic effects of K+ and Na+ on the apparent Km for Mg2+-ATP.
• Utilized a quantitative model to describe the binding interactions between ions and the enzyme.
• Measured K+ affinity using K+-accelerated inactivation by fluoride and K+-dependent phosphatase activity.

Main Results:

• K+ increases the apparent Km for Mg2+-ATP, an effect competitively antagonized by Na+.
• A model was developed where K+ binding to alpha-sites inhibits Mg2+-ATP binding.
• Mg2+-ATP decreases the apparent affinity of K+ for the alpha-sites, consistent with observed K+ inhibition at low substrate concentrations.

Conclusions:

• The study provides a quantitative model for K+ and Mg2+-ATP interactions with Na+/K+-ATPase.
• K+ binding to alpha-sites negatively impacts Mg2+-ATP binding, influencing enzyme kinetics.
• The findings support a model where K+ can inhibit Na+/K+-ATPase activity, particularly at low substrate levels, and explain the stimulatory effect of Li+ at high K+ concentrations.