Stoichiometry of a Cl(-)-translocating ATPase

Area of Science:

• Biochemistry
• Cellular Physiology

Background:

• The chloride pump plays a crucial role in cellular ion transport.
• Understanding its reaction sequence and energy coupling is vital for cellular function.

Purpose of the Study:

• To elucidate the roles of Mg2+ and Cl- in the chloride pump's reaction sequence.
• To determine the stoichiometry of chloride transport per ATP hydrolyzed.
• To calculate the operant free energy of ATP hydrolysis for the pump.

Main Methods:

• Investigated the effect of Mg2+ on phosphorylation and Cl- on dephosphorylation.
• Quantified ATP hydrolysis and Cl- transport stoichiometry.
• Measured intracellular ATP, ADP, and inorganic phosphate concentrations.
• Calculated the standard and operant free energy of ATP hydrolysis.

Main Results:

• Mg2+ was found to stimulate phosphorylation, while Cl- stimulated dephosphorylation.
• The stoichiometry of ATP hydrolysis to Cl- transport was determined.
• Operant free energy for ATP hydrolysis was calculated based on intracellular concentrations and standard free energy.
• The pump's operating free energy is approximately half the total free energy from ATP hydrolysis.

Conclusions:

• The chloride pump utilizes approximately half the energy from ATP hydrolysis.
• Possible integral stoichiometries for net charge transport per ATP hydrolyzed are one or two.