Voltage-dependent proton fluxes in liposomes

Area of Science:

• Biochemistry
• Biophysics
• Membrane Biology

Background:

• Liposomes are widely used to study membrane transport.
• Diffusion potentials can influence proton movement across lipid bilayers.
• Bacterial lipids can form functional liposomes.

Purpose of the Study:

• To investigate proton influx into liposomes driven by a diffusion potential (delta psi).
• To characterize the conditions under which delta psi-induced proton flux occurs.
• To assess the implications of this phenomenon for reconstituted membrane systems.

Main Methods:

• Preparation of liposomes from bacterial lipids and synthetic lipids containing buffered KCl.
• Induction of a diffusion potential (delta psi) using valinomycin in K+-free media.
• Assay of proton influx by 9-aminoacridine fluorescence quenching.
• Manipulation of membrane potential by altering ion concentrations and using chemical agents.

Main Results:

• Substantial proton influx was observed upon formation of a diffusion potential (delta psi).
• Proton influx was dependent on the magnitude of delta psi, occurring above -60 mV.
• Proton influx was also observed with a sodium diffusion potential and in synthetic liposomes.
• Proton flux was not inhibited by N,N'-dicyclohexylcarbodiimide, suggesting a non-ATP-dependent mechanism.

Conclusions:

• Diffusion potentials can induce significant proton fluxes in liposomes, independent of lipid source.
• These proton fluxes can complicate the interpretation of studies involving reconstituted membrane proteins and proton translocation.
• The magnitude of the electrochemical proton gradient in reconstituted systems may be affected by diffusion potential-driven proton movement.