The proton electrochemical gradient in Escherichia coli cells

Area of Science:

• Microbiology
• Biochemistry
• Cell Physiology

Background:

• Understanding the proton motive force (PMF) in bacteria is crucial for comprehending cellular energy transduction.
• Escherichia coli's internal pH regulation and its dependence on respiration are key areas of investigation.

Purpose of the Study:

• To quantify the internal pH and proton motive force (delta muH) in Escherichia coli under various conditions.
• To elucidate the roles of delta pH and membrane potential (delta psi) in maintaining PMF.
• To investigate the specific requirement of potassium for E. coli respiration.

Main Methods:

• Internal pH estimation using radiolabeled compounds (5,5-[14C]dimethyl-2,4-oxazolidinedione or [14C]methylamine).
• Measurement of membrane potential (delta psi) and delta pH using 86Rb+ distribution in EDTA/valinomycin-treated cells.
• Assessment of PMF components under varying extracellular pH and potassium concentrations.
• Inhibition studies using KCN and carbonylcyanide p-trifluoromethoxyphenylhydrazone.

Main Results:

• Intact respiring E. coli cells maintain an internal pH 0.63-0.75 units more alkaline than the external environment at pH 7.
• Delta pH decreases with inhibited respiration; PMF components are influenced by extracellular potassium concentration.
• Potassium is specifically required for the respiration of EDTA-treated E. coli K12 cells, with valinomycin not being necessary in this context.

Conclusions:

• The proton motive force in E. coli is composed of both delta pH and delta psi, with their relative contributions varying based on external conditions.
• Potassium plays a critical and specific role as a counter-ion in E. coli respiration, particularly in EDTA-treated cells.
• These findings support models of electronic proton pumps and highlight the complex regulation of intracellular pH and energy metabolism in bacteria.