Substrate selectivity in glutamate-dependent acid resistance in enteric bacteria

Area of Science:

• Microbiology
• Biochemistry
• Molecular Biology

Background:

• Enteric bacteria possess a glutamate-dependent acid resistance system to survive stomach acidity.
• The antiporter GadC is crucial for this system, importing glutamate and exporting gamma-aminobutyric acid.
• Previous models of GadC function oversimplified substrate protonation states and their impact on proton transport.

Purpose of the Study:

• To elucidate the specific substrate exchange mechanisms employed by GadC under acidic conditions.
• To determine how GadC differentiates between protonated and deprotonated substrate forms.
• To understand the role of substrate charge in GadC's selectivity and proton extrusion efficiency.

Main Methods:

• Development of an oriented GadC liposome system.
• Creation of an inward pH gradient to simulate gastric conditions.
• Assessment of substrate transport electrogenicity to determine proton stoichiometry.

Main Results:

• GadC selectively exchanges glutamate (Glu(-) or Glu(0)) with gamma-aminobutyric acid (GABA(+)).
• This selective exchange results in effective proton extrusion (>0.9 H(+) per turnover).
• GadC utilizes a charge-based recognition mechanism for substrate selection, not direct protonation status recognition.

Conclusions:

• GadC actively extrudes protons by selectively transporting specific charged forms of glutamate and gamma-aminobutyric acid.
• The antiporter's charge-based selection mechanism is key to its function in acid resistance.
• This study provides a foundation for investigating the molecular basis of GadC's substrate selectivity.