title: Substrate selectivity of the acid-activated glutamate/γ-aminobutyric acid (GABA) antiporter GadC from Escherichia coli.
abstract: GadC, a central component of the Escherichia coli acid resistance system, is a Glu/GABA antiporter. A previous structural study and biochemical characterization showed that GadC exhibits a stringent pH dependence for substrate transport, with no detectable activity at pH values above 6.5. However, the substrate selectivity and the mechanism of pH-dependent transport activity of GadC remain enigmatic. In this study, we demonstrate that GadC selectively transports Glu with no net charge and GABA with a positive charge. A C-plug-truncated variant of GadC (residues 1-470) transported Gln (a mimic of Glu with no net charge), but not Glu, even at pH 8.0. The pH-dependent transport of Gln by this GadC variant was shifted ~1 unit toward a higher pH compared with Glu transport. Taken together, the results identify the substrate selectivity for GadC and show that the protonation states of substrates are crucial for transport.
authors.affiliation: Ministry of Education Protein Science Laboratory and the Joint Center for Life Sciences, Center for Structural Biology, Schools of Life Sciences and Medicine, Tsinghua University, Beijing 100084, China.
authors.forename: Dan
authors.initials: D
authors.lastname: Ma
authors.forename: Peilong
authors.initials: P
authors.lastname: Lu
authors.forename: Yigong
authors.initials: Y
authors.lastname: Shi
mesh_terms: D001693:Biological Transport, Active / Q000502:physiology*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D005973:Glutamine / Q000235:genetics / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D013379:Substrate Specificity / Q000502:physiology
chemical_list: D029968:Escherichia coli Proteins; C491226:GadC protein, E coli; D008565:Membrane Proteins; D005973:Glutamine
keywords: Acid Resistance; Amino Acid Transport; GadC; Glutamate; Glutamine; Membrane Antiporter; Membrane Proteins; Membrane Transporter Reconstitution; Protonation/Deprotonation; Substrate Selectivity