title: Synergistic substrate binding determines the stoichiometry of transport of a prokaryotic H(+)/Cl(-) exchanger.
abstract: Active exchangers dissipate the gradient of one substrate to accumulate nutrients, export xenobiotics and maintain cellular homeostasis. Mechanistic studies have suggested that two fundamental properties are shared by all exchangers: substrate binding is antagonistic, and coupling is maintained by preventing shuttling of the empty transporter. The CLC H(+)/Cl(-) exchangers control the homeostasis of cellular compartments in most living organisms, but their transport mechanism remains unclear. We show that substrate binding to CLC-ec1 is synergistic rather than antagonistic: chloride binding induces protonation of a crucial glutamate. The simultaneous binding of H(+) and Cl(-) gives rise to a fully loaded state that is incompatible with conventional transport mechanisms. Mutations in the Cl(-) transport pathway identically alter the stoichiometries of H(+)/Cl(-) exchange and binding. We propose that the thermodynamics of synergistic substrate binding, rather than the kinetics of conformational changes and ion binding, determine the stoichiometry of transport.
authors.affiliation: Department of Anesthesiology, Weill Cornell Medical College, New York, New York, USA.
authors.forename: Alessandra
authors.initials: A
authors.lastname: Picollo
authors.forename: Yanyan
authors.initials: Y
authors.lastname: Xu
authors.forename: Niklaus
authors.initials: N
authors.lastname: Johner
authors.forename: Simon
authors.initials: S
authors.lastname: Bernèche
authors.forename: Alessio
authors.initials: A
authors.lastname: Accardi
mesh_terms: D018118:Chloride Channels / Q000235:genetics / Q000378:metabolism*; D002712:Chlorides / Q000378:metabolism; D002713:Chlorine / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D009154:Mutation; D011485:Protein Binding; D011522:Protons; D013379:Substrate Specificity; D013816:Thermodynamics
chemical_list: D018118:Chloride Channels; D002712:Chlorides; C508370:ClC protein, E coli; D029968:Escherichia coli Proteins; D011522:Protons; D002713:Chlorine