Synergistic substrate binding determines the stoichiometry of transport of a prokaryotic H(+)/Cl(-) exchanger

Nat Struct Mol Biol. 2012 Apr 8;19(5):525-31, S1. doi: 10.1038/nsmb.2277.

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.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Chloride Channels / genetics
  • Chloride Channels / metabolism*
  • Chlorides / metabolism
  • Chlorine / metabolism*
  • Escherichia coli / genetics
  • Escherichia coli / metabolism*
  • Escherichia coli Proteins / genetics
  • Escherichia coli Proteins / metabolism*
  • Models, Molecular
  • Mutation
  • Protein Binding
  • Protons*
  • Substrate Specificity
  • Thermodynamics

Substances

  • Chloride Channels
  • Chlorides
  • ClC protein, E coli
  • Escherichia coli Proteins
  • Protons
  • Chlorine