The dimerization equilibrium of a ClC Cl−/H+ antiporter in lipid bilayers
نویسندگان
چکیده
Interactions between membrane protein interfaces in lipid bilayers play an important role in membrane protein folding but quantification of the strength of these interactions has been challenging. Studying dimerization of ClC-type transporters offers a new approach to the problem, as individual subunits adopt a stable and functionally verifiable fold that constrains the system to two states - monomer or dimer. Here, we use single-molecule photobleaching analysis to measure the probability of ClC-ec1 subunit capture into liposomes during extrusion of large, multilamellar membranes. The capture statistics describe a monomer to dimer transition that is dependent on the subunit/lipid mole fraction density and follows an equilibrium dimerization isotherm. This allows for the measurement of the free energy of ClC-ec1 dimerization in lipid bilayers, revealing that it is one of the strongest membrane protein complexes measured so far, and introduces it as new type of dimerization model to investigate the physical forces that drive membrane protein association in membranes.
منابع مشابه
A model-free method for measuring dimerization free energies of CLC-ec1 in lipid bilayers.
The thermodynamic reasons why membrane proteins form stable complexes inside the hydrophobic lipid bilayer remain poorly understood. This is largely because of a lack of membrane-protein systems amenable for equilibrium studies and a limited number of methods for measuring these reactions. Recently, we reported the equilibrium dimerization of the CLC-ec1 Cl-/H+ transporter in lipid bilayers (Ch...
متن کاملFluoride-dependent interruption of the transport cycle of a CLC Cl−/H+ antiporter
Cl(-)/H(+) antiporters of the CLC superfamily transport anions across biological membranes in varied physiological contexts. These proteins are weakly selective among anions commonly studied, including Cl(-), Br(-), I(-), NO3(-) and SCN(-), but they seem to be very selective against F(-). The recent discovery of a new CLC clade of F(-)/H(+) antiporters, which are highly selective for F(-) over ...
متن کاملIonic Currents Mediated by a Prokaryotic Homologue of CLC Cl− Channels
CLC-ec1 is an E. coli homologue of the CLC family of Cl- channels, which are widespread throughout eukaryotic organisms. The structure of this membrane protein is known, and its physiological role has been described, but our knowledge of its functional characteristics is severely limited by the absence of electrophysiological recordings. High-density reconstitution and incorporation of crystall...
متن کاملExtracellular Determinants of Anion Discrimination of the Cl−/H+ Antiporter Protein CLC-5*
Mammalian CLC proteins comprise both Cl- channels and Cl-/H+ antiporters that carry out fundamental physiological tasks by transporting Cl- across plasma membrane and intracellular compartments. The NO3- over Cl- preference of a plant CLC transporter has been pinpointed to a conserved serine residue located at Scen and it is generally assumed that the other two binding sites of CLCs, Sext and S...
متن کاملF−/Cl− selectivity in CLCF-type F−/H+ antiporters
Many bacterial species protect themselves against environmental F(-) toxicity by exporting this anion from the cytoplasm via CLC(F) F(-)/H(+) antiporters, a subclass of CLC superfamily anion transporters. Strong F(-) over Cl(-) selectivity is biologically essential for these membrane proteins because Cl(-) is orders of magnitude more abundant in the biosphere than F(-). Sequence comparisons rev...
متن کامل