Tuning the Stability of Membrane Protein Dimerization by Changing the Lipid Solvent

Abstract

In cell membranes, proteins encounter a unique problem where the solvent is a mixture of diverse chemical composition. The question arises, how do changes in lipid composition affect a simple reaction such as protein association? Previously, we experimentally measured the equilibrium dimerization reaction of the CLC-ec1 Cl−/H+antiporter in a synthetic mimic of the E. colimembrane - 2:1 POPE/POPG comprised of C16:0/C18:1 acyl chains (Chadda et al., eLife 2016). In this condition, the free energy of association is −11 kcal/mole, one of the strongest membrane protein complexes studied so far. Examination of the dimerization interface shows that the monomer introduces a region of hydrophobic mismatch ∼10 Å shorter than the surrounding membrane. Coarse-grained molecular dynamics (MD) simulations show that C16:0/C18:1 lipids adopt constrained configurations at the dimerization interface that introduce an energetic penalty in the dissociated state. Based on this, we hypothesized that short chain lipids, e.g. 2:1 DLPE/DLPG with C12:0 acyl chains, could solvate the shorter interface without penalty, and thus stabilize the dissociated state. Chloride transport function was measured in the DL membranes demonstrating that the protein is folded. On the other hand, dimerization stability is shifted, with a gradual increase in monomeric population for DL < 1%, and a complete shift to monomers at higher DL. Small-angle neutron scattering experiments were carried out on the mixed DL/PO membranes, showing a reduction in bilayer thickness from 0 - 40% DL that correlates strongly with the changes in dimerization. MD simulations of mixed PO/DL membranes show preferential accumulation of short-chain lipids along the dimerization interface. Altogether, these results indicate that short-chain lipids modulate CLC dimer stability by local hydrophobic matching, with preferential solvation capable of tuning the reaction across a wide-range of compositions.