Membrane proteins assemble and form stable complexes in the two-dimensional solvent structure of the lipid bilayer. Here, proteins encounter a unique problem where the chemical composition of the solvent is diverse and always changing under physiological pressures. The question arises, how do changes in lipid composition affect the free energy of membrane protein assembly in membranes? Previously, we experimentally determined that the CLC-ec1 Cl-/H+ antiporter participates in an equilibrium dimerization reaction in a synthetic mimic of the E. coli membrane - 2:1 POPE/POPG comprised of C16:0/C18:1 acyl chains (Chadda et al., eLife 2016). In this condition, the free energy of dimerization is measured to be -11 kcal/mole, one of the strongest complexes studied so far in membranes. Examination of the dimerization interface shows that the monomer introduces a region of hydrophobic mismatch that is ~10 Å shorter than the surrounding membrane. In turn, C16:0/C18:1 lipids adopt constricted configurations at this interface, resulting in an energetic penalty for the dissociated state. Based on this, we hypothesize that addition of short chain lipids, e.g. 2:1 DLPE/DLPG with C12:0 acyl chains, will stabilize the monomer by reducing the energetic penalty from lipid solvation. Small-angle neutron scattering experiments were conducted on mixed DL/PO membranes, showing a reduction in bilayer thickness as the DL proportion increases from 0-100%. The dimerization free energy of CLC was measured in the same membranes, showing a shift in stability to the monomeric state that occurs in two phases: (1) a gradual change at low DL (&lt; 1 %) and (2) an abrupt shift at high DL % that correlates with the decrease in bilayer thickness. Chloride transport function was preserved in short-chain membranes, confirming that the protein was folded. Coarse-grained molecular dynamics simulations of mixed DL/PO membranes shows an accumulation of short-chain lipids around the shortened dimerization interface, with a preferential sequestering of the negatively charged DLPG in the inner leaflet. Altogether, these results indicate that short-chain lipids preferentially solvate the dimerization interface and shift the stability of membrane protein association in lipid bilayers. The nature of this effect suggests a physical basis for tuning protein interactions in membranes that biology may take advantage of when physiologically regulating the membrane composition.

Tuning the stability of membrane protein dimerization by changing the lipid solvent

256th National Meeting (2018)

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