Mechanism of selectivity in aquaporins and aquaglyceroporins

Abstract

Aquaporins and aquaglyceroporins form a family of pore proteins that facilitate the efficient and selective flux of small solutes across biological membranes. We studied the selectivity of aquaporin-1 (AQP1) and the bacterial glycerol facilitator, GlpF, for O(2), CO(2), NH(3), glycerol, urea, and water. Using molecular dynamics simulations, we calculated potentials of mean force for solute permeation along the aquaporin channels and compared them with the alternative pathway across the lipid bilayer. For small solutes permeating through AQP1, a remarkable anticorrelation between permeability and solute hydrophobicity was observed, whereas the opposite trend was observed for permeation through the membrane. This finding renders AQP1 a selective filter for small polar solutes, whereas GlpF was found to be highly permeable for small solutes and permeable for larger solutes. Surprisingly, not solute-channel but water-channel interactions were found to be the key determinant underlying the selectivity mechanism of aquaporins. Hence, a hydrophobic effect, together with steric restraints, determines the selectivity of aquaporins.

Fig. 1.

Glycerol facilitator GlpF. (A) Snapshot of an MD simulation showing a single file of water inside the pore of GlpF. Some water-interacting residues are shown in stick representation. (B) Simulation box of a GlpF tetramer solvated in a membrane of POPE and water in side view (Left) and top view (Right). Glycerol molecules (displayed in sphere representation) are placed along the channel axes as starting configurations for umbrella sampling simulations.

Fig. 2.

Potentials of mean force G(z) for O₂, CO₂, NH₃, glycerol, and urea permeating through the monomeric water pores of human AQP1 (black curves), GlpF (red curves), and the H180A/R195V mutant of AQP1 (green curves). z = 0 corresponds to the center of the NPA motifs. The NPA region is highlighted by a blue bar, the aromatic/arginine constriction region by an orange bar. The red circles correspond to the glycerol positions in the GlpF crystal structure (6).

Fig. 3.

PMFs for permeation of O₂, CO₂, NH₃, H₂O, glycerol, and urea through membranes of POPE (Upper) and POPC (Lower). z = 0 corresponds to the membrane center.

Fig. 4.

Permeability as a function of hydrophobicity of the solute. (A and B) Free-energy barrier ΔG_max for urea (+), glycerol (inverted triangle), H₂O (triangle), NH₃ (diamond), CO₂ (square), and O₂ (circle) permeating along the pores of AQP1 (A) and GlpF (B). ΔG_max is plotted versus the logarithm of the hexadecane–water partition coefficient log K_hex of the solute, which is a common measure for hydrophobicity. (C) Solvation free-energy difference ΔG_tails between the solute in water and the solute in the hydrophobic tail region of the POPC membrane as determined by umbrella sampling simulations. The dashed line indicates the energetic cost ΔG_hex = −k_BT ln K_hex for moving the solute from bulk water into hexadecane.

Fig. 5.

Water–protein interaction as selectivity mechanism for aquaporins. Comparison between hAQP1 (Left) and GlpF (Right). (A) PMFs for O₂ permeating through the ar/R regions of hAQP1 and GlpF versus the position of O₂. (B) Interactions between water and the ar/R residues as a function of the position of a permeating O₂ molecule (magenta curves). In AQP1, water–ar/R interactions are reduced by ≈60 kJ/mol when an O₂ molecule is present in the ar/R region, and the loss of water–protein interaction cannot be compensated by O₂–protein interactions, which are displayed in C. In GlpF, water–ar/R interactions are hardly affected by the O₂. The water–ar/R interaction can be decomposed into interactions between water and single residues (blue, orange, green, and red curves). (D) Number of hydrogen bonds between water and protein residues versus the position of a permeating O₂ molecule. (E) MD snapshots of the ar/R regions of hAQP1 and GlpF, including several water molecules. The residues are colored according to the curves in B and D. Possible water–protein hydrogen bonds are denoted by dashed lines. The red arrow indicates the pore coordinate.