Chapter 9 : Towards an Understanding of Membrane Channels

(Left) An AQP monomer shown in cartoon representation. The reentrant loops described in the text are very important structural elements in AQP's architecture. They are shown in a darker shade and are surrounded by the helical bundle formed by transmembrane helices of the protein. (Right) Hydrogen bonds between the two NPA motifs are essential for the stability of the structure and for the function of AQPs. Due to two stable hydrogen bonds between the amino group of the asparagine side chain of each NPA motif with neighboring side chains, one of the amino hydrogens of the asparagine is restrained to be fully exposed toward the interior of the channel, where it forms hydrogen bonds with the permeating substrate. Hydrogen bonds between the two NPA motifs are also important for the stability of the two reentrant loops.

Comparison of ribitol (a) and arabitol (b) at the selectivity filter of GlpF. Hydrogen bonds are shown as dotted lines. Ribitol is able to form an optimal number of hydrogen bonds with the channel without losing its linear conformation in this region. Formation of the same number of hydrogen bonds for arabitol requires a tilted conformation of the molecule that is unfavorable due to strong steric hindrance of the filter region.

Illustration of the method to produce a pressure gradient across the membrane through forces, shown by small arrows, applied to individual water molecules in the bulk region. Either all or only some water molecules may be selected for force application. The total pressure difference,ΔP, is determined by the number of water molecules, n; the applied force on each water molecule, f ; and the area of the membrane, A. In this case, P drives the water downward.

(a) View of the ClC dimer showing the broken helix architecture and the position of the Cl− ions in the crystal structure. Each monomer and pair of ions is displayed in a different shade. (b) Vertical cross-section of the solvent-accessible surface of the ClC protein embedded in a lipid bilayer. The simulated model comprises 97,000 atoms. In the narrowest part of the protein, where the Cl− ions permeate, the residues that define the selectivity filter are shown.

Detailed view of residues forming the ClC selectivity filter. The locations of hydrophobic residues are indicated as balls, whereas polar and charged residues are drawn explicitly. The positions of the Cl− ions as they permeate across the selectivity filter are plotted as small spheres, and the locations of the three Cl−-binding sites (Sint, Scen, and Sext) identified by X-ray crystallography ( Dutzler et al., 2003 ) are indicated as large circles.

Map of the PMF for a pair of Cl− ions moving across one of ClC's two pores as a function of the independent positions of the top and bottom Cl− ion. The thick line represents the coordinated motion of the pair of ions that follows the path of minimum energy (i.e., the most probable path). Each contour represents an energy difference of 1 kcal/mol. The axes correspond to the distance of the top and bottom permeating ions along a line perpendicular to the membrane, with the three binding sites from the crystal structure indicated as Sin, Scen, and Sext for reference.

Sequence of motion of two ions in the ClC pore, resulting in the conduction of one Cl− ion across ClC, as inferred from the calculated PMF.

Current-voltage characteristics of alpha-hemolysin computed with MD. Each data point is derived from a 288,680-atom simulation of the system shown in Fig. 8 . The dashed line indicates the linear fit through the data points at 120 and 240 mV and the origin. In accordance with experimental studies ( Menestrina, 1986 ; Krasilnikov and Sabirov, 1989 ), the I-V curve is sublinear at V 0. The absolute value of the ionic current at 120 mV is also in good agreement with experiment ( Meller and Branton, 2002 ). The inset shows cumulative currents through alpha-hemolysin at 120 and 240 mV. The table shows the number of ion permeations computed by dividing the total charge transported through the alphahemolysin pore by e, the unitary charge (1.6 ×10−19 C).

Microscopic model of the alphahemolysin channel in its native environment, a lipid bilayer membrane. The channel is drawn as a molecular surface separating the protein from the membrane and water. This surface is cut by the plane normal to the lipid bilayer passing through the geometrical center of the protein. All atoms but phosphorus of the dipalmitoyl phosphatidylcholine lipid bilayer are shown as lines; the phosphorus atoms are shown as spheres. Water and ions are not shown. The model comprises 288,680 atoms.

Computing the osmotic permeability of alpha-hemolysin with MD. (Top) Collective coordinate of all water molecules inside the channel n(t) ( equation 2 ) versus time; n(t) quantifies the net amount of water permeation through the channel (see the text). Neither the sign nor the magnitude of the transmembrane potential has a noticeable deterministic effect on water permeability. (Bottom) Mean square displacement of n(t) versus time. The slope of the curve yields the collective diffusion constant of water at 310/ns, which gives, after taking into account a correction for the low viscosity of TIP3P water, the osmotic permeability for alpha-hemolysin of 1.9 ×10−12cm3/s (see text).