Nanosecond-timescale conformational dynamics of the human alpha7 nicotinic acetylcholine receptor

Abstract

We explore the conformational dynamics of a homology model of the human alpha7 nicotinic acetylcholine receptor using molecular dynamics simulation and analyses of root mean-square fluctuations, block partitioning of segmental motion, and principal component analysis. The results reveal flexible regions and concerted global motions of the subunits encompassing extracellular and transmembrane domains of the subunits. The most relevant motions comprise a bending, hinged at the beta10-M1 region, accompanied by concerted tilting of the M2 helices that widens the intracellular end of the channel. Despite the nanosecond timescale, the observations suggest that tilting of the M2 helices may initiate opening of the pore. The results also reveal direct coupling between a twisting motion of the extracellular domain and dynamic changes of M2. Covariance analysis of interresidue motions shows that this coupling arises through a network of residues within the Cys and M2-M3 loops where Phe135 is stabilized within a hydrophobic pocket formed by Leu270 and Ile271. The resulting concerted motion causes a downward shift of the M2 helices that disrupts a hydrophobic girdle formed by 9' and 13' residues.

FIGURE 1

Snapshot of the simulation system with the human α7 receptor (ribbon) inserted into the POPC lipid bilayer (licorice) and fully hydrated with water molecules (not shown).

FIGURE 2

(a) RMSFs of the C^α atoms from the MD simulation presented here (black line) along with those from a previous MD simulation (red line) and from the block normal mode analysis (green line). The color bar on the top of Fig. 2 a indicates sequence position of important structural elements; these structural elements are also labeled in Fig. 2 b. (b) The computed B-factor values are color-coded onto a single subunit of the receptor, with red corresponding to the most mobile region and blue corresponding to the most stable region. For clarity, the RMSFs are averaged over five subunits.

FIGURE 3

Global motions of the α7 receptor during the simulation, as suggested by the PCA analysis. The model structures are generated from some small displacements along (a) the first, (b) the second, and (c) the third principal modes. Some key regions are highlighted with red circles.

FIGURE 4

“Block” motion of the α7 receptor as suggested by the TLSMD analysis based on the simulation data. Fourteen blocks have been identified within a single α7 subunit, and are shown in different colors. The color bar on the bottom indicates the sequence information. (a) Front view, taken from the periphery of the receptor. (b) Rotated ∼180° from the position in a.

FIGURE 5

Correlated fluctuations of the C^α atoms in the α7 receptor, calculated from the MD simulation. The correlation maps are shown for correlated residues from (a) the A subunit, (b) the Cys-loop and the β10-M1 region, with the two primary regions of correlation highlighted by red circles, (c) the Cys-loop and the M2-M3 linker, with a coupled hydrophobic tetrad highlighted by a purple rectangular box, (d) the β10-M1 and the β8-β9 linker. The color bars on the right indicate the extent of the correlation. Residue pairs with a high level of correlated motions are shown in yellow, orange, and red. Anticorrelated motions are represented by the blue/cyan regions. Green indicates no correlation.

FIGURE 6

Correlated motion of Phe¹³⁵ (located in the Cys-loop) and Ile²⁷¹ (located in the M2-M3 linker) during the MD simulation. (a) Distance between Phe¹³⁵ and Ile²⁷¹ as a function of time. (b) Averaged z axis position of the M2 helix as a function of time. (c and d) Detailed views of the membrane interface region in the initial structure (c) and in a snapshot after 6 ns of MD simulation (d).

FIGURE 7

(a) Minimum pore radius and (b) z axis position of the minimum pore radius as a function of time during the MD simulation (with the conventional numbering for M2 residues depicted on the right axis). The center of the pore domain is set to zero, with positive z values toward the extracellular end.

FIGURE 8

(a and b) Schematic representations of lateral tilting θ₁ and radial tilting θ₂. See Methods section for the definitions of these two angles. (c) Lateral tilting and (d) radial tilting motions of the five M2 helices as a function of time during the MD simulation.

FIGURE 9

Isosurfaces of time-averaged water density during the simulation. The surface corresponds to the isodensity contour, ∼0.4 of the bulk water density. The receptor structure is shown in cartoon representation, with each subunit colored differently.

FIGURE 10

Detailed view of two hydrophobic residues, Leu⁹′ (248) and Val¹³′ (252), (a) in the initial structure, and (b) in a snapshot after 10 ns of MD simulation.