Similarity of force-induced unfolding of apomyoglobin to its chemical-induced unfolding: an atomistic molecular dynamics simulation approach

Abstract

We have compared force-induced unfolding with traditional unfolding methods using apomyoglobin as a model protein. Using molecular dynamics simulation, we have investigated the structural stability as a function of the degree of mechanical perturbation. Both anisotropic perturbation by stretching two terminal atoms and isotropic perturbation by increasing the radius of gyration of the protein show the same key event of force-induced unfolding. Our primary results show that the native structure of apomyoglobin becomes destabilized against the mechanical perturbation as soon as the interhelical packing between the G and H helices is broken, suggesting that our simulation results share a common feature with the experimental observation that the interhelical contact is more important for the folding of apomyoglobin than the stability of individual helices. This finding is further confirmed by simulating both helix destabilizing and interhelical packing destabilizing mutants.

FIGURE 1

The equilibration of the initial structure using the Nose-Hoover thermostat at 300 K for 1 ns. The change of helices with time is plotted every 4 ps by representing the helix forming residue as a black dot. The alphabetical order on the y axis of the right side represents the eight helices of apomyoglobin and the solid lines divide the region of each helix.

FIGURE 2

The averaged force-extension profile simulated under the distance restraint. The r_NC in the plot denotes the distance between the N atom of Val¹ and the C atom of Gly¹⁵³. The biasing potential is applied to the two atoms to increase the distance between the two terminal atoms at a stretching rate of 0.5 Å/ps. The averaged end-to-end distance of the initial structure is 25.9 Å.

FIGURE 3

The changes of helix structure and external force under the distance restraint with time. The black dots and solid lines in this plot have the same meaning as in Fig. 1.

FIGURE 4

The solvent-accessible surface area of Trp⁷ and Trp¹⁴ residues in the A-helix. Note that these hydrophobic residues of the A-helix in the native state are buried in the hydrophobic core consisting of the AGH helices.

FIGURE 5

The disruption of hydrophobic core. (a) The change of external force and axis angles between helices as a function of time during stretching. The axis angle is defined as the angle between the two helices vectors that corresponds to the vector from the N-terminus atom to the C-terminus atom in these two helices. (b) Hydrogen bond distance between Ile⁹⁹ in the G-helix and Tyr¹⁴⁶ in the H-helix. (c) The packing between the G (white) and H (black) helices in the native state. The fragment of the GH hairpin is drawn only using C_α of the backbone chain.

FIGURE 6

The contact maps and their corresponding snapshots of force-induced unfolding of apomyoglobin under distance restraint (a) at 200 ps, r_NC = 99 Å, (b) at 400 ps, r_NC = 197 Å, and (c) at 500 ps, r_NC = 268 Å. In the contact maps, two nonconsecutive residues are considered to make a contact if any side-chain atoms are within 7 Å of each other. For comparison, the contact map for the native state is represented by open squares and those for the snapshot of the force-induced unfolding simulation are represented by filled circles. All snapshots are visualized using WebLab ViewerPro4.0 of Accelrys Inc. (San Diego, CA). In the snapshots, two black ribbons correspond to the G and H helices.

FIGURE 7

The change of end-to-end distance between the N atom of Val¹ and the C atom of Gly¹⁵³ as a function of time when a constant force is applied to the terminal atoms.

FIGURE 8

Snapshots of force-induced unfolding of apomyoglobin under constant force applied to the N atom of Val¹ and the C atom of Gly¹⁵³. Two black ribbons correspond to the G and H helices. (a) 200 pN, r_NC = 143 Å at 100 ps; (b) 100 pN, r_NC = 117 Å at 1000 ps; (c) 100 pN, r_NC = 135 Å at 2000 ps; (d) 50 pN, r_NC = 72 Å at 1400 ps; (e) 50 pN, r_NC = 103 Å at 1600 ps. All snapshots are visualized using WebLab ViewerPro4.0 of Accelrys Inc.

FIGURE 9

Unfolding simulation using a biasing force for increasing the radius of gyration from 16 Å to 21 Å at a constant rate of 0.005 Å/ps. (a) The change of the radius of gyration (R_g) as a function of time. The dotted lines indicate the region where the value of R_g abruptly increases. (b) The snapshot at 800 ps. (c) The snapshot at 1200 ps. All snapshots are visualized using WebLab ViewerPro4.0 of Accelrys Inc.

FIGURE 10

Force-induced unfolding for two types of mutants under the constant force of 200 pN applied to the N atom of Val¹ and the C atom of Gly¹⁵³. The mutation at L115A, F123A, and L135A is used for destabilizing interhelical packing and the mutation at N132G and E136G is used for destabilizing H-helix.