Molecular dynamics simulation of the neuroglobin crystal: comparison with the simulation in solution

Abstract

Neuroglobin (Ngb) is a monomeric protein that, despite the small sequence similarity with other globins, displays the typical globin fold. In the absence of exogenous ligands, the ferric and the ferrous forms of Ngb are both hexacoordinated to the distal and proximal histidines. In the ferrous form, oxygen, nitric oxide or carbon monoxide can displace the distal histidine, yielding a reversible adduct. Crystallographic data show that the binding of an exogenous ligand is associated to structural changes involving heme sliding and a topological reorganization of the internal cavities. Molecular dynamics (MD) simulations in solution show that the heme oscillates between two positions, much as the ones observed in the crystal structure, although the occupancy is different. The simulations also suggest that ligand binding in solution can affect the flexibility and conformation of residues connecting the C and D helices, referred to as the CD corner, which is coupled to the configuration adopted by the distal histidine. In this study, we report the results of 30 ns MD simulations of CO-bound Ngb in the crystal. Our goal was to compare the protein dynamical behavior in the crystal with the results supplied by the previous MD simulation of CO-bound Ngb in solution and the x-ray experimental data. The results show that the different environments (crystal or solution) affect the dynamics of the heme group and of the CD corner.

FIGURE 1

Sample configuration of NgbCO as extracted from MD simulations in solution. The heme, distal His⁶⁴(E7) and proximal His⁹⁶(F8) are depicted by sticks. The CD corner, composed of helices C and D and the CD loop, is highlighted.

FIGURE 2

Heme displacement distributions along its first essential eigenvector in carboxy Ngb in the crystal (solid line) and in solution (dashed line). The values corresponding to the experimental heme positions in carboxy Ngb (−0.72 nm) and metNgb (0.13 nm) in the crystal are indicated by vertical bars. Note that, with a single value in the abscissa, the distribution converges to a Dirac function.

FIGURE 3

Mean force with their standard deviations (upper panel) and heme group free energy landscape (bottom panel) for carboxy Ngb in the crystal (solid line) and in solution (dashed line). In brief, the method is based on the choice of a direction and on the calculation of the mean force acting on the heme center of mass along this direction. The mean force corresponds to the free energy gradient.

FIGURE 4

Heme displacement distributions along its first essential eigenvector in the carboxy Ngb crystal. The distributions are shown for each protein molecule in the crystal unit cell (–18). The vertical lines represent the principal configurations detected in the crystal (dashed line) and in solution (dotted lines).

FIGURE 5

Distribution of the projection onto the CD corner first essential eigenvector for carboxy Ngb in the crystal (lower panel) and in solution (upper panel). The distribution corresponding to the distal His64(E7) “closed” (solid line) or “open” (dashed line) configurations are shown. The value corresponding to the experimental CD corner position in carboxy Ngb (0.20 nm) is indicated by a vertical bar.

FIGURE 6

Distribution of the projection onto the CD corner first essential eigenvector for each protein molecule in the crystal unit cell (–18). The vertical dashed lines represent the projection values corresponding to the CD corner configurations detected in solution (a and b) and in the crystal structure (c).

FIGURE 7

CD corner interactions in the crystal unit cell. The CD corner (indicated by X) is surrounded by three symmetry-related protein molecules (marked by A, B, and C) whose positions correspond to as many symmetry transformations (see text). The protein “A” interacts with the CD corner through the helices G and H and the BC loop, the protein “B” through the CD loop, and the protein “C” through the helices B and G.