Forced detachment of the CD2-CD58 complex - PubMed (original) (raw)

Comparative Study

Forced detachment of the CD2-CD58 complex

M V Bayas et al. Biophys J. 2003 Apr.

Abstract

The force-induced detachment of the adhesion protein complex CD2-CD58 was studied by steered molecular dynamics simulations. The forced detachment of CD2 and CD58 shows that the system can respond to an external force by two mechanisms, which depend on the loading rate. At the rapid loading rates of 70 and 35 pN/ps (pulling speeds of 1 and 0.5 A/ps) the two proteins unfold before they separate, whereas at slower loading rates of 7 and 3.5 pN/ps (pulling speeds of 0.1 and 0.05 A/ps), the proteins separate before the domains can unfold. When subjected to a constant force of 400 pN, the two proteins separated without significant structural distortion. These findings suggest that protein unfolding is not coupled to the adhesive function of CD2 and CD58. The simulations further confirm that salt bridges primarily determine the tensile strength of the protein-to-protein bond, and that the order of salt bridge rupture depends mainly on the position of the bond, relative to the line of action of the applied force. Salt bridges close to this line break first. The importance of each of the salt bridges for adhesion, determined from the simulations, correlates closely with their role in cell-to-cell adhesion and equilibrium binding determined by site-directed mutagenesis experiments.

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Figures

FIGURE 1

(a) Predicted structure of the adhesive CD2-CD58 complex. The Brookhaven Protein Data Bank entries 1HNF (Bodian et al., 1994) and 1CCZ (Ikemizu et al., 1999) were used to prepare the representations of CD2 and CD58 respectively. (b) Topology of the binding domains of CD2 and CD58. (c) Representation of the adhesive complex involving domains D1. This corresponds to the entry 1QA9 (Wang et al., 1999), with most of the charge amino acids in the binding interface in bond representation. All of the charged residues, except K91 and D33, form salt bridges. The different salt bridges are shown from the front (d), the left (e), and from the back (f). Several amino acids in the binding interface form multiple contacts. This is the case of CD2 K51, which forms salt bridges with CD58 E39 and CD58 E42 (d). CD2 R48 similarly forms salt bridges with CD58 E37 and CD58 E39 (f).

FIGURE 2

Evolution of the backbone RMSD as a function of time during the equilibration process, for SET 1 (a) and SET 2 (b) set of conditions.

FIGURE 3

Snapshots of the complex during the simulation performed at 1 Å/ps. The inset shows the corresponding force-extension profile. The charge residues in the binding region are highlighted using a VDW representation. The sequential rupture of the salt bridges and the partial unraveling of the proteins are visualized in these figures.

FIGURE 4

Force-extension profile and displacements at 1 Å/ps (a) and 0.5 Å/ps (b). In both cases the separation (Δ_z_ centers of mass) remains close to zero at the beginning, whereas the extension (Δ_z_) increases rapidly. This is accompanied by a uniform increase of the force until point a in both figures. After this period the main rupture events occur. At point b in Fig. 4 a, strand G of CD2 separates. This generates a plateau in the force profile. At point b in Fig. 4 b strand F of CD58 partially separates from strand C and at point c strand G separates completely. For both pulling speeds the complete separation of strands G of CD2 and CD58 occurs around the maximum of the force (point c in Fig. 4 a and point d in Fig. 4 b). The difference between the extension and separation at the end of the simulations are due primarily to the protein unraveling.

FIGURE 5

Snapshots and force-extension profile from the simulation at 0.05 Å/ps. The charged residues in the binding region are highlighted using a VDW representation. The structures show the sequential breaking of the salt bridges that lead to the detachment of the complex. After detachment the protein structures are only slightly deformed.

FIGURE 6

(a) Force-extension profile and displacements determined at 0.1 Å/ps. (b) Variation of the RMSDs of the protein backbones versus time. At point a, the force decreases due to the partial separation of the G strand of CD58. CD2 did not rupture. (c) Distance between salt bridge pairs versus time. The rupture of the salt bridges E95K32, D32K34, and R44E37 are associated with the peaks at points b, c, and d, respectively. (d) Distance between salt bridge pairs versus time. At points e and f, salt bridges K51E39 and D31R44 broke.

FIGURE 7

(a) Force-extension profile and displacements for 0.05 Å/ps. (b) Variation of the backbone RMSD of the proteins versus time. At point b the partial separation of the _G_-strand of CD58 caused a peak in the force profile. CD2 did not exhibit any abrupt rupture. (c) Distance between salt bridge pairs versus time. The rupture of the salt bridge D32K34 caused the force peak observed in point a. At point c the salt bridges E43E25 and D31K44 broke. (d) Distance between salt bridge pairs versus time. At point d, the salt bridge K51E39 broke, after which the proteins separated quickly.

FIGURE 8

Force (a) and extension (b) as a function of the spring displacement determined at the four pulling speeds for the SET 1 conditions.

FIGURE 9

Snapshots showing the formation and rupture of the salt bridge K91D33 in the FGCC′ region when the pulling speed was 0.1 Å/ps. The salt bridge formed at 300 ps. The neighboring salt bridge, E95K32, broke at 400 ps. The separation of the CD2-FC loop from the CD58-CC′ loop was possible after the rupture of this salt bridge at 522 ps.

FIGURE 10

Snapshots and separation profile determined from the simulation at a constant force of 400 pN. In this case, the detachment occurred in seven steps; most of them involved ruptures of single salt bridges. The snapshots show the sequential breakage of the salt bridges leading to the detachment of the complex. A and B provide views of the CD2 and CD58 complex from two different angles. Charged residues in the binding region are highlighted using a VDW representation.

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Forced detachment of the CD2-CD58 complex - PubMed (original) (raw)