Energy Landscapes of Ligand Motion Inside the Tunnel-Like Cavity of Lipid Transfer Proteins: The Case of the Pru p 3 Allergen

Abstract

Allergies are a widespread problem in western countries, affecting a large part of the population, with levels of prevalence increasingly rising due to reasons still not understood. Evidence accumulated in recent years points to an essential role played by ligands of allergen proteins in the sensitization phase of allergies. In this regard, we recently identified the natural ligand of Pru p 3, a lipid transfer protein, a major allergen from peach fruit and a model of food allergy. The ligand of Pru p 3 has been shown to play a key role in the sensitization to peach and to other plant food sources that provoke cross-reactivity in a large proportion of patients allergic to peach. However, the question of which is the binding pose of this ligand in its carrier protein, and how it can be transferred to receptors of the immune system where it develops its function as a coadjuvant was not elucidated. In this work, different molecular dynamics simulations have been considered as starting points to study the properties of the ligand⁻protein system in solution. Besides, an energy landscape based on collective variables that describe the process of ligand motion within the cavity of Pru p 3 was obtained by using well-tempered metadynamics. The simulations revealed the differences between distinct binding modes, and also revealed important aspects of the motion of the ligand throughout its carrier protein, relevant to its binding⁻unbinding process. Our findings are potentially interesting for studying protein⁻ligand systems beyond the specific case of the allergen protein dealt with here.

Keywords: allergy; enhanced sampling; lipid transfer proteins; metadynamics; molecular dynamics.

Figure 1

(A) Structural formulas of the ligands addressed in this work: the natural ligand of Pru p 3 (formed by the union of phytosphingosine and 10-hydroxy-camptothecin); phytosphingosine, and sphingosine. (B) Two possible orientations for the location of the ligand in the hydrophobic cavity (tunnel) of Pru p 3. The surface colored in red encloses the volume of a constant pocket inside the protein, obtained using MDPocket over molecular dynamics simulations.

Figure 2

RMSD of protein (backbone atoms) and ligands along 100 ns trajectories for the orientations A and B inside the cavity of Pru p 3 of the three ligands studied: the phytosphingosine tail of the natural ligand of Pru p 3 (“ligand” label), phytosphingosine, and sphingosine.

Figure 3

(A) Scheme of the collective variables (CVs) setup. (B) Rendering of the volume in which the ligand tail is confined using harmonic walls (see the text in Section 2.3).

Figure 4

CV1 and CV2 along the unbiased molecular dynamics trajectory for the three ligands in orientations A and B calculated only for aliphatic carbons. “Lig” = natural ligand of Pru p 3, “phs” = phytosphingosine, and “sph” = sphingosine.

Figure 5

Free energy landscape obtained by well-tempered metadynamics simulations along the collective variable CV1. Dotted lines mark the different basins detected.

Figure 6

Convergence of metadynamics simulations illustrated as energy deposition along time in the diffusion axis. Latest contributions added little energy to the landscape. (a) Energy landscape for the orientation A. (b) Ibid. for the orientation B.

Figure 7

Cavity volume for each position of the ligand along the diffusion axis during metadynamics simulations. Color of orientation A was changed for visualization purposes.