Molecular mechanism of secreted amyloid-β precursor protein in binding and modulating GABABR1a

Abstract

A recent phenomenal study discovered that the extension domain of secreted amyloid-β precursor protein (sAPP) can bind to the intrinsically disordered sushi 1 domain of the γ-aminobutyric acid type B receptor subunit 1a (GABA_BR1a) and modulate its synaptic transmission. The work provided an important structural foundation for the modulation of GABA_BR1a; however, the detailed molecular interaction mechanism, crucial for future drug design, remains elusive. Here, we further investigated the dynamical interactions between sAPP peptides and the natively unstructured sushi 1 domain using all-atom molecular dynamics simulations, for both the 17-residue sAPP peptide (APP 17-mer) and its minimally active 9 residue segment (APP 9-mer). We then explored mutations of the APP 9-mer with rigorous free energy perturbation (FEP) calculations. Our in silico mutagenesis studies revealed key residues (D4, W6, and W7) responsible for the binding with the sushi 1 domain. More importantly, one double mutation based on different vertebrate APP sequences from evolution exhibited a stronger binding (ΔΔG = -1.91 ± 0.66 kcal mol^-1), indicating a potentially enhanced GABA_BR1a modulator. These large-scale simulations may provide new insights into the binding mechanism between sAPP and the sushi 1 domain, which could open new avenues in the development of future GABA_BR1a-specific therapeutics.

Fig. 1. Sushi 1 domain of GABA_BR1a binding with the APP 17-mer peptide (A) and APP 9-mer peptide (B). The APP 17-mer peptide and APP 9-mer peptide are represented by sticks, while the sushi 1 domain is shown in cartoon and surface representations. In (B), non-polar residues are colored white, polar uncharged residues green, positively charged residues blue, and negatively charged residues red. (C) APP 17-mer–sushi 1 domain average residue contact probabilities (black) and average contact area ratios (blue) extracted from the MD simulation. (D) Per-residue average interaction energy between the APP 17-mer peptide and the sushi 1 domain.

Fig. 2. Structural fluctuations and key binding residues for the sAPP–GABA_BR1a complexes. (A) RMSD of the backbone atoms of APP 17-mer (green), APP 9-mer (blue) and the first nine residues of APP 17-mer (red) with respect to the reference structure at t = 0. (B) Time-dependent end-to-end distances of the APP 17-mer, APP 9-mer, and the first nine residues of APP 17-mer peptide. (C) APP 9-mer–sushi 1 binding interface, APP 9-mer and sushi 1 domain are represented by orange and purple NewCartoon, respectively. The contact residues are highlighted as sticks.

Fig. 3. Structural comparison of sushi 1 domain bound to WT APP 9-mer (left) and mutated APP 9-mer's (right) at the end of 300 ns of MD simulation: (A) for D4A mutation; (B) for W6A; (C) for W7A.

Fig. 4. RMSD of the backbone atoms of APP 9-mer (red) and sushi 1 domain (black) for WT APP 9-mer (A) and APP 9-mer's with mutations D4A (B), W6A (C), and W7A (D). Final structure of sushi 1 domain binding to WT and mutated APP 9-mer's (E–H). APP 9-mer is colored in orange and sushi 1 domain in gray. The shallow binding groove of sushi 1 is marked by the red circle.

Fig. 5. The binding complex structural changes due to the double mutation D1AD2N of APP 9-mer. All the sushi 1 domain residues within 6 Å from the first two APP 9-mer residues are shown in sticks, with the rest shown in ribbon view. The figure clearly shows the strong repulsion between D1 and D2 in the wild-type, with both D1 and D2's negatively charged sidechains sticking into solvent (left). On the other hand, the double mutant A1N2 displays a backbone hydrogen bond between N2 and Gly22, and meanwhile, both A1 and N2 residues are pushed closer to the sushi 1 domain (right).