Constant pH Molecular Dynamics Reveals pH-Modulated Binding of Two Small-Molecule BACE1 Inhibitors

Abstract

Targeting BACE1 with small-molecule inhibitors offers a promising route for treatment of Alzheimer’s disease. However, the intricate pH dependence of BACE1 function and inhibitor efficacy has posed major challenges for structure-based drug design. Here we investigate two, structurally similar BACE1 inhibitors that have dramatically different binding affinity and inhibitory activity using continuous constant pH molecular dynamics (CpHMD). At high pH, both inhibitors are stably bound to BACE1, however, within the enzyme active pH range, only the iminopyrimidinone-based inhibitor remains bound, while the aminothiazine-based inhibitor becomes partially dissociated following the loss of hydrogen bonding with the active site and change of the 10s loop conformation. The drastically lower affinity of the second inhibitor is due to the protonation of a catalytic aspartate and the lack of a propyne tail. This work demonstrates that CpHMD can be used for screening pH-dependent binding profiles of small-molecule inhibitors, providing a new tool for structure-based drug design and optimization.

Proton-Coupled Conformational Allostery Modulates the Inhibitor Selectivity for β-Secretase

“…We followed the double decoupling scheme of Boresch et al 20 (Figure 2) to calculate the absolute binding free energies of BACE1 and CatD to the inhibitor LY2811376 11,12 at the reference pH (pH 8). The free energy perturbation (FEP) method was used to compute $\mathrm{\Delta }{G}_{\text{dsolv}}^{\mathrm{normalL}},\mathrm{\Delta }{G}_{\text{coup}}^{\mathrm{normalC}}$, and $\mathrm{\Delta }{G}_{\text{res}}^{\mathrm{normalC}}$.…”

“…36 The proteins were modeled with the CHARMM22/CMAP force field, 37,38 and the inhibitor LY2811376 was modeled with the force field obtained previously by us. 11,12 The FEP protocols made use of 14–31 λ windows, and each window was run for 1 ns.…”

“…8,11,12 In these simulations, the Asp, Glu and His sidechains as well as the inhibitor pyrimidin nitrogen (Fig. 1, circled red) were allowed to titrate, while Cys and Tyr were kept neutral and Lys and Arg were kept charged.…”

“…11,12 For both BACE1 and CatD systems, except for the catalytic aspartic acids and inhibitor titratable site, most residues display small p K a shifts with absolute values below 0.2 units (Fig. 3a) and block standard errors of 0–0.2 units (Table S1 and S2).…”

Conformational dynamics of cathepsin D and binding to a small‐molecule BACE1 inhibitor

“…At high pH when the catalytic dyad is di‐deprotonated the conserved tyrosine of the flap forms a stable hydrogen bond with the dyad and prevents substrate access, while within the active pH range 3.5–5.5, protonation of one Asp residue destabilizes this interaction and a binding competent state becomes populated. Additionally, we demonstrated how subtle changes in the protonation state of the catalytic dyad as well as the inhibitor can significantly alter the strength of binding between inhibitor and the target enzyme …”