Abstract
Inhibition of the insulin-regulated aminopeptidase (IRAP) improves memory and cognition in animal models. The enzyme has recently been crystallized and several series of inhibitors reported. We herein focused on one series of benzopyran-based inhibitors of IRAP known as the HFI series, with unresolved binding mode to IRAP, and developed a robust computational model to explain the structure-activity relationship (SAR) and potentially guide their further optimization. The binding model here proposed places the benzopyran ring in the catalytic binding site, coordinating the Zn2+ ion through the oxygen in position 3, in contrast to previous hypothesis. The whole series of HFI compounds was then systematically simulated, starting from this binding mode, using molecular dynamics and binding affinity estimated with the linear interaction energy (LIE) method. The agreement with experimental affinities supports the binding mode proposed, which was further challenged by rigorous free energy perturbation (FEP) calculations. Here, we found excellent correlation between experimental and calculated binding affinity differences, both between selected compound pairs and also for recently reported experimental data concerning the site directed mutagenesis of residue Phe544. The computationally derived structure-activity relationship of the HFI series and the understanding of the involvement of Phe544 in the binding of this scaffold provide valuable information for further lead optimization of novel IRAP inhibitors.
Highlights
Insulin-regulated aminopeptidase (IRAP, EC 3.4.11.3), known as leucyl-cystinyl aminopeptidase, placental leucine aminopeptidase, and oxytocinase is a transmembrane zinc metalloenzyme that belongs to the M1 family of aminopeptidases (Lew et al, 2003)
The crystal structure of human insulin-regulated aminopeptidase (IRAP) was retrieved from the protein data bank (PDB code 4PJ6) (Hermans et al, 2015), and monomer A from the crystal dimer was retained for docking and molecular dynamics (MD) simulations
The same MD sampling used in the previous stage was replicated 10 times per ligand-IRAP complex, accompanied by the same MD sampling of each ligand in a similar water sphere, all of which was used as a basis to calculate the binding free energy with the linear interaction energy (LIE) method (Aqvist et al, 1994)
Summary
Insulin-regulated aminopeptidase (IRAP, EC 3.4.11.3), known as leucyl-cystinyl aminopeptidase, placental leucine aminopeptidase, and oxytocinase is a transmembrane zinc metalloenzyme that belongs to the M1 family of aminopeptidases (Lew et al, 2003). Inhibition of IRAP by Benzopyranes (Albiston et al, 2007), and regulation of oxytocin and vasopressin levels in the brain (Chai et al, 2004; Stragier et al, 2008; Hattori and Tsujimoto, 2013). It has been shown that inhibiting IRAP with Ang IV (1, Figure 1) and other structurally related peptidomimetics like HA08 (2) (Diwakarla et al, 2016b) is linked with improved memory and learning in vivo (Braszko et al, 1988; Wright et al, 1993, 1996, 1999; O’Malley et al, 1998; De Bundel et al, 2009; Fu et al, 2012), including enhancement of dendritic spine density (DSD) exerted by HA08 in hippocampal cells (O’Malley et al, 1998; Fu et al, 2012), as well as drug mitigation and lesion-induced memory deficits in rodents (Vauquelin et al, 2002; Albiston et al, 2003; Chai et al, 2004). It was demonstrated that HFI compounds, exemplified by HFI-419 (8), enhance spatial working memory possibly by promoting the formation of functional dendritic spines by facilitating GLUT4-mediated glucose uptake into hippocampal neurons (Seyer et al, 2020)
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