Computational Analysis of Energy Landscapes Reveals Dynamic Features That Contribute to Binding of Inhibitors to CFTR-Associated Ligand

Abstract

The CFTR-associated ligand PDZ domain (CALP) binds to the cystic fibrosis transmembrane conductance regulator (CFTR) and mediates lysosomal degradation of mature CFTR. Inhibition of this interaction has been explored as a therapeutic avenue for cystic fibrosis. Previously, we reported the ensemble-based computational design of a novel peptide inhibitor of CALP, which resulted in the most binding-efficient inhibitor to date. This inhibitor, kCAL01, was designed using OSPREY and evinced significant biological activity in in vitro cell-based assays. Here, we report a crystal structure of kCAL01 bound to CALP and compare structural features against iCAL36, a previously developed inhibitor of CALP. We compute side-chain energy landscapes for each structure to not only enable approximation of binding thermodynamics but also reveal ensemble features that contribute to the comparatively efficient binding of kCAL01. Finally, we compare the previously reported design ensemble for kCAL01 vs the new crystal structure and show that, despite small differences between the design model and crystal structure, significant biophysical features that enhance inhibitor binding are captured in the design ensemble. This suggests not only that ensemble-based design captured thermodynamically significant features *

Computational Engineering of Protein L to Achieve an Optimal Affinity Chromatography Resin for Purification of Antibody Fragments

“…Osprey software was used to predict mutations in the B domain to increase binding affinity. , In the first B domain–Fab interface, the best score was predicted for conformation mutations Lys824Arg, Ilu834Leu, Ala837Thr, Lys840Arg, and Leu856Trp with the binding energy of −2603.446 kcal mol –1 . In the second B domain–Fab interface, the lowest-energy predicted conformation with the top-ranked mutations included Phe843Tyr, Thr847Met, Ala848Arg, Thr865Trp, and Gly871Trp with the binding energy of −3006.986 kcal mol –1 .…”

“…It has been shown that mutation prediction using Ospery not only led to an increase in the binding affinity but also increased the protein stability. , Lys824Arg and Lys840Arg mutations in the first binding site and Phe843Tyr and Thr847Met mutations in the second binding site predicted improvement of the protein stability. Sokalingam et al reported that the lysine to arginine mutagenesis could increase the mutant protein stability in the presence of chemical denaturants such as urea, alkaline pH, and ionic detergents .…”

Specificity in PDZ-peptide interaction networks: Computational analysis and review

“…In general, we and others are attempting to use available sequence and structural information to predict binding affinities with the goal of identifying cellular interaction networks (e.g., refs. Gerek et al, 2009;Gerek and Ozkan, 2010;Gfeller et al, 2011;Holt et al, 2019;Landgraf et al, 2004;te Velthuis et al, 2011;Tian et al, 2011;Valgardson et al, 2019). Understanding structural flexibility is important to these studies (Roberts et al, 2012;Thomas et al, 2009).…”

Proteostasis Regulators in Cystic Fibrosis: Current Development and Future Perspectives

“… 174 Structure and energy landscape analysis of the crystal structure of 41 :CALP (PDB ID: 6OV7) showed that the tighter binding efficiency of 41 could stem from entropic effect at P0 and favorable substitutions at P-1 and P-4 with long polar and charged residues (from Ile and Pro of 40 to Arg and Gln of 41 , respectively). 176 …”

“…Furthermore, 41 restored F508del-CFTR-mediated chloride efflux in CFBE cells in Ussing Chamber experiments, similarly to the previously available inhibitor F *- 40 or to the corrector 5 . Structure and energy landscape analysis of the crystal structure of 41 :CALP (PDB ID: 6OV7) showed that the tighter binding efficiency of 41 could stem from entropic effect at P0 and favorable substitutions at P-1 and P-4 with long polar and charged residues (from Ile and Pro of 40 to Arg and Gln of 41 , respectively) …”