title: Computational design of a PDZ domain peptide inhibitor that rescues CFTR activity.
abstract: The cystic fibrosis transmembrane conductance regulator (CFTR) is an epithelial chloride channel mutated in patients with cystic fibrosis (CF). The most prevalent CFTR mutation, ΔF508, blocks folding in the endoplasmic reticulum. Recent work has shown that some ΔF508-CFTR channel activity can be recovered by pharmaceutical modulators ("potentiators" and "correctors"), but ΔF508-CFTR can still be rapidly degraded via a lysosomal pathway involving the CFTR-associated ligand (CAL), which binds CFTR via a PDZ interaction domain. We present a study that goes from theory, to new structure-based computational design algorithms, to computational predictions, to biochemical testing and ultimately to epithelial-cell validation of novel, effective CAL PDZ inhibitors (called "stabilizers") that rescue ΔF508-CFTR activity. To design the "stabilizers", we extended our structural ensemble-based computational protein redesign algorithm K* to encompass protein-protein and protein-peptide interactions. The computational predictions achieved high accuracy: all of the top-predicted peptide inhibitors bound well to CAL. Furthermore, when compared to state-of-the-art CAL inhibitors, our design methodology achieved higher affinity and increased binding efficiency. The designed inhibitor with the highest affinity for CAL (kCAL01) binds six-fold more tightly than the previous best hexamer (iCAL35), and 170-fold more tightly than the CFTR C-terminus. We show that kCAL01 has physiological activity and can rescue chloride efflux in CF patient-derived airway epithelial cells. Since stabilizers address a different cellular CF defect from potentiators and correctors, our inhibitors provide an additional therapeutic pathway that can be used in conjunction with current methods.
authors.affiliation: Department of Computer Science, Duke University, Durham, North Carolina, United States of America.
authors.forename: Kyle E
authors.initials: KE
authors.lastname: Roberts
authors.forename: Patrick R
authors.initials: PR
authors.lastname: Cushing
authors.forename: Prisca
authors.initials: P
authors.lastname: Boisguerin
authors.forename: Dean R
authors.initials: DR
authors.lastname: Madden
authors.forename: Bruce R
authors.initials: BR
authors.lastname: Donald
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D001665:Binding Sites; D002352:Carrier Proteins / Q000037:antagonists & inhibitors* / Q000737:chemistry*; D003198:Computer Simulation; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000037:antagonists & inhibitors* / Q000648:ultrastructure*; D015195:Drug Design; D000077614:Golgi Matrix Proteins; D008565:Membrane Proteins / Q000037:antagonists & inhibitors* / Q000737:chemistry*; D026901:Membrane Transport Proteins; D008956:Models, Chemical; D008958:Models, Molecular; D054731:PDZ Domains; D010455:Peptides / Q000737:chemistry*; D011485:Protein Binding
chemical_list: D048868:Adaptor Proteins, Signal Transducing; C505032:CFTR protein, human; D002352:Carrier Proteins; C434484:GOPC protein, human; D000077614:Golgi Matrix Proteins; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D010455:Peptides; D019005:Cystic Fibrosis Transmembrane Conductance Regulator