Cysteine modifiers suggest an allosteric inhibitory site on the CAL PDZ domain.

Bioscience Reports
Yu ZhaoDean R Madden

Abstract

Protein-protein interactions have become attractive targets for both experimental and therapeutic interventions. The PSD-95/Dlg1/ZO-1 (PDZ) domain is found in a large family of eukaryotic scaffold proteins that plays important roles in intracellular trafficking and localization of many target proteins. Here, we seek inhibitors of the PDZ protein that facilitates post-endocytic degradation of the cystic fibrosis (CF) transmembrane conductance regulator (CFTR): the CFTR-associated ligand (CAL). We develop and validate biochemical screens and identify methyl-3,4-dephostatin (MD) and its analog ethyl-3,4-dephostatin (ED) as CAL PDZ inhibitors. Depending on conditions, MD can bind either covalently or non-covalently. Crystallographic and NMR data confirm that MD attacks a pocket at a site distinct from the canonical peptide-binding groove, and suggests an allosteric connection between target residue Cys319 and the conserved Leu291 in the GLGI motif. MD and ED thus appear to represent the first examples of small-molecule allosteric regulation of PDZ:peptide affinity. Their mechanism of action may exploit the known conformational plasticity of the PDZ domains and suggests that allosteric modulation may represent a strategy for targeti...Continue Reading

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Citations

Jan 16, 2020·International Journal of Molecular Sciences·Samuel Estabrooks, Jeffrey L Brodsky
Dec 15, 2020·Journal of Medicinal Chemistry·Patrick G DoughertyDehua Pei
Oct 21, 2020·Bioconjugate Chemistry·Michael J Swierczynski, Zachary T Ball

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Methods Mentioned

BETA
PCR
size-exclusion chromatography
FRET
AlphaScreen
NMR
transfection
Assay
chemical shift

Software Mentioned

Phenix
PyMOL
Sparky
CALP
MD
nlme
Wincoot
R

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