Development of Phosphodiesterase–Protein-Kinase Complexes as Novel Targets for Discovery of Inhibitors with Enhanced Specificity

Phosphodiesterases (PDEs) hydrolyze cyclic nucleotides to modulate multiple signaling events in cells. PDEs are recognized to actively associate with cyclic nucleotide receptors (protein kinases, PKs) in larger macromolecular assemblies referred to as signalosomes. Complexation of PDEs with PKs gene...

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Bibliographic Details
Published in:International journal of molecular sciences 2021-05, Vol.22 (10), p.5242
Main Authors: Tulsian, Nikhil K., Sin, Valerie Jia-En, Koh, Hwee-Ling, Anand, Ganesh S.
Format: Article
Language:English
Subjects:
Binding sites
Cross-reactivity
Cyclic GMP
Cyclic nucleotides
Fluorescence
Fluorescence polarization
inhibitors
Kinases
Macromolecules
Mammals
natural products
Nucleotides
Phosphodiesterase
phosphodiesterase (PDE)
Protein interaction
Protein kinase
Proteins
selectivity
Signalosomes
Therapeutic targets
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Summary:Phosphodiesterases (PDEs) hydrolyze cyclic nucleotides to modulate multiple signaling events in cells. PDEs are recognized to actively associate with cyclic nucleotide receptors (protein kinases, PKs) in larger macromolecular assemblies referred to as signalosomes. Complexation of PDEs with PKs generates an expanded active site that enhances PDE activity. This facilitates signalosome-associated PDEs to preferentially catalyze active hydrolysis of cyclic nucleotides bound to PKs and aid in signal termination. PDEs are important drug targets, and current strategies for inhibitor discovery are based entirely on targeting conserved PDE catalytic domains. This often results in inhibitors with cross-reactivity amongst closely related PDEs and attendant unwanted side effects. Here, our approach targeted PDE–PK complexes as they would occur in signalosomes, thereby offering greater specificity. Our developed fluorescence polarization assay was adapted to identify inhibitors that block cyclic nucleotide pockets in PDE–PK complexes in one mode and disrupt protein-protein interactions between PDEs and PKs in a second mode. We tested this approach with three different systems—cAMP-specific PDE8–PKAR, cGMP-specific PDE5–PKG, and dual-specificity RegA–RD complexes—and ranked inhibitors according to their inhibition potency. Targeting PDE–PK complexes offers biochemical tools for describing the exquisite specificity of cyclic nucleotide signaling networks in cells.
ISSN:1422-0067
1661-6596
1422-0067
DOI:10.3390/ijms22105242