Mechanism of Action of an EPAC1-Selective Competitive Partial Agonist

Shao, Hongzhao; Mohamed, Hebatallah; Boulton, Stephen; Huang, Jinfeng; Wang, Pingyuan; Chen, Haiying; Zhou, Jia; Luchowska-Stańska, Urszula; Jentsch, Nicholas G.; Armstrong, Alison; Magolan, Jakob; Yarwood, Stephen J.; Melacini, Giuseppe

doi:10.1021/acs.jmedchem.9b02151

Cited by 15 publications

(33 citation statements)

References 81 publications

(230 reference statements)

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“…Measurements of cNMP-binding affinities for HCN4 were achieved via competitive binding with the fluorescent cAMP analog, 8-NBD-cAMP, using an approach similar to the protocols described previously (49)(50)(51). The K d of 8-NBD-cAMP for each HCN4 construct was initially determined via a titration in which the 8-NBD-cAMP concentration was fixed at 500 nM and the HCN4 construct was added at concentrations ranging from 0.1 to 50 mM.…”

Section: Affinity Measurementsmentioning

confidence: 99%

Allosteric Mechanisms of Nonadditive Substituent Contributions to Protein-Ligand Binding

Boulton

Van

VanSchouwen

et al. 2020

Biophysical Journal

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Quantifying chemical substituent contributions to ligand-binding free energies is challenging due to nonadditive effects. Protein allostery is a frequent cause of nonadditivity, but the underlying allosteric mechanisms often remain elusive.Here, we propose a general NMR-based approach to elucidate such mechanisms and we apply it to the HCN4 ion channel, whose cAMP-binding domain is an archetypal conformational switch. Using NMR, we show that nonadditivity arises not only from concerted conformational transitions, but also from conformer-specific effects, such as steric frustration. Our results explain how affinity-reducing functional groups may lead to affinity gains if combined. Surprisingly, our approach also reveals that nonadditivity depends markedly on the receptor conformation. It is negligible for the inhibited state but highly significant for the active state, opening new opportunities to tune potency and agonism of allosteric effectors.

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Section: Affinity Measurementsmentioning

confidence: 99%

Allosteric Mechanisms of Nonadditive Substituent Contributions to Protein-Ligand Binding

Boulton

Van

VanSchouwen

et al. 2020

Biophysical Journal

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“…Interestingly, depletion of ANXA2 in ECs reduced the enhanced binding force between reOmpB and ECs in the presence of I942 (versus the WT group, p = 0.09), independent of its concentration in the culture medium (Figure S2). Taking advantage of a recently developed non-cAMP analogue, an EPAC1 specific activator named I942 that competitively binds the cyclic nucleotide-binding domain of EPAC1 [50][51][52], we observed that I942 at 5 µM in culture media enhanced the interacting force between reOmpB and a live WT BMEC (p < 0.05) (Figure 4). Depletion of ANXA2 in ECs diminished reOmpB binding forces to the ANXA2-KO BMEC surface (p < 0.01) (Figure 4), corroborating our previous report that anti-ANXA2 antibody weakens the interacting force between reOmpB and a living EC [44].…”

Section: Rickettsial Ompb Shows a Host Epac1-dependent Binding Strength On The Surface Of A Living Ecmentioning

confidence: 98%

Host EPAC1 Modulates Rickettsial Adhesion to Vascular Endothelial Cells via Regulation of ANXA2 Y23 Phosphorylation

Shelite

Qiu

et al. 2021

Pathogens

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Introduction: Intracellular cAMP receptor exchange proteins directly activated by cAMP 1 (EPAC1) regulate obligate intracellular parasitic bacterium rickettsial adherence to and invasion into vascular endothelial cells (ECs). However, underlying precise mechanism(s) remain unclear. The aim of the study is to dissect the functional role of the EPAC1-ANXA2 signaling pathway during initial adhesion of rickettsiae to EC surfaces. Methods: In the present study, an established system that is anatomically based and quantifies bacterial adhesion to ECs in vivo was combined with novel fluidic force microscopy (FluidFM) to dissect the functional role of the EPAC1-ANXA2 signaling pathway in rickettsiae–EC adhesion. Results: The deletion of the EPAC1 gene impedes rickettsial binding to endothelium in vivo. Rickettsial OmpB shows a host EPAC1-dependent binding strength on the surface of a living brain microvascular EC (BMEC). Furthermore, ectopic expression of phosphodefective and phosphomimic mutants replacing tyrosine (Y) 23 of ANXA2 in ANXA2-knock out BMECs results in different binding force to reOmpB in response to the activation of EPAC1. Conclusions: EPAC1 modulates rickettsial adhesion, in association with Y23 phosphorylation of the binding receptor ANXA2. Underlying mechanism(s) should be further explored to delineate the accurate role of cAMP-EPAC system during rickettsial infection.

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“…Recently, Yarwood’s group identified, in a high throughput screening (HTS) study, the first known non-cyclic nucleotide (NCN) EPAC1 agonist, I942 with in vitro application [ 32 , 33 ]. NMR spectrometry analysis demonstrated that I942 can stabilize an inhibition-incompetent activation intermediate state of EPAC1 distinct from both active and inactive states [ 34 ]. Additionally, in a study carried out by the Zhou’s group, a series of non-cyclic nucleotide activators named 25g, 25q, 25n, 25u, 25e, and 25f with good selectivity over PKA were identified.…”

Section: Epac1 Pharmacological Modulatorsmentioning

confidence: 99%

Role of EPAC1 Signalosomes in Cell Fate: Friends or Foes?

Formoso

Lezoualc’h

Mialet‐Perez

2020

Cells

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The compartmentation of signaling processes is accomplished by the assembly of protein complexes called signalosomes. These signaling platforms colocalize enzymes, substrates, and anchoring proteins into specific subcellular compartments. Exchange protein directly activated by cAMP 1 (EPAC1) is an effector of the second messenger, 3′,5′-cyclic adenosine monophosphate (cAMP) that is associated with multiple roles in several pathologies including cardiac diseases. Both EPAC1 intracellular localization and molecular partners are key players in the regulation of cell fate, which may have important therapeutic potential. In this review, we summarize the recent findings on EPAC1 structure, regulation, and pharmacology. We describe the importance of EPAC1 subcellular distribution in its biological action, paying special attention to its nuclear localization and mechanism of action leading to cardiomyocyte hypertrophy. In addition, we discuss the role of mitochondrial EPAC1 in the regulation of cell death. Depending on the cell type and stress condition, we present evidence that supports either a protective or detrimental role of EPAC1 activation.

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Mechanism of Action of an EPAC1-Selective Competitive Partial Agonist

Cited by 15 publications

References 81 publications

Allosteric Mechanisms of Nonadditive Substituent Contributions to Protein-Ligand Binding

Allosteric Mechanisms of Nonadditive Substituent Contributions to Protein-Ligand Binding

Host EPAC1 Modulates Rickettsial Adhesion to Vascular Endothelial Cells via Regulation of ANXA2 Y23 Phosphorylation

Role of EPAC1 Signalosomes in Cell Fate: Friends or Foes?

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