A Computational Study of the Oligosaccharide Binding Sites in the Lectin-Like Domain of Tumor Necrosis Factor and the TNF-derived TIP Peptide

Dulebo, Alexander; Ettrich, Rüdiger; Lucas, Rudolf; Kaftan, David

doi:10.2174/138161212802430549

Cited by 8 publications

(8 citation statements)

References 30 publications

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“…These results correspond to data from a study of molecular docking between TNF and N,N9-diacetylchitobiose, indicating that oligosaccharide binding to TNF sterically hinders its association with residues involved in the Na 1 uptake stimulating effect of the cytokine (40).…”

Section: As Shown Insupporting

confidence: 86%

“…The puzzling observation that oligosaccharides can blunt the binding of the TIP peptide to recombinant, and thus nonglycosylated, subunits can be explained by our previously published molecular docking study indicating that residues involved in the binding of human TNF to N,N9-diacetylchitobiose are different from, but spatially close to, those necessary for activation of Na 1 uptake (40). Indeed, the mutant TIP peptide, which has a strongly reduced capacity to bind to ENaC-a and activate ENaC, still efficiently binds to N,N9-diacetylchitobiose (59).…”

Section: Discussionmentioning

confidence: 99%

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A Novel Tumor Necrosis Factor–mediated Mechanism of Direct Epithelial Sodium Channel Activation

Czikora

Alli

Bao

et al. 2014

Am J Respir Crit Care Med

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Rationale: Alveolar liquid clearance is regulated by Na 1 uptake through the apically expressed epithelial sodium channel (ENaC) and basolaterally localized Na 1 -K 1 -ATPase in type II alveolar epithelial cells. Dysfunction of these Na 1 transporters during pulmonary inflammation can contribute to pulmonary edema.Objectives: In this study, we sought to determine the precise mechanism by which the TIP peptide, mimicking the lectin-like domain of tumor necrosis factor (TNF), stimulates Na 1 uptake in a homologous cell system in the presence or absence of the bacterial toxin pneumolysin (PLY).Methods: We used a combined biochemical, electrophysiological, and molecular biological in vitro approach and assessed the physiological relevance of the lectin-like domain of TNF in alveolar liquid clearance in vivo by generating triple-mutant TNF knock-in mice that express a mutant TNF with deficient Na 1 uptake stimulatory activity.Measurements and Main Results: TIP peptide directly activates ENaC, but not the Na 1 -K 1 -ATPase, upon binding to the carboxyterminal domain of the a subunit of the channel. In the presence of PLY, a mediator of pneumococcal-induced pulmonary edema, this binding stabilizes the ENaC-PIP2-MARCKS complex, which is necessary for the open probability conformation of the channel and preserves ENaC-a protein expression, by means of blunting the protein kinase C-a pathway. Triple-mutant TNF knock-in mice are more prone than wild-type mice to develop edema with low-dose intratracheal PLY, correlating with reduced pulmonary ENaC-a subunit expression.Conclusions: These results demonstrate a novel TNF-mediated mechanism of direct ENaC activation and indicate a physiological role for the lectin-like domain of TNF in the resolution of alveolar edema during inflammation.

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Section: As Shown Insupporting

confidence: 86%

Section: Discussionmentioning

confidence: 99%

A Novel Tumor Necrosis Factor–mediated Mechanism of Direct Epithelial Sodium Channel Activation

Czikora

Alli

Bao

et al. 2014

Am J Respir Crit Care Med

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“…This was demonstrated in whole-cell recordings and in singlechannel experiments in both endogenously as well as heterologously expressed hENaC. Recent docking and molecular dynamics simulation experiments indicated that the TIP peptide AP301 represents a partial binding motif for chitobiose (Dulebo et al, 2012). By definition, lectins bind to glycans, and our data clearly show that glycosylation of ENaC is necessary for activation with AP301.…”

Section: Ap301-induced Activation Of Henacsupporting

confidence: 70%

“…Deglycosylation with -(N-Acetyl-b-D-Glucosaminyl)Asparagine Amidase F. Docking and molecular dynamics simulation experiments indicated that the TIP peptide AP301 represents a partial binding motif for chitobiose (Dulebo et al, 2012). To investigate a possible interaction of TIP peptides with sugar moieties on the cell membrane, deglycosylation of A549 as well as transfected and nontransfected HEK-293 cell membranes was performed with peptide-N 4 -(N-acetyl-b-D-glucosaminyl)asparagine amidase F (PNGase F, peptide N-glycanase) at room temperature.…”

Section: Electrophysiologymentioning

confidence: 99%

Mechanism of Action of Novel Lung Edema Therapeutic AP301 by Activation of the Epithelial Sodium Channel

et al. 2013

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AP301 [Cyclo(CGQRETPEGAEAKPWYC)], a cyclic peptide comprising the human tumor necrosis factor lectin-like domain (TIP domain) sequence, is currently being developed as a treatment for lung edema and has been shown to reduce extravascular lung water and improve lung function in mouse, rat, and pig models. The current paradigm for liquid homeostasis in the adult mammalian lung is that passive apical uptake of sodium via the amiloride-sensitive epithelial Na 1 channel (ENaC) and nonselective cyclic-nucleotide-gated cation channels creates the major driving force for reabsorption of water through the alveolar epithelium in addition to other ion channels such as potassium and chloride channels. AP301 can increase amiloride-sensitive current in A549 cells as well as in freshly isolated type II alveolar epithelial cells from different species. ENaC is expressed endogenously in all of these cell types. Consequently, this study was undertaken to determine whether ENaC is the specific target of AP301. The effect of AP301 in A549 cells as well as in human embryonic kidney cells and Chinese hamster ovary cells heterologously expressing human ENaC subunits (a, b, g, and d) was measured in patch clamp experiments. The congener TIP peptide AP318 [Cyclo(4-aminobutanoic acid-GQRETPEGAEAKPWYD)] activated ENaC by increasing single-channel open probability. AP301 increased current in proteolytically activated (cleaved) but not near-silent (uncleaved) ENaC in a reversible manner. abg-or dbg-ENaC coexpression was required for maximal activity. No increase in current was observed after deglycosylation of extracellular domains of ENaC. Thus, our data suggest that the specific interaction of AP301 with both endogenously and heterologously expressed ENaC requires precedent binding to glycosylated extracellular loop(s).

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“…All energy calculations and minimizations/dynamics used periodic boundary conditions and the YAMBER3 force field34. The system was then energy-minimized using steepest descent minimization, in order to remove conformational stress, followed by a simulated annealing minimization until convergence (<0.05 kJ per mol per 200 steps).…”

Section: Methodsmentioning

confidence: 99%

Deconvolution of Buparlisib’s mechanism of action defines specific PI3K and tubulin inhibitors for therapeutic intervention

et al. 2017

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BKM120 (Buparlisib) is one of the most advanced phosphoinositide 3-kinase (PI3K) inhibitors for the treatment of cancer, but it interferes as an off-target effect with microtubule polymerization. Here, we developed two chemical derivatives that differ from BKM120 by only one atom. We show that these minute changes separate the dual activity of BKM120 into discrete PI3K and tubulin inhibitors. Analysis of the compounds cellular growth arrest phenotypes and microtubule dynamics suggest that the antiproliferative activity of BKM120 is mainly due to microtubule-dependent cytotoxicity rather than through inhibition of PI3K. Crystal structures of BKM120 and derivatives in complex with tubulin and PI3K provide insights into the selective mode of action of this class of drugs. Our results raise concerns over BKM120's generally accepted mode of action, and provide a unique mechanistic basis for next-generation PI3K inhibitors with improved safety profiles and flexibility for use in combination therapies.

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A Computational Study of the Oligosaccharide Binding Sites in the Lectin-Like Domain of Tumor Necrosis Factor and the TNF-derived TIP Peptide

Cited by 8 publications

References 30 publications

A Novel Tumor Necrosis Factor–mediated Mechanism of Direct Epithelial Sodium Channel Activation

A Novel Tumor Necrosis Factor–mediated Mechanism of Direct Epithelial Sodium Channel Activation

Mechanism of Action of Novel Lung Edema Therapeutic AP301 by Activation of the Epithelial Sodium Channel

Deconvolution of Buparlisib’s mechanism of action defines specific PI3K and tubulin inhibitors for therapeutic intervention

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