Structural and biochemical differences between the Notch and the amyloid precursor protein transmembrane domains

Deatherage, Catherine L.; Lu, Zhenwei; Kroncke, Brett M.; Ma, Si‐Rui; Smith, Jarrod A.; Voehler, Markus; McFeeters, Robert L.; Sanders, Charles R.

doi:10.1126/sciadv.1602794

Cited by 40 publications

(46 citation statements)

References 95 publications

(171 reference statements)

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“…A recent structural study of γ-secretase cleavage 17 , reported that when native Notch is recognized by γ-secretase, the C-terminus of the TMD must be precisely positioned to form a hybrid β-sheet with the protease. Another structural study suggested that the RAM sequence behind the C-terminus of TMD could form a membrane docking domain 18 , which may help the positioning of TMD. Because the RAM sequence was not included in any published synNotch, we hypothesized that adding this natural motif might suppress LIA.…”

Section: Resultsmentioning

confidence: 99%

Engineering of an enhanced synthetic Notch receptor by reducing ligand-independent activation

Yang

Yue

et al. 2020

Commun Biol

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Notch signaling is highly conserved in most animals and plays critical roles during neurogenesis as well as embryonic development. Synthetic Notch-based systems, modeled from Notch receptors, have been developed to sense and respond to a specific extracellular signal. Recent advancement of synNotch has shown promise for future use in cellular engineering to treat cancers. However, synNotch from Morsut et al. (2016) has a high level of ligandindependent activation, which limits its application. Here we show that adding an intracellular hydrophobic sequence (QHGQLWF, named as RAM7) present in native Notch, significantly reduced ligand-independent activation. Our enhanced synthetic Notch receptor (esNotch) demonstrates up to a 14.6-fold reduction in ligand-independent activation, without affecting its antigen-induced activation efficiency. Our work improves a previously reported transmembrane receptor and provides a powerful tool to develop better transmembrane signaling transduction modules for further advancement of eukaryotic synthetic biology.

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Section: Resultsmentioning

confidence: 99%

Engineering of an enhanced synthetic Notch receptor by reducing ligand-independent activation

Yang

Yue

et al. 2020

Commun Biol

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“…The FAD mutations consistently increase H-bond fluctuations in the region upstream to the ε-site while maintaining strong α-helicity near the cleavage sites. A stable helix near scissile bonds was found previously not only for the C99-TMD (Chen et al, 2014; Dominguez et al, 2016; Pester, Barrett, et al, 2013; Pester, Götz, et al, 2013; Takeshi Sato et al, 2009; Scharnagl et al, 2014), but also for Notch (Deatherage et al, 2017) and the SREBP-1 substrate of the intramembrane site-2 protease (Linser et al, 2015). The vicinity of the γ-site was shown, by solid-state NMR, to exist in a mixture of helical and non-helical conformations (Itkin et al, 2017; J.-X.…”

Section: Resultsmentioning

confidence: 67%

“…While the repertoire of conformational motions of the substrate TMDs is mainly determined by their α-helical fold architecture, there is reason to suspect that the relative importance of individual motions reflects differences in sequence and local fold. The TMD of NOTCH, another prominent substrate of GSEC (Beel & Sanders, 2008), appears to be a straight helix (Deatherage et al, 2017) missing the large-scale bending flexibility that characterizes the C99 TMD. As a result, the TMDs of NOTCH and C99 adapt differently to the hydrophobic thickness of the membrane environment (Deatherage et al, 2017).…”

Section: Resultsmentioning

confidence: 99%

“…The TMD of NOTCH, another prominent substrate of GSEC (Beel & Sanders, 2008), appears to be a straight helix (Deatherage et al, 2017) missing the large-scale bending flexibility that characterizes the C99 TMD. As a result, the TMDs of NOTCH and C99 adapt differently to the hydrophobic thickness of the membrane environment (Deatherage et al, 2017). In a similar manner, the coexistence of multiple dynamic features of substrate TMDs might determine how they adapt to interactions with the enzyme.…”

Section: Resultsmentioning

confidence: 99%

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Dissecting Conformational Changes in APP’s Transmembrane Domain Linked to ε-Efficiency in Familial Alzheimer’s Disease

Goetz

Scharnagl

2018

Preprint

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1The mechanism by which familial Alzheimer's disease (FAD) mutations within the transmembrane domain 2 (TMD) of the Amyloid Precursor Protein (APP) affect ε-endoproteolysis is only poorly understood. Thereby, motions, obscured by large-scale motions in the pre-bound state, provide (i) a dynamic mechanism underlying 10 the proposed coupling between binding and ε-cleavage, (ii) key sites consistent with experimentally determined 11 docking sites, and (iii) the distinction between mutants and wild-type.

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“…Notably, the TMD of Notch1, as well as the TMD of the insulin receptor (IR), two other substrates of γ-secretase (9), have conformations very different from that of the C99 TMD as determined from solution-NMR of single-span TM helices in membrane mimics (100,101). In particular, Notch1 appears to be a straight helix, while the TMD helix of the might nevertheless provide the repertoire of functionally important motions necessary to adapt to interactions with the enzyme at different stages of the catalytic cycle.…”

Section: Discussionmentioning

confidence: 99%

Modulating hinge flexibility in the APP transmembrane domain alters γ-secretase cleavage

Götz¹,

Mylonas²,

Högel³

et al. 2018

Preprint

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Intramembrane cleavage of the β-amyloid precursor protein C99 substrate by γsecretase is implicated in Alzheimer´s disease pathogenesis. Since conformational flexibility of a di-glycine hinge in the C99 transmembrane domain (TMD) might be critical for γ-secretase cleavage, we mutated one of the glycine residues, G38, to a helixstabilizing leucine and to a helix-distorting proline. CD, NMR and hydrogen/deuterium exchange measurements as well as MD simulations showed that the mutations distinctly altered the intrinsic structural and dynamical properties of the TMD. However, although helix destabilization/unfolding was not observed at the initial ε-cleavage sites of C99, both mutants impaired γ-secretase cleavage and altered its cleavage specificity. Moreover, helix flexibility enabled by the di-glycine hinge translated to motions of other helix parts. Our data suggest that both local helix stabilization and destabilization in the di-glycine hinge may decrease the occurrence of enzyme-substrate complex conformations required for normal catalysis and that hinge mobility can be conducive for productive substrate-enzyme interactions.

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Structural and biochemical differences between the Notch and the amyloid precursor protein transmembrane domains

Abstract: The Notch transmembrane domain exhibits significant structural differences from that of the amyloid precursor protein.

Cited by 40 publications

References 95 publications

Engineering of an enhanced synthetic Notch receptor by reducing ligand-independent activation

Engineering of an enhanced synthetic Notch receptor by reducing ligand-independent activation

Dissecting Conformational Changes in APP’s Transmembrane Domain Linked to ε-Efficiency in Familial Alzheimer’s Disease

Modulating hinge flexibility in the APP transmembrane domain alters γ-secretase cleavage

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