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Allosteric Modulation of Intact ?-Secretase Structural Dynamics.

ABSTRACT: As a protease complex involved in the cleavage of amyloid precursor proteins that lead to the formation of amyloid ? fibrils implicated in Alzheimer's disease, ?-secretase is an important target for developing therapeutics against Alzheimer's disease. ?-secretase is composed of four subunits: nicastrin (NCT) in the extracellular (EC) domain, presenilin-1 (PS1), anterior pharynx defective 1, and presenilin enhancer 2 in the transmembrane (TM) domain. NCT and PS1 play important roles in binding amyloid ? precursor proteins and modulating PS1 catalytic activity. Yet, the molecular mechanisms of coupling between substrate/modulator binding and catalytic activity remain to be elucidated. Recent determination of intact human ?-secretase cryo-electron microscopy structure has opened the way for a detailed investigation of the structural dynamics of this complex. Our analysis, based on a membrane-coupled anisotropic network model, reveals two types of NCT motions, bending and twisting, with respect to PS1. These underlie the fluctuations between the "open" and "closed" states of the lid-like NCT with respect to a hydrophilic loop 1 (HL1) on PS1, thus allowing or blocking access of the substrate peptide (EC portion) to HL1 and to the neighboring helix TM2. In addition to this alternating access mechanism, fluctuations in the volume of the PS1 central cavity facilitate the exposure of the catalytic site for substrate cleavage. Druggability simulations show that ?-secretase presents several hot spots for either orthosteric or allosteric inhibition of catalytic activity, consistent with experimental data. In particular, a hinge region at the interface between the EC and TM domains, near the interlobe groove of NCT, emerges as an allo-targeting site that would impact the coupling between HL1/TM2 and the catalytic pocket, opening, to our knowledge, new avenues for structure-based design of novel allosteric modulators of ?-secretase protease activity.

SUBMITTER: Lee JY

PROVIDER: S-EPMC5770568 | biostudies-literature | 2017 Dec

REPOSITORIES: biostudies-literature

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Allosteric Modulation of Intact γ-Secretase Structural Dynamics.

Lee Ji Young JY Feng Zhiwei Z Xie Xiang-Qun XQ Bahar Ivet I

Biophysical journal 20171201 12

As a protease complex involved in the cleavage of amyloid precursor proteins that lead to the formation of amyloid β fibrils implicated in Alzheimer's disease, γ-secretase is an important target for developing therapeutics against Alzheimer's disease. γ-secretase is composed of four subunits: nicastrin (NCT) in the extracellular (EC) domain, presenilin-1 (PS1), anterior pharynx defective 1, and presenilin enhancer 2 in the transmembrane (TM) domain. NCT and PS1 play important roles in binding am ...[more]

PMID: 29262358

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Project description:BACKGROUND:γ-Secretase is a multiprotein protease that cleaves amyloid protein precursor (APP) and other type I transmembrane proteins. It has two catalytic subunits, presenilins 1 and 2 (PS1 and 2). In our previous report, we observed subtle differences in PS1- and PS2-mediated cleavages of select substrates and slightly different potencies of PS1 versus PS2 inhibition for select γ-secretase inhibitors (GSIs) on various substrates. In this study, we investigated whether γ-secretase modulators (GSMs) and inverse γ-secretase modulators (iGSMs) modulate γ-secretase processivity using multiple different substrates. We next used HEK 293T cell lines in which PSEN1 or PSEN2 was selectively knocked out to investigate processivity and response to GSMs and iGSMs. METHODS:For cell-free γ-secretase cleavage assay, recombinant substrates were incubated with CHAPSO-solubilized CHO or HEK 293T cell membrane with GSMs or iGSMs in suitable buffer. For cell-based assay, cDNA encoding substrates were transfected into HEK 293T cells. Cells were then treated with GSMs or iGSMs, and conditioned media were collected. Aβ and Aβ-like peptide production from cell-free and cell-based assay were measured by ELISA and mass spectrometry. RESULT:These studies demonstrated that GSMs are highly selective for effects on APP, whereas iGSMs have a more promiscuous effect on many substrates. Surprisingly, iGSMs actually appear to act as like GSIs on select substrates. The data with PSEN1 or PSEN2 knocked out HEK 293T reveal that PS1 has higher processivity and response to GSMs than PS2, but PS2 has higher response to iGSM. CONCLUSION:Collectively, these data indicate that GSMs are likely to have limited target-based toxicity. In addition, they show that iGSMs may act as substrate-selective GSIs providing a potential new route to identify leads for substrate-selective inhibitors of certain γ-secretase-mediated signaling events. With growing concerns that long-term β-secretase inhibitor is limited by target-based toxicities, such data supports continued development of GSMs as AD prophylactics.

Dataset Information

Allosteric Modulation of Intact ?-Secretase Structural Dynamics.

Publications

Allosteric Modulation of Intact γ-Secretase Structural Dynamics.

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