The Backbone Dynamics of the Amyloid Precursor Protein Transmembrane Helix Provides a Rationale for the Sequential Cleavage Mechanism of γ-Secretase

Pester, Oxana; Barrett, Paul J.; Hornburg, Daniel; Hornburg, Philipp; Pröbstle, Rasmus; Widmaier, Simon; Kutzner, Christoph; Dürrbaum, Milena; Kapurniotu, Aphrodite; Sanders, Charles R.; Scharnagl, Christina; Langosch, Dieter

doi:10.1021/ja3112093

Cited by 74 publications

(206 citation statements)

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“…The recent NMR investigations of the TMD of ␤CTF lend substantial support to the model (35,36). However, the pathways are not as simple as we previously thought.…”

Section: Discussionmentioning

confidence: 72%

γ-Secretase Associated with Lipid Rafts

Matsumura

Takami

Okochi

et al. 2014

Journal of Biological Chemistry

101

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Background: Intramembranous cleavages of ␤-carboxyl-terminal fragment (␤CTF) by ␥-secretase generate amyloid ␤-protein (A␤). Results: Three-to six-residue peptides are released successively along with A␤ generation by lipid raft-associated ␥-secretase. Conclusion: ␥-Secretase cleaves ␤CTF through multiple interactive pathways for stepwise successive processing to generate A␤.Significance: This cleavage model provides insights into the precise molecular mechanism of A␤ generation.

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“…The recent NMR investigations of the TMD of ␤CTF lend substantial support to the model (35,36). However, the pathways are not as simple as we previously thought.…”

Section: Discussionmentioning

confidence: 72%

γ-Secretase Associated with Lipid Rafts

Matsumura

Takami

Okochi

et al. 2014

Journal of Biological Chemistry

101

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“…This key prediction was validated in a recent study involving H/D exchange experiments on the C99 peptide complemented with molecular dynamics (MD) simulations of the TM region, C99 28−55 , in a 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) bilayer (20), which also provided detailed insights into the stability of the TM helical region of C99. It was found that the portion of the TM helix on the N-terminal side of the Gly 37 /Gly 38 hinge was more dynamic and showed enhanced H/D exchange relative to the C-terminal portion of the TM helix (20), a finding in agreement with studies of the C99 monomer in lyso-myristoyl phosphatidylglycerol (LMPG) micelles (21,22).…”

mentioning

confidence: 71%

Impact of membrane lipid composition on the structure and stability of the transmembrane domain of amyloid precursor protein

Domı́nguez

Foster

Straub

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

116

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Cleavage of the amyloid precursor protein (APP) by γ-secretase is a crucial first step in the evolution of Alzheimer's disease. To discover the cleavage mechanism, it is urgent to predict the structures of APP monomers and dimers in varying membrane environments. We determined the structures of the C99 23−55 monomer and homodimer as a function of membrane lipid composition using a multiscale simulation approach that blends atomistic and coarsegrained models. We demonstrate that the C99 23−55 homodimer structures form a heterogeneous ensemble with multiple conformational states, each stabilized by characteristic interpeptide interactions. The relative probabilities of each conformational state are sensitive to the membrane environment, leading to substantial variation in homodimer peptide structure as a function of membrane lipid composition or the presence of an anionic lipid environment. In contrast, the helicity of the transmembrane domain of monomeric C99 1−55 is relatively insensitive to the membrane lipid composition, in agreement with experimental observations. The dimer structures of human EphA2 receptor depend on the lipid environment, which we show is linked to the location of the structural motifs in the dimer interface, thereby establishing that both sequence and membrane composition modulate the complete energy landscape of membrane-bound proteins. As a by-product of our work, we explain the discrepancy in structures predicted for C99 congener homodimers in membrane and micelle environments. Our study provides insight into the observed dependence of C99 protein cleavage by γ-secretase, critical to the formation of amyloid-β protein, on membrane thickness and lipid composition.nderstanding the structural and thermodynamic properties of transmembrane (TM) helical dimers is of fundamental importance to molecular biology. It is known that the association of "bitopic" proteins, having single pass TM helical domains, is essential to immunoreceptors and protein kinases that play critical roles in cellular function. Computational and experimental studies have provided insight into the role of sequence-specific interactions stabilizing helix dimerization (1, 2). Examples include the heptad repeat motif responsible for the stability of coiled-coils in GCN4 phospholamban (3) and the M2 proton channel (4), the role of the GxxxG motif in stabilizing TM helix-helix association in systems including the glycophorin A (GpA) homodimer (5, 6), found in the human erythrocyte membrane, and GxxxG and heptad repeat motifs, which play a role in stabilizing homodimers of APP-C99 (C99), the 99-aa C-terminal fragment of the amyloid precursor protein (APP) (7,8).The amyloid β (Aβ) peptide aggregation pathway, known to be crucial to the evolution of Alzheimer's disease (AD), begins with the cleavage of C99 by γ-secretase leading to the formation of a number of isoforms of Aβ. The formation of homodimers of C99 has been proposed to be critical to the mechanism by which C99 is cleaved by γ-secretase, a process that is known to dep...

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“…2E-G). LC-MS cleavage profiles for MCS10 reveal highly specific cut sites -Ala 42 -Thr 43 and Thr 43 -Val 44 -both in DDM and bicelles (Fig. 2H).…”

Section: Resultsmentioning

confidence: 97%

Both positional and chemical variables control in vitro proteolytic cleavage of a presenilin ortholog

Naing

Kalyoncu

Smalley³

et al. 2018

Journal of Biological Chemistry

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Mechanistic details of intramembrane aspartyl protease (IAP) chemistry, which is central to many biological and pathogenic processes, remain largely obscure. Here, we investigated the in vitro kinetics of a microbial intramembrane aspartyl protease (mIAP) fortuitously acting on the renin substrate angiotensinogen and the C-terminal transmembrane segment of amyloid precursor protein (C100), which is cleaved by the presenilin subunit of γ-secretase, an Alzheimer disease (AD)-associated IAP. mIAP variants with substitutions in active-site and putative substrate gating residues generally exhibit impaired, but not abolished, activity toward angiotensinogen, and retain the predominant cleavage site (His-Thr). The aromatic ring, but not its hydroxyl substituent, in Tyr of the catalytic Tyr-Asp (YD) motif plays a catalytic role, and the hydrolysis reaction incorporates bulk water as in soluble aspartyl proteases. mIAP hydrolyzes the transmembrane region of C100 at two major presenilin cleavage sites, one corresponding to the AD-associated Aβ42 peptide (Ala-Thr) and the other the nonpathogenic Aβ48 (Thr-Leu). For the former site, we observed more favorable kinetics in lipid bilayer-mimicking bicelles than in detergent solution, indicating that substrate-lipid and substrate-enzyme interactions both contribute to catalytic rates. High-resolution MS analyses across four substrates support a preference for threonine at the scissile bond. However, results from threonine-scanning mutagenesis of angiotensinogen indicate a competing positional preference for cleavage. Our results indicate that IAP cleavage is controlled by both positional and chemical factors, opening up new avenues for selective IAP inhibition for therapeutic interventions. INTRODUCTIONIntramembrane proteases (IPs) cleave within a transmembrane (TM) helix of membranebound substrates to release cytoplasmic or extracellular proteins/peptides, which in turn translocate to different regions of the cell where they elicit their corresponding biological response related to, for example, cell differentiation, development, and metabolism (1). Despite their broad biomedical reach, basic questions surrounding the structure of the active enzyme, how substrates are presented, and how hydrolysis chemistry occurs in an active site sequestered within the hydrophobic lipid membrane, remain active areas of research.The least biochemically understood IP is the intramembrane aspartyl protease (IAP) enzyme class, one of just three bona fide IP types that hydrolyze substrates within the hydrophobic lipid environment by using different catalytic nucleophiles (2). IAPs employ membrane-

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The Backbone Dynamics of the Amyloid Precursor Protein Transmembrane Helix Provides a Rationale for the Sequential Cleavage Mechanism of γ-Secretase

Cited by 74 publications

References 73 publications

γ-Secretase Associated with Lipid Rafts

γ-Secretase Associated with Lipid Rafts

Impact of membrane lipid composition on the structure and stability of the transmembrane domain of amyloid precursor protein

Both positional and chemical variables control in vitro proteolytic cleavage of a presenilin ortholog

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