Toward the structure of presenilin/γ-secretase and presenilin homologs

Wolfe, Michael S.

doi:10.1016/j.bbamem.2013.04.015

Cited by 38 publications

(32 citation statements)

References 156 publications

(176 reference statements)

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“…In terms of how substrate may gain entry into the active site, replacing the mIAP sequence AGL 275 with PAL 435 found in presenilin (23)(24)(25) reduces catalytic efficiency by ~80% compared to WT, indicating PAL is not a favorable substitute (Fig. 1D, Table 1).…”

Section: Resultsmentioning

confidence: 99%

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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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Section: Resultsmentioning

confidence: 99%

“…The TM helices proximal to the proposed C-terminal substrate-gating motif PAL in presenilin (Fig. 1A) (23)(24)(25) need to undergo a substantial conformational change to enable entry of an exogenous TM substrate to the active site, a motion that is chemically incompatible with the potential presence of water in the active site.…”

Section: Introductionmentioning

confidence: 99%

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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“…We can make a distinction here between mature and functional (or active) ␥-secretase complexes (14,19,75). Maturation is related to NCT glycosylation.…”

Section: Discussionmentioning

confidence: 99%

“…␥-Secretase assembly is accompanied by an activating step in which presenilin is endoproteolysed and converted into a heterodimer formed by its N-and C-terminal fragments (PS-NTF/PS-CTF) (16 -18). In addition to this activating step, ␥-secretase complex has two catalytic activities: (i) endoproteolytic, releasing intracellular domains from type I integral membrane protein substrates, and (ii) carboxypeptidase, trimming the remaining TM domains from the substrate until the final product is released (19).…”

Section: A␤mentioning

confidence: 99%

Presenilin Transmembrane Domain 8 Conserved AXXXAXXXG Motifs Are Required for the Activity of the γ-Secretase Complex

Marinangeli¹,

Tasiaux²,

Opsomer³

et al. 2015

Journal of Biological Chemistry

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Background: Presenilin TMD8 contains a conserved AXXXAXXXG motif, involved in transmembrane protein interactions. Results: Mutation of PS AXXXAXXXG motifs strongly impacts ␥-secretase activity. Conclusion: The geometry and activity of ␥-secretase depend on the integrity of the conserved AXXXAXXXG motifs in TMD8. Significance: The PS AXXXAXXXG motif is a key determinant for understanding the structure, assembly, and function of the ␥-secretase complex.

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“…The increased ratio of Aβ42 over Aβ40 is thought to have a causal link with Alzheimer's disease (23). Considerable effort has been dedicated to the discovery of modulators that, upon binding to γ-secretase, may decrease the Aβ42/Aβ40 ratio.…”

Section: Significancementioning

confidence: 99%

Cleavage of amyloid precursor protein by an archaeal presenilin homologue PSH

Dang

Wang

et al. 2015

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

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Aberrant cleavage of amyloid precursor protein (APP) by γ-secretase contributes to the development of Alzheimer's disease. More than 200 disease-derived mutations have been identified in presenilin (the catalytic subunit of γ-secretase), making modulation of γ-secretase activity a potentially attractive therapeutic opportunity. Unfortunately, the technical challenges in dealing with intact γ-secretase have hindered discovery of modulators and demand a convenient substitute approach. Here we report that, similar to γ-secretase, the archaeal presenilin homolog PSH faithfully processes the substrate APP C99 into Aβ42, Aβ40, and Aβ38. The molar ratio of the cleavage products Aβ42 over Aβ40 by PSH is nearly identical to that by γ-secretase. The proteolytic activity of PSH is specifically suppressed by presenilin-specific inhibitors. Known modulators of γ-secretase also modulate PSH similarly in terms of the Aβ42/Aβ40 ratio. Structural analysis reveals association of a known γ-secretase inhibitor with PSH between its two catalytic aspartate residues. These findings identify PSH as a surrogate protease for the screening of agents that may regulate the protease activity and the cleavage preference of γ-secretase.A myloid precursor protein (APP) is initially cleaved by β-secretase in the extracellular space, producing a membrane-tethered, 99-residue carboxyl-terminal fragment known as APP C99 (1). APP C99 then undergoes sequential cleavages by γ-secretase, first at the e sites, yielding Aβ49/Aβ48, and eventually at the γ-sites, generating Aβ42/Aβ40/Aβ38 (2-4) (Fig. 1A). The 230-kDa γ-secretase contains four components: presenilin (PS), Pen-2, Aph-1, and nicastrin (NCT), of which PS1 is the target of most mutations derived from early-onset familial Alzheimer's disease (FAD) patients. Rather than abolishing the protease activity of γ-secretase, these mutations are thought to increase the molar ratio of Aβ42 over Aβ40 (2).Among the cleavage products of γ-secretase, Aβ42 is particularly prone to aggregation, leading to formation of β-amyloid plaque in the brain and presumably causing Alzheimer's disease (3). Other FAD-derived mutations map to APP and PS2, lending support to the causal relationship between formation of β-amyloid plaque and Alzheimer's disease (5). Therapeutic intervention of Alzheimer's disease may directly benefit from in vitro investigation of γ-secretase and discovery of its potential modulators (6). Unfortunately, such effort has been hampered by the difficulty in expression, purification, and manipulation of the complex protease. This problem persists despite recent breakthroughs in structural elucidation of γ-secretase and nicastrin (7,8).The intramembrane aspartate protease PSH from the archaeon Methanoculleus marisnigri JR1 shares 19% sequence identity and 53% sequence similarity with human PS1 (hPS1). The signature motifs for catalysis, ΦYDΦΦ (Φ for a hydrophobic residue) on transmembrane segment 6 (TM6) and ΦGΦGD on TM7, are identical between PSH and hPS1. PSH can be readily overexpressed in Escheri...

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Toward the structure of presenilin/γ-secretase and presenilin homologs

Cited by 38 publications

References 156 publications

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

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

Presenilin Transmembrane Domain 8 Conserved AXXXAXXXG Motifs Are Required for the Activity of the γ-Secretase Complex

Cleavage of amyloid precursor protein by an archaeal presenilin homologue PSH

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