Presenilin 1 is linked with γ-secretase activity in the detergent solubilized state

Li, Yueming; Lai, Ming‐Tain; Xu, Min; Huang, Qian; DiMuzio-Mower, Jillan; Sardana, Mohinder K.; Shi, Xiao‐Ping; Yin, Kuo‐Chang

doi:10.1016/s0197-4580(00)83199-1

Cited by 36 publications

(62 citation statements)

References 12 publications

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“…In vitro γ-secretase assays were performed as previously described (30,32) except for samples in which no SDS was added to the C100Flag substrate preparations before the activity assays.…”

Section: Multistep Purification Of γ-Secretase From S-1 Cells (1-3) mentioning

confidence: 99%

Purification and Characterization of the Human γ-Secretase Complex

et al. 2004

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Gamma-secretase is a member of an unusual class of proteases with intramembrane catalytic sites. This enzyme cleaves many type I membrane proteins, including the amyloid beta-protein (Abeta) precursor (APP) and the Notch receptor. Biochemical and genetic studies have identified four membrane proteins as components of gamma-secretase: heterodimeric presenilin (PS) composed of its N- and C-terminal fragments (PS-NTF/CTF), a mature glycosylated form of nicastrin (NCT), Aph-1, and Pen-2. Recent data from studies in Drosophila, mammalian, and yeast cells suggest that PS, NCT, Aph-1, and Pen-2 are necessary and sufficient to reconstitute gamma-secretase activity. However, many unresolved issues, in particular the possibility of other structural or regulatory components, would be resolved by actually purifying the enzyme. Here, we report a detailed, multistep purification procedure for active gamma-secretase and an initial characterization of the purified protease. Extensive mass spectrometry of the purified proteins strongly suggests that PS-NTF/CTF, mNCT, Aph-1, and Pen-2 are the components of active gamma-secretase. Using the purified gamma-secretase, we describe factors that modulate the production of specific Abeta species: (1) phosphatidylcholine and sphingomyelin dramatically improve activity without changing cleavage specificity within an APP substrate; (2) increasing CHAPSO concentrations from 0.1 to 0.25% yields a approximately 100% increase in Abeta42 production; (3) exposure of an APP-based recombinant substrate to 0.5% SDS modulates cleavage specificity from a disease-mimicking pattern (high Abeta42/43) to a physiological pattern (high Abeta40); and (4) sulindac sulfide directly and preferentially decreases Abeta42 cleavage within the purified complex. Taken together, our results define a procedure for purifying active gamma-secretase and suggest that the lipid-mediated conformation of both enzyme and substrate regulate the production of the potentially neurotoxic Abeta42 and Abeta43 peptides.

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“…In vitro γ-secretase assays were performed as previously described (30,32) except for samples in which no SDS was added to the C100Flag substrate preparations before the activity assays.…”

Section: Multistep Purification Of γ-Secretase From S-1 Cells (1-3) mentioning

confidence: 99%

Purification and Characterization of the Human γ-Secretase Complex

et al. 2004

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“…26 That some γ-secretase activity resides in PS1 has been supported by studies using known transition state specific inhibitors of γ-secretase activity to bind, label, and identify PS1 and also by detecting γ-secretase activity in PS1 associated complexes. [27][28][29] Taken together, these findings have led to the hypothesis that PS1 might directly or indirectly regulate γ-secretase, perhaps as a diaspartyl cofactor or by post-translational modification of γ-secretase and/or APP, or indeed might actually be γ-secretase itself. However, the therapeutic viability of inhibiting γ-secretase could be limited as this may deleteriously affect Notch signalling pathways that are involved in haematopoeisis and myogenesis as well as neurodevelopment.…”

Section: The Presenilins and γ-Secretasementioning

confidence: 99%

Protein aggregates and dementia: is there a common toxicity?

Lovestone

2002

Journal of Neurology, Neurosurgery &Amp; Psychiatry

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“…Perhaps the most striking biochemical result came from the use of radiolabeled photoactivatable transition state analog inhibitors; the only proteins to be labeled in a membrane preparation enriched in γ-secretase activity were the PS1 cleavage fragments (36,37). Detergent solubilization and immunoprecipitation of PS1 from such a membrane preparation revealed clear coprecipitation with γ-secretase activity (38). While ultimate proof will consist of reconstituting γ-secretase activity from recombinant proteins, this series of experiments appears to many to be definitive evidence that PS1 is γ-secretase.…”

Section: γ-Secretase As a Therapeutic Targetmentioning

confidence: 99%

Spotlight on BACE: The secretases as targets for treatment in Alzheimer disease

Dingwall¹

2001

J. Clin. Invest.

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Alzheimer disease (AD) is a progressive degenerative disease of the brain that is characterized by neocortical atrophy, neuron and synapse loss, and the presence of extracellular senile plaques and intracellular neurofibrillary tangles (NFTs). The primary clinical manifestation of AD is a profound global dementia that is marked by severe amnesia with additional deficits in language, "executive" functions, attention, and visiospatial and constructional abilities. The neurodegenerative changes occur primarily in the hippocampus and entorhinal cortex and in the association cortices of the frontal, temporal, and parietal lobes. Although the temporal progression of the neuropathological changes of AD is not fully known, recent studies suggest that the hippocampus and entorhinal cortex are involved in the earliest stage of the disease, and that frontal, temporal, and parietal association cortices develop pathology as the disease progresses. This "spreading" of the pathology from those regions of the brain in which hallmarks of the disease (amyloid plaques, reactive gliosis, NFTs) can be first detected, to other regions of the brain is notable, and while generally accepted as being a genuine feature of the pathology, no explanation for it has yet emerged (1).As the population ages, the projected number of individuals that will be affected by dementia, and AD in particular, indicates that a serious public health problem is looming. However, intense research over the past decade has begun to uncover some of the cell and molecular processes leading to neuronal loss with the discovery of possible targets for therapeutic intervention, raising the hope that we may be able at least to halt the progression of the disease (2, 3).The major constituent of senile plaques is the β-amyloid peptide, which is derived from the amyloid precursor protein (APP) by proteolytic cleavage (3). This peptide is invariably described as a 40-to 42-amino acid peptide, although numerous shorter x-40 and x-42 forms are found, particularly in the AD brain, and many of these peptides have a strong tendency to aggregate (4). By contrast, the intracellular NFTs are fibrillar aggregates of the microtubule-associated protein tau

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Presenilin 1 is linked with γ-secretase activity in the detergent solubilized state

Cited by 36 publications

References 12 publications

Purification and Characterization of the Human γ-Secretase Complex

Purification and Characterization of the Human γ-Secretase Complex

Protein aggregates and dementia: is there a common toxicity?

Spotlight on BACE: The secretases as targets for treatment in Alzheimer disease

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