title: Development and characterization of a novel membrane assay for full-length BACE-1 at pH 6.0.
abstract: β-Site amyloid precursor protein cleaving enzyme-1 (BACE-1) is a transmembrane aspartic protease that mediates the initial cleavage of the amyloid precursor protein (APP), leading to the generation of amyloid-β (Aβ) peptides that are thought to be causative of Alzheimer's disease (AD). Consequently, inhibition of BACE-1 is an attractive therapeutic approach for the treatment of AD. In general, in vitro biochemical assays to monitor BACE-1 activity have used the extracellular domain of the protein that contains the catalytic active site. This form of BACE-1 is catalytically active at acidic pH and cleaves APP-based peptide substrates at the β-site. However, this form of BACE-1 does not mimic the natural physiology of BACE-1 and shows minimal activity at pH 6.0, which is more representative of the pH within the intracellular compartments where BACE-1 resides. Moreover, high-throughput screens with recombinant BACE-1 at pH 4.5 have failed to identify tractable leads for drug discovery, and hence, BACE-1 inhibitor development has adopted a rational drug design approach. Here we describe the development and validation of a novel membrane assay comprising full-length BACE-1 with measurable activity at pH 6.0, which could be used for the identification of novel inhibitors of BACE-1.
authors.affiliation: Global Research & Early Development, Merck Serono S.A., Geneva, Switzerland.
authors.forename: Arman
authors.initials: A
authors.lastname: Saric
authors.forename: Lars zur
authors.initials: Lz
authors.lastname: Brügge
authors.forename: Dirk
authors.initials: D
authors.lastname: Müller-Pompalla
authors.forename: Thomas
authors.initials: T
authors.lastname: Rysiok
authors.forename: Solenne
authors.initials: S
authors.lastname: Ousson
authors.forename: Bruno
authors.initials: B
authors.lastname: Permanne
authors.forename: Anna
authors.initials: A
authors.lastname: Quattropani
authors.forename: Michael
authors.initials: M
authors.lastname: Busch
authors.forename: Dirk
authors.initials: D
authors.lastname: Beher
authors.forename: Ishrut
authors.initials: I
authors.lastname: Hussain
mesh_terms: D000544:Alzheimer Disease / Q000188:drug therapy / Q000378:metabolism; D053829:Amyloid Precursor Protein Secretases / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000378:metabolism; D016282:Aspartic Acid Endopeptidases / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000378:metabolism; D020134:Catalytic Domain; D002462:Cell Membrane / Q000737:chemistry* / Q000378:metabolism*; D057809:HEK293 Cells; D057166:High-Throughput Screening Assays / Q000379:methods*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism
chemical_list: D011994:Recombinant Proteins; D053829:Amyloid Precursor Protein Secretases; D016282:Aspartic Acid Endopeptidases; C509280:BACE1 protein, human