Full-length cellular β-secretase has a trimeric subunit stoichiometry, and its sulfur-rich transmembrane interaction site modulates cytosolic copper compartmentalization

• Published in: The Journal of biological chemistry Vol. 292; no. 32; pp. 13258 - 13270
• Main Authors: Liebsch, Filip, Aurousseau, Mark R.P., Bethge, Tobias, McGuire, Hugo, Scolari, Silvia, Herrmann, Andreas, Blunck, Rikard, Bowie, Derek, Multhaup, Gerd
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 11.08.2017; American Society for Biochemistry and Molecular Biology
• Subjects: Alanine - chemistry; Amino Acid Substitution; Amyloid Precursor Protein Secretases - antagonists & inhibitors; Amyloid Precursor Protein Secretases - chemistry; Amyloid Precursor Protein Secretases - genetics; Amyloid Precursor Protein Secretases - metabolism; Aspartic Acid Endopeptidases - antagonists & inhibitors; Aspartic Acid Endopeptidases - chemistry; Aspartic Acid Endopeptidases - genetics; Aspartic Acid Endopeptidases - metabolism; Bacterial Proteins - chemistry; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; beta-secretase 1 (BACE1); Biological Transport; Cell Biology; Copper - metabolism; copper transport; Cysteine - chemistry; Cytosol - metabolism; Fluorescence Resonance Energy Transfer; Green Fluorescent Proteins - chemistry; Green Fluorescent Proteins - genetics; Green Fluorescent Proteins - metabolism; HEK293 Cells; Humans; Luminescent Proteins - chemistry; Luminescent Proteins - genetics; Luminescent Proteins - metabolism; Microscopy, Fluorescence; Models, Molecular; Mutagenesis, Site-Directed; Point Mutation; Protein Conformation; protein cross-linking; Protein Folding; Protein Interaction Domains and Motifs; Protein Multimerization; Recombinant Fusion Proteins - chemistry; Recombinant Fusion Proteins - metabolism; RNA Interference; single-molecule biophysics; stoichiometry; single-molecule biophysics; stoichiometry; protein cross-linking; beta-secretase 1 (BACE1); copper transport
• DOI: 10.1074/jbc.M117.779165
• ISSN: 0021-9258

The β-secretase (BACE1) initiates processing of the amyloid precursor protein (APP) into Aβ peptides, which have been implicated as central players in the pathology of Alzheimer disease. BACE1 has been described as a copper-binding protein and its oligomeric state as being monomeric, dimeric, and/or multimeric, but the native cellular stoichiometry has remained elusive. Here, by using single-molecule fluorescence and in vitro cross-linking experiments with photo-activatable unnatural amino acids, we show that full-length BACE1, independently of its subcellular localization, exists as trimers in human cells. We found that trimerization requires the BACE1 transmembrane sequences (TMSs) and cytoplasmic domains, with residues Ala463 and Cys466 buried within the trimer interface of the sulfur-rich core of the TMSs. Our 3D model predicts that the sulfur-rich core of the trimeric BACE1 TMS is accessible to metal ions, but copper ions did not trigger trimerization. The results of functional assays of endogenous BACE1 suggest that it has a role in intracellular copper compartmentalization by transferring cytosolic copper to intracellular compartments, while leaving the overall cellular copper concentration unaltered. Adding to existing physiological models, our results provide novel insight into the atypical interactions between copper and BACE1 and into its non-enzymatic activities. In conclusion, therapeutic Alzheimer disease prevention strategies aimed at decreasing BACE1 protein levels should be regarded with caution, because adverse effects in copper homeostasis may occur.