title: Competition between homodimerization and cholesterol binding to the C99 domain of the amyloid precursor protein.
abstract: The 99-residue transmembrane C-terminal domain (C99, also known as β-CTF) of the amyloid precursor protein (APP) is the product of the β-secretase cleavage of the full-length APP and is the substrate for γ-secretase cleavage. The latter cleavage releases the amyloid-β polypeptides that are closely associated with Alzheimer's disease. C99 is thought to form homodimers; however, the free energy in favor of dimerization has not previously been quantitated. It was also recently documented that cholesterol forms a 1:1 complex with monomeric C99 in bicelles. Here, the affinities for both homodimerization and cholesterol binding to C99 were measured in bilayered lipid vesicles using both electron paramagnetic resonance (EPR) and Förster resonance energy transfer (FRET) methods. Homodimerization and cholesterol binding were seen to be competitive processes that center on the transmembrane G₇₀₀XXXG₇₀₄XXXG₇₀₈ glycine-zipper motif and adjacent Gly709. On one hand, the observed Kd for cholesterol binding (Kd = 2.7 ± 0.3 mol %) is on the low end of the physiological cholesterol concentration range in mammalian cell membranes. On the other hand, the observed K(d) for homodimerization (K(d) = 0.47 ± 0.15 mol %) likely exceeds the physiological concentration range for C99. These results suggest that the 1:1 cholesterol/C99 complex will be more highly populated than C99 homodimers under most physiological conditions. These observations are of relevance for understanding the γ-secretase cleavage of C99.
authors.affiliation: Department of Biochemistry and Center for Structural Biology, Vanderbilt University School of Medicine, Nashville, Tennessee 37232, United States.
authors.forename: Yuanli
authors.initials: Y
authors.lastname: Song
authors.forename: Eric J
authors.initials: EJ
authors.lastname: Hustedt
authors.forename: Suzanne
authors.initials: S
authors.lastname: Brandon
authors.forename: Charles R
authors.initials: CR
authors.lastname: Sanders
mesh_terms: D020816:Amino Acid Motifs; D019943:Amino Acid Substitution; D053829:Amyloid Precursor Protein Secretases / Q000378:metabolism; D016564:Amyloid beta-Protein Precursor / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D002784:Cholesterol / Q000737:chemistry / Q000378:metabolism*; D019281:Dimerization; D004578:Electron Spin Resonance Spectroscopy; D031541:Fluorescence Resonance Energy Transfer; D005998:Glycine / Q000737:chemistry; D006801:Humans; D007700:Kinetics; D008051:Lipid Bilayers; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D054730:Protein Interaction Domains and Motifs; D017434:Protein Structure, Tertiary; D059748:Proteolysis; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism
chemical_list: D016564:Amyloid beta-Protein Precursor; D008051:Lipid Bilayers; D050505:Mutant Proteins; D010446:Peptide Fragments; D011994:Recombinant Proteins; C543304:amyloid beta-protein precursor C-terminal fragment beta, human; D002784:Cholesterol; D053829:Amyloid Precursor Protein Secretases; D005998:Glycine