The amyloidogenic pathway is widely believed to be closely linked to most forms of Alzheimer's disease. In this pathway the full length amyloid precursor protein (APP) is cleaved by ?-secretase to release a 99 residue transmembrane C-terminal domain known as """"""""C99"""""""". C99 is then cleaved by ?-secretase to release the amyloid-? (A?) polypeptides. There is a considerable body of data that elevated cholesterol in neuronal membranes promotes the amyloidogenic pathway, but there has not been a mechanistic explanation. In our recent work we have shown that C99 forms a specific 1:1 complex with cholesterol, with a dissociation constant well within the physiological concentration range of cholesterol in mammalian membranes. This observation, combined with a large body of literature evidence that the ?- and ?-secretases tend to be associated with cholesterol-rich membrane domains often referred to as """"""""lipid rafts"""""""", suggests a compelling hypothesis for how cholesterol promotes amyloidogenesis. We hypothesize that formation of a complex between cholesterol and C99 (or full length APP) results in enhanced partitioning of C99/APP to lipid rafts, where ?- and ?-secretase reside. This enhances the rate of amyloid-? production relative to conditions in which C99/APP is not complexed with cholesterol and the protein resides in bulk membranes.
Aims are:
Aim 1. Determine the structure of the C99/cholesterol complex in both bulk membranes and in """"""""lipid rafts"""""""".
This aim will provide the structural basis for molecular recognition of cholesterol by C99 and will also provide the first ever comparison of the structure of a membrane protein under model membrane conditions that mimic lipid rafts versus bulk membranes.
Aim 2. Elucidate the structural determinants in cholesterol that drive its association with C99. This will involve binding studies between C99 and a variety of cholesterol analogs/metabolites and will further illuminate the basis for molecular recognition between C99 and cholesterol. It will also provide a starting point for developing compounds that mimic cholesterol but that bind even more avidly to C99. Moreover, we will test the possibility that cholesterol analogs known to be """"""""raft-phobic"""""""" can compete effectively with cholesterol for binding to C99.
Aim 3. Determine whether binding of cholesterol to C99 and APP increases partitioning of these proteins into lipid rafts. Both giant unilamellar vesicles and cell-derived vesicles will be employed. These studies will test the hypothesis that association of cholesterol with the C99 and APP drives partitioning of these protein into rafts. Moreover, using selected compounds from Aim 2 that compete effectively with cholesterol but that have no avidity for rafts, we will also test whether raft association of C99/APP can be suppressed.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM106672-02
Application #
8642200
Study Section
Special Emphasis Panel (ZRG1)
Program Officer
Chin, Jean
Project Start
2013-05-01
Project End
2017-04-30
Budget Start
2014-05-01
Budget End
2015-04-30
Support Year
2
Fiscal Year
2014
Total Cost
Indirect Cost
Name
Vanderbilt University Medical Center
Department
Biochemistry
Type
Schools of Medicine
DUNS #
City
Nashville
State
TN
Country
United States
Zip Code
37212
Hutchison, James M; Lu, Zhenwei; Li, Geoffrey C et al. (2017) Dodecyl-?-melibioside Detergent Micelles as a Medium for Membrane Proteins. Biochemistry 56:5481-5484
Taylor, Keenan C; Sanders, Charles R (2017) Regulation of KCNQ/Kv7 family voltage-gated K(+) channels by lipids. Biochim Biophys Acta 1859:586-597
Cao, Zheng; Hutchison, James M; Sanders, Charles R et al. (2017) Backbone Hydrogen Bond Strengths Can Vary Widely in Transmembrane Helices. J Am Chem Soc 139:10742-10749
Kim, Ji-Hun; Schlebach, Jonathan P; Lu, Zhenwei et al. (2016) A pH-Mediated Topological Switch within the N-Terminal Domain of Human Caveolin-3. Biophys J 110:2475-2485
Schlebach, Jonathan P; Barrett, Paul J; Day, Charles A et al. (2016) Topologically Diverse Human Membrane Proteins Partition to Liquid-Disordered Domains in Phase-Separated Lipid Vesicles. Biochemistry 55:985-8
Li, Xinyi; Song, Yuanli; Sanders, Charles R et al. (2016) Transthyretin Suppresses Amyloid-? Secretion by Interfering with Processing of the Amyloid-? Protein Precursor. J Alzheimers Dis 52:1263-75
Mercredi, Peter Y; Bucca, Nadine; Loeliger, Burk et al. (2016) Structural and Molecular Determinants of Membrane Binding by the HIV-1 Matrix Protein. J Mol Biol 428:1637-55
Schlebach, Jonathan P; Sanders, Charles R (2015) The safety dance: biophysics of membrane protein folding and misfolding in a cellular context. Q Rev Biophys 48:1-34
Kroncke, Brett M; Vanoye, Carlos G; Meiler, Jens et al. (2015) Personalized biochemistry and biophysics. Biochemistry 54:2551-9
Kenworthy, Anne K (2015) Lipid Domains. Preface. Curr Top Membr 75:xiii-xvii

Showing the most recent 10 out of 15 publications