A growing number of neurodegenerative diseases, including Alzheimer's disease (AD), Parkinson's disease, Huntington's disease, and Creutzfeldt-Jakob disease (CJD), involve the misfolding of normal proteins in the brain, which has recently been associated with the binding of metal ions such as iron, copper, and zinc. It is thought that metal dyshomeostasis is involved in protein misfolding and leads to oxidative damage and neuron degeneration. Yet, the functions of these metal ions and the misfolded proteins in the disease process are not well understood. To date, metal concentrations in brain tissue are generally measured via macroscopic (bulk) techniques, which cannot provide any spatial information on localized metal accumulation. Stains and/or fluorescently-tagged antibodies are used to identify misfolded proteins in tissue, but do not provide direct information on the protein's structure. Structural studies involving metal-binding and protein misfolding are primarily done in vitro. Thus, this proposal aims to bridge the gap between macroscopic methods of analyzing brain tissue and in vitro studies of metal-protein binding. It describes the development and utilization of a combination of spectroscopic imaging tools that directly investigate metal content and protein structure within intact tissues. The overall goal of this proposal is to obtain an in situ structural and mechanistic picture of how metal ions in the brain are involved in protein misfolding and aggregation in two protein-folding diseases: Alzheimer's disease and scrapie. We hypothesize that elevated concentrations of metal ions (notably Cu, Fe, and Zn) in the brain are involved in the disease pathogenesis. Using x-ray microspectroscopy, we will image the metal ion distribution, concentration, oxidation state, and local structure as a function of disease severity in intact brain tissue. By combining fluorescence microscopy and infrared microspectroscopy, we will image the location and secondary structure of the associated misfolded proteins. By combining these results, we will identify which metal ions accumulate before, concurrently, or after protein misfolding. We will test whether metal ion reduction is correlated with oxidation markers such as lipid peroxidation in the brain. Finally, we use the local structure and homogeneity of the in situ metal-protein complex to develop a possible mechanism for the complex formation and toxicity.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM066873-05
Application #
7194977
Study Section
Metallobiochemistry Study Section (BMT)
Program Officer
Fabian, Miles
Project Start
2003-03-15
Project End
2010-02-28
Budget Start
2007-03-01
Budget End
2010-02-28
Support Year
5
Fiscal Year
2007
Total Cost
$189,396
Indirect Cost
Name
Brookhaven National Laboratory
Department
Type
DUNS #
027579460
City
Upton
State
NY
Country
United States
Zip Code
11973
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Bourassa, Megan W; Miller, Lisa M (2012) Metal imaging in neurodegenerative diseases. Metallomics 4:721-38
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Miller, Lisa M; Dumas, Paul (2010) From structure to cellular mechanism with infrared microspectroscopy. Curr Opin Struct Biol 20:649-56
Leskovjan, Andreana C; Kretlow, Ariane; Miller, Lisa M (2010) Fourier transform infrared imaging showing reduced unsaturated lipid content in the hippocampus of a mouse model of Alzheimer's disease. Anal Chem 82:2711-6
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Leskovjan, Andreana C; Lanzirotti, Antonio; Miller, Lisa M (2009) Amyloid plaques in PSAPP mice bind less metal than plaques in human Alzheimer's disease. Neuroimage 47:1215-20
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Kretlow, Ariane; Wang, Qi; Beekes, Michael et al. (2008) Changes in protein structure and distribution observed at pre-clinical stages of scrapie pathogenesis. Biochim Biophys Acta 1782:559-65

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