The long-term objective of this research project is to develop a detailed atomic-resolution structural and functional understanding of protein aggregation and fibrillation relevant to human disease, principally with studies of alpha-synuclein. Wild type alpha-synuclein and its mutants (A30P, E46K and A53T) associated with early-onset Parkinson's disease (PD) will be examined in several structural states, including protofibrils, fibrils and membrane-associatedcomplexes. We propose that the chemical details of these structures determine the natural physiological function of alpha-synuclein and, in the mutants or under environmental stress, contribute to PD pathology. Moreover, we propose that conversion among several structural states is an essential feature of synucleins; this structural plasticity is likely to be required for neuronal development and maintenance. To examine these structures, magic-angle spinning (MAS) solid- state NMR (SSNMR) experiments will be employed, elucidating details of covalent chemistry, conformation and dynamics that are inaccessible to other experimental techniques. Complementary analysis and characterization techniques will include kinetic measurements (by thioflavin T fluorescence, light scattering and sedimentation analysis), high-resolution mass spectrometry, atomic force and electron microscopy, solution NMR and phage display. We will leverage these technologies along with sample preparation and isotopic labeling strategies to improve our understanding of protein aggregation relevant to disease in the specific case of alpha-synuclein, including: (1) the fundamental biophysical principles of alpha-synuclein aggregation and membrane association, (2) specific structural information that will enable rational design strategies for small molecules that modulate alpha-synuclein function and/or serve as diagnostic tools for PD, and (3) elucidation of specific structural interactions between alpha-synuclein and other brain proteins. Relevance to public health: Alpha-synuclein is a central player in Parkinson's disease. The precise relationships between alpha-synuclein structure and this disease are not yet well understood. Our proposed studies aim to advance this fundamental knowledge, which will assist the broader research community in developing improved diagnostic tools and therapies for Parkinson's disease.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
3R01GM073770-02S1
Application #
7535413
Study Section
Macromolecular Structure and Function C Study Section (MSFC)
Program Officer
Wehrle, Janna P
Project Start
2007-01-15
Project End
2010-12-31
Budget Start
2008-01-01
Budget End
2008-12-31
Support Year
2
Fiscal Year
2008
Total Cost
$49,450
Indirect Cost
Name
University of Illinois Urbana-Champaign
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
041544081
City
Champaign
State
IL
Country
United States
Zip Code
61820
Barclay, Alexander M; Dhavale, Dhruva D; Courtney, Joseph M et al. (2018) Resonance assignments of an ?-synuclein fibril prepared in Tris buffer at moderate ionic strength. Biomol NMR Assign 12:195-199
Courtney, Joseph M; Rienstra, Chad M (2016) Efficient dipolar double quantum filtering under magic angle spinning without a (1)H decoupling field. J Magn Reson 269:152-156
Hisao, Grant S; Harland, Michael A; Brown, Robert A et al. (2016) An efficient method and device for transfer of semisolid materials into solid-state NMR spectroscopy rotors. J Magn Reson 265:172-6
Tuttle, Marcus D; Comellas, Gemma; Nieuwkoop, Andrew J et al. (2016) Solid-state NMR structure of a pathogenic fibril of full-length human ?-synuclein. Nat Struct Mol Biol 23:409-15
Tuttle, Marcus D; Courtney, Joseph M; Barclay, Alexander M et al. (2016) Preparation of Amyloid Fibrils for Magic-Angle Spinning Solid-State NMR Spectroscopy. Methods Mol Biol 1345:173-83
Shi, Xiangyan; Rienstra, Chad M (2016) Site-Specific Internal Motions in GB1 Protein Microcrystals Revealed by 3D ²H-¹³C-¹³C Solid-State NMR Spectroscopy. J Am Chem Soc 138:4105-19
Courtney, Joseph M; Ye, Qing; Nesbitt, Anna E et al. (2015) Experimental Protein Structure Verification by Scoring with a Single, Unassigned NMR Spectrum. Structure 23:1958-1966
Lin, Wayne; Insley, Thomas; Tuttle, Marcus D et al. (2015) Control of protein orientation on gold nanoparticles. J Phys Chem C Nanomater Interfaces 119:21035-21043
Rienstra, Chad M (2013) Amyloid structures from Alzheimer's disease patients. Structure 21:1722-3
Tang, Ming; Nesbitt, Anna E; Sperling, Lindsay J et al. (2013) Structure of the disulfide bond generating membrane protein DsbB in the lipid bilayer. J Mol Biol 425:1670-82

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