Many debilitating human diseases are associated with the extracellular misfolding and aggregation of globular proteins to form fibrillar deposits that are rich in ?-sheet. There is considerable evidence that the process is initiated by local unfolding of the native structure to form aggregation-prone amyloidogenic intermediates. Aggregation then proceeds via nucleation-growth or downhill polymerization mechanisms. Despite their key role in amyloidogenesis, influencing the kinetic partitioning between aggregation and refolding pathways, very little is known about the structure of amyloidogenic intermediates because of their strong propensity to aggregate. The goals of the present proposal are to apply state-of-the-art NMR methods to elucidate the fundamental molecular events involved in initiation of transthyretin amyloidogenesis. Transthyretin amyloidosis is associated with numerous neurodegenerative diseases and cardiomyopathies. Misfolding and aggregation of transthyretin leads to fibrous deposits in the peripheral nerves and heart. Deposition of wild type protein is age related, whereas the familial diseases associated with genetic mutations that destabilize the quaternary and/or tertiary structure are early onset. This research will provide novel insights into the fundamental molecular mechanisms by which familial mutations destabilize the native transthyretin tetramer and initiate the aggregation cascade. Recently developed NMR relaxation dispersion experiments will be utilized to elucidate the structure of an aggregation-prone, early amyloidogenic intermediate and determine the kinetics and thermodynamics of the processes by which it is formed by partial unfolding of the monomeric transthyretin subunit.

Public Health Relevance

The proposed research will address the fundamental molecular mechanisms by which transthyretin misfolds and aggregates to cause human disease. Transthyretin amyloidosis plays a causative role in numerous age-related and familial neurodegenerative diseases and cardiomyopathies. This research will provide novel insights into the fundamental molecular mechanisms by which wild type transthyretin unfolds to form an aggregation-prone intermediate, and by which familial mutations destabilize the native transthyretin tetramer to initiate the aggregation cascade.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK034909-26
Application #
8642171
Study Section
Macromolecular Structure and Function B Study Section (MSFB)
Program Officer
Bishop, Terry Rogers
Project Start
1984-12-01
Project End
2018-03-31
Budget Start
2014-04-01
Budget End
2015-03-31
Support Year
26
Fiscal Year
2014
Total Cost
$412,163
Indirect Cost
$194,663
Name
Scripps Research Institute
Department
Type
DUNS #
781613492
City
La Jolla
State
CA
Country
United States
Zip Code
92037
Sun, Xun; Jaeger, Marcus; Kelly, Jeffery W et al. (2018) Mispacking of the Phe87 side chain reduces the kinetic stability of human transthyretin. Biochemistry :
Sun, Xun; Dyson, H Jane; Wright, Peter E (2018) Kinetic analysis of the multistep aggregation pathway of human transthyretin. Proc Natl Acad Sci U S A 115:E6201-E6208
Leach, Benjamin I; Zhang, Xin; Kelly, Jeffery W et al. (2018) NMR Measurements Reveal the Structural Basis of Transthyretin Destabilization by Pathogenic Mutations. Biochemistry 57:4421-4430
Sun, Xun; Dyson, H Jane; Wright, Peter E (2017) Fluorotryptophan Incorporation Modulates the Structure and Stability of Transthyretin in a Site-Specific Manner. Biochemistry 56:5570-5581
Dyson, H Jane; Wright, Peter E (2017) How Does Your Protein Fold? Elucidating the Apomyoglobin Folding Pathway. Acc Chem Res 50:105-111
Lim, Kwang Hun; Dasari, Anvesh K R; Hung, Ivan et al. (2016) Solid-State NMR Studies Reveal Native-like ?-Sheet Structures in Transthyretin Amyloid. Biochemistry 55:5272-8
Aoto, Phillip C; Nishimura, Chiaki; Dyson, H Jane et al. (2014) Probing the non-native H helix translocation in apomyoglobin folding intermediates. Biochemistry 53:3767-80
Li, Xinyi; Zhang, Xin; Ladiwala, Ali Reza A et al. (2013) Mechanisms of transthyretin inhibition of ?-amyloid aggregation in vitro. J Neurosci 33:19423-33
Lim, Kwang Hun; Dyson, H Jane; Kelly, Jeffery W et al. (2013) Localized structural fluctuations promote amyloidogenic conformations in transthyretin. J Mol Biol 425:977-88
Nishimura, Chiaki; Dyson, H Jane; Wright, Peter E (2011) Consequences of stabilizing the natively disordered f helix for the folding pathway of apomyoglobin. J Mol Biol 411:248-63

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