Very little is currently known about the molecular mechanisms of protein aggregation. The increasing public concern about protein deposition diseases, both systemic and neurodegenerative, emphasize the growing recognition of the critical importance of protein aggregation and makes this a very important area for study. This proposal is aimed at elucidating the detailed molecular mechanisms of protein aggregation that lead to ordered aggregates such as amyloid fibrils, as well as disordered or amorphous aggregates such as inclusion bodies. Understanding the mechanisms of deposition will facilitate the rational design of clinical therapies that are based on inhibition of protein aggregation. There is a critical need for developing therapeutic solutions to protein deposition diseases. For this proposal we have chosen to focus on one particular protein system, the variable domain of the immunoglobulin light chain, VL, which has a number of features which make it well-suited for our purposes. Each year thousands of people die in the US from several types of light chain deposition disease. These disorders pose challenging biophysical questions regarding the relationship between protein stability, folding and aggregation. We hypothesize that partially-folded intermediates are the critical precursors to protein aggregation.
The specific aims of the proposal are (1) to determine why some proteins have a strong propensity to aggregate, and why some form amyloid and others amorphous aggregates, (2) to determine the mechanisms of protein aggregation, especially amyloid fibrils, and (3) to identify inhibitors of light chain aggregation and fibril formation, which could lead to the development of molecules of potential therapeutic value. Techniques to be used include mutagenesis, chromatography, kinetics, ATR FTIR spectroscopy, atomic force and electron microscopy, and other biophysical/biochemical methods.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK055675-03
Application #
6381518
Study Section
Molecular and Cellular Biophysics Study Section (BBCA)
Program Officer
Haft, Carol R
Project Start
1999-05-01
Project End
2003-04-30
Budget Start
2001-05-01
Budget End
2002-04-30
Support Year
3
Fiscal Year
2001
Total Cost
$227,994
Indirect Cost
Name
University of California Santa Cruz
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
City
Santa Cruz
State
CA
Country
United States
Zip Code
95064
Meng, Xiaoyun; Fink, Anthony L; Uversky, Vladimir N (2008) The effect of membranes on the in vitro fibrillation of an amyloidogenic light-chain variable-domain SMA. J Mol Biol 381:989-99
Qin, Zhijie; Hu, Dongmei; Zhu, Min et al. (2007) Structural characterization of the partially folded intermediates of an immunoglobulin light chain leading to amyloid fibrillation and amorphous aggregation. Biochemistry 46:3521-31
Khurana, Ritu; Coleman, Chris; Ionescu-Zanetti, Cristian et al. (2005) Mechanism of thioflavin T binding to amyloid fibrils. J Struct Biol 151:229-38
Uversky, Vladimir N; Fink, Anthony L (2004) Conformational constraints for amyloid fibrillation: the importance of being unfolded. Biochim Biophys Acta 1698:131-53
Zhu, Min; Han, Shubo; Zhou, Feimeng et al. (2004) Annular oligomeric amyloid intermediates observed by in situ atomic force microscopy. J Biol Chem 279:24452-9
Khurana, Ritu; Souillac, Pierre O; Coats, Alisa C et al. (2003) A model for amyloid fibril formation in immunoglobulin light chains based on comparison of amyloidogenic and benign proteins and specific antibody binding. Amyloid 10:97-109
Souillac, Pierre O; Uversky, Vladimir N; Fink, Anthony L (2003) Structural transformations of oligomeric intermediates in the fibrillation of the immunoglobulin light chain LEN. Biochemistry 42:8094-104
Zhu, Min; Souillac, Pierre O; Ionescu-Zanetti, Cristian et al. (2002) Surface-catalyzed amyloid fibril formation. J Biol Chem 277:50914-22
Goers, John; Permyakov, Sergei E; Permyakov, Eugene A et al. (2002) Conformational prerequisites for alpha-lactalbumin fibrillation. Biochemistry 41:12546-51