The physical basis of protein folding defines a second """"""""genetic code"""""""". Interest in mechanisms of folding has recently been stimulated by the discovery of human diseases caused by the misfolding and aggregation of diverse proteins. Although a protein's native state is encoded in its amino-acid sequence, chaperonin-assisted protein folding is universal among life's three Kingdoms. How chaperonins interact with and assist in the folding of nascent polypeptides represent problems of fundamental significance in cell biology and biophysics. This application proposes a novel strategy to study a model mini-chaperonin -- the apical domain of GroEL - by exploiting a thermophilic homologue. The essential idea is to use temperature (rather than chemical denaturants) to unfold mesophilic protein substrates in the presence of native mini-chaperonin. The thermophilic apical domain is derived from the bacterium Thermus thermophilus and retains native structure up to 80 degree C. This strategy will enable us to test the hypothesis that the apical domain contains a single and universally conserved surface for protein-substrate recognition. The proposed approach of study employs an interdisciplinary combination of synchrotron-based X-ray crystallography at cryogenic temperature and high-resolution MNR at physiologic temperature. Function will be correlated with structure and dynamics by mutagenesis and biochemical analysis of in vitro chaperonin-assisted protein-folding reactions. Taken together, the proposed studies promise to provide an atomic-level view of a fundamental step in chaperonin-assisted protein folding.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM061446-02
Application #
6181479
Study Section
Molecular and Cellular Biophysics Study Section (BBCA)
Program Officer
Wehrle, Janna P
Project Start
1999-09-01
Project End
2004-08-31
Budget Start
2000-09-01
Budget End
2001-08-31
Support Year
2
Fiscal Year
2000
Total Cost
$183,953
Indirect Cost
Name
Case Western Reserve University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
077758407
City
Cleveland
State
OH
Country
United States
Zip Code
44106