We propose to study protein folding intermediates at two levels (a) under suitable equilibrium conditions where we can stabilize partially folded states of proteins and (b) by direct kinetic measurements using stopped-flow time-resolved SAXS. SAXS data yield overall size and shape indication of the conformational state of a protein. High-quality, high-angle scattering profiles for cytochrome c have been measured at varying denaturant concentrations. We will use the combination of singular value decomposition analysis and a denaturant binding model to determine whether an equilibrium intermediate state exists. This analysis follows the method used to distinguish a folding intermediate for lysozyme (Chen, J. Mol. Biol. 261, 658-671). The A-states, partially folded intermediate states obtained at low pH with the addition of salt, of apomyoglobin have been characterized with SAXS. The apomyoglobin A-states stabilized with TFA, TCA, and KCl demonstrated different degrees of compaction, suggesting that there is not a unique A-state.

Agency
National Institute of Health (NIH)
Institute
National Center for Research Resources (NCRR)
Type
Biotechnology Resource Grants (P41)
Project #
5P41RR001209-23
Application #
6586788
Study Section
Project Start
2002-03-01
Project End
2003-02-28
Budget Start
Budget End
Support Year
23
Fiscal Year
2002
Total Cost
$143,176
Indirect Cost
Name
Stanford University
Department
Type
DUNS #
800771545
City
Stanford
State
CA
Country
United States
Zip Code
94305
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