![]() 1A), show a strong dependence on guanidinium chloride (GdmCl) concentration, reflecting a very large unfolding reaction ( Eq. ![]() ( 11) These hydrogens, exemplified by Leu 98 ( Fig. The Δ G op values for the most slowly exchanging hydrogens in native cytochrome c can be computed ( Eq. The opening reactions that determine protein hydrogen exchange may expose much or little new surface. The slope ( m) depends on the additional denaturant sensitive surface exposed in the unfolding reaction. These appear to represent the major intermediates in the unfolding and refolding pathways of cytochrome c. Results for cytochrome c reveal a small sequence of distinct partially unfolded forms with progressively increasing free energy and degree of unfolding. From this we can infer, respectively, the structure, the free energy, and the surface exposure of each protein form. Hydrogen exchange experiments can then determine the hydrogens exposed in each higher energy form, their rates of exchange with solvent, and their sensitivity to the perturbant. The energy levels and therefore the occupation of these conformationally excited states can be manipulated by denaturants and temperature. The study of these partially unfolded forms (intermediates) may illuminate the fundamental cooperative nature of protein structure and define the unfolding and refolding pathways of a protein even though the intermediates are normally invisible to measurement. Under native conditions, a small fraction of any population of protein molecules occupies each possible higher energy, partially unfolded state, including even the fully unfolded state, as described by the Boltzmann distribution.
0 Comments
Leave a Reply. |
AuthorWrite something about yourself. No need to be fancy, just an overview. ArchivesCategories |