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Three‐state protein folding: Experimental determination of free‐energy profile
Author(s) -
Baryshnikova Ekateri.,
Melnik Bogdan S.,
Finkelstein Alexei V.,
Semisotnov Gennady V.,
Bychkova Valentina E.
Publication year - 2005
Publication title -
protein science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.353
H-Index - 175
eISSN - 1469-896X
pISSN - 0961-8368
DOI - 10.1110/ps.051402705
Subject(s) - protein folding , chemistry , kinetic energy , intermediate state , folding (dsp implementation) , urea , native state , equilibrium unfolding , thermodynamics , chemical physics , crystallography , phase transition , physics , atomic physics , biochemistry , electrical engineering , engineering , quantum mechanics
When considering protein folding with a transient intermediate, a difficulty arises as to determination of the rates of separate transitions. Here we overcome this problem, using the kinetic studies of the unfolding/refolding reactions of the three‐state protein apomyoglobin as a model. Amplitudes of the protein refolding kinetic burst phase corresponding to the transition from the unfolded (U) to intermediate (I) state, that occurs prior to the native state (N) formation, allow us to estimate relative populations of the rapidly converting states at various final urea concentrations. On the basis of these proportions, a complicated experimental chevron plot has been deconvolved into the urea‐dependent rates of the I↔N and U↔N transitions to give the dependence of free energies of the main transition state and of all three (N, I, and U) stable states on urea concentration.