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Hydrogen exchange kinetics of proteins in denaturants: a generalized two-process model.

Authors
Type
Published Article
Journal
Journal of Molecular Biology
0022-2836
Publisher
Elsevier
Publication Date
Volume
286
Issue
2
Pages
607–616
Identifiers
PMID: 9973574
Source
Medline

Abstract

The recent progress in measurements on the amide hydrogen exchange (HX) in proteins under varying denaturing conditions, both at equilibrium and in transient relaxation, necessitates the development of a unifying theory which quantitatively relates the HX rates to the conformational energetics of the proteins. We present here a comprehensive kinetic model for the site-specific HX of proteins under varying solvent denaturing conditions based on the two-state protein folding model. The generalized two-process model considers both conformational fluctuations and residual protections, respectively, within the folded and unfolded states of a protein, as well as a global kinetic folding-unfolding transition between the two states. The global transition can be either rapid or slow, depending on the solvent condition for the protein. This novel model is applicable to the traditional equilibrium HX measurements in both EX2 and EX1 regimes, and also the recently introduced transient pulse-labeling HX experiments. A set of simple analytical equations is provided for quantitative interpretation of experimental data. The model emphasizes the use of full time-course of bi-exponential HX kinetics, rather than fitting time-course data to single rate constants, to obtain quantitative information about fluctuating conformers within the folded and unfolded states of proteins. This HX kinetic model naturally unfolds into a simple two-state and two-stage kinetic interpretation for protein folding. It suggests that the various observed intermediates of a protein can be interpreted as dominant isomers of either the folded or the unfolded state under different solvent conditions. This simple, minimalist's view of protein folding is consistent with various recent experimental observations on folding kinetics by HX.

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