Better biomolecule thermodynamics from kinetics

Kiran Girdhar, University of Illiinois
Gregory Scott, University of Illinois
Yann R. Chemla, University of Illinois
Martin Gruebele, University of Illinois

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7 pages.

Copyright © 2011 American Institute of Physics. This article may be downloaded for personal use only. Any other use requires prior permission of the author and the American Institute of Physics. The following article appeared in Journal of Chemical Physics and may be found at http://dx.doi.org/10.1063/1.3607605.

NOTE: At the time of publication, the author Gregory Scott was not yet affiliated with Cal Poly.

Abstract

Protein stability is measured by denaturation: When solvent conditions are changed (e.g., temperature, denaturant concentration, or pH) the protein population switches between thermodynamic states. The resulting denaturation curves have baselines. If the baselines are steep, nonlinear, or incomplete, it becomes difficult to characterize protein denaturation. Baselines arise because the chromophore probing denaturation is sensitive to solvent conditions, or because the thermodynamic states evolve structurally when solvent conditions are changed, or because the barriers are very low (downhill folding). Kinetics can largely eliminate such baselines: Relaxation of chromophores, or within thermodynamic states, is much faster than the transition over activation barriers separating states. This separation of time scales disentangles population switching between states (desired signal) from chromophore or population relaxation within states (baselines).We derive simple formulas to extract unfolding thermodynamics from kinetics. The formulas are tested with model data and with a difficult experimental test case: the apparent two-state folder PI3K SH3 domain. Its melting temperature T_m can be extracted reliably by our “thermodynamics from kinetics approach,” even when conventional fitting is unreliable.