Experimental measurement of binding energy, selectivity, and allostery using fluctuation theorems

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Science  27 Jan 2017:
Vol. 355, Issue 6323, pp. 412-415
DOI: 10.1126/science.aah4077

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Pulling macromolecules apart

Many biological processes involve macromolecular interactions, and knowing the binding energies of these interactions is key to a functional understanding. There are several experimental approaches to calculate binding energies in bulk solutions, but these require that the binding is at equilibrium. Bulk measurements may also mask different binding modes or concerted binding by several subunits. Camunas-Soler et al. used a fluctuation theorem for binding reactions to extract ligand binding energies directly from force experiments that probed a single binding reaction. They resolved binding energies of peptides to specific and non-specific DNA binding sites with affinities spanning six orders of magnitude.

Science, this issue p. 412


Thermodynamic bulk measurements of binding reactions rely on the validity of the law of mass action and the assumption of a dilute solution. Yet, important biological systems such as allosteric ligand-receptor binding, macromolecular crowding, or misfolded molecules may not follow these assumptions and may require a particular reaction model. Here we introduce a fluctuation theorem for ligand binding and an experimental approach using single-molecule force spectroscopy to determine binding energies, selectivity, and allostery of nucleic acids and peptides in a model-independent fashion. A similar approach could be used for proteins. This work extends the use of fluctuation theorems beyond unimolecular folding reactions, bridging the thermodynamics of small systems and the basic laws of chemical equilibrium.

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