Long-lived conformational isomerism of protein dimers: the role of the free energy of subunit association.

The association of protein subunits to form N-mers (N >or= 3) does not follow the dependence on the law of mass action predicted by the classical thermodynamic description used for the equilibrium of association of small molecules. For those anomalous cases, a so-called deterministic model has been previously proposed. The latter model was based on the empirical observation that the dynamics of subunit exchange between protein oligomers can be very slow, leading to the existence of long-lived conformational isomers and to a persistently heterogeneous ensemble of oligomers in solution. Contrary to the expectation for a protein dimer, we have recently shown that the subunit association of triosephosphate isomerase (TIM) could also be described as a deterministic process and that long-lived conformational isomers of TIM could be isolated in solution. Here we show that a), observation of hysteresis in pressure dissociation curves is an additional indicator of deterministic behavior; b), the extent of deviation from the classical thermodynamic behavior correlates with the free-energy change of subunit association; and c), experimental manipulation of the free energy of subunit association through the addition of a subdenaturing concentration of a chaotropic agent restores the concentration dependence of subunit association of TIM. A model that explains these features and its biological relevance is discussed.