The interactions discussed in this dissertation have different effects on the folded and unfolded states. We are relatively familiar with the structure of the folded state of a protein; it is after all the subject of the PPS course. However, in order to discuss factors which lead to different degrees of stabilization between the folded and unfolded states, we need some sort of description of the unfolded state.
Until recently, most studies and theoretical treatments of protein stability have assumed that the unfolded state of a protein is a random coil, an ensemble of extended conformations, in which the protein chain is extensively hydrated and the individual residues do not interact with each other. This random coil ensemble can be treated as a single state for a thermodynamic description. One consequence of this view of the unfolded state is that any mutation that effects stability must have its effect almost exclusively via the native (folded) structure.
However, there is now mounting evidence that this simplistic view of the unfolded state is not always true (reviewed by Shortle, 1996). For example, small angle X-ray scattering studies have shown that the radius of gyration of the unfolded state of ribonuclease A is smaller than if it is in a random coil (Sosnick & Trewhella, 1992). Further, NMR studies have shown that interactions do occur between residues, particularly between aromatic residues, in the unfolded state and that these interactions can be non-native (e.g. Pan et al, 1995). In addition, there is strong evidence that mutations can affect the degree of interaction in the unfolded state (discussed in detail in Sturtevant, 1994). For example, in one mutant of Staphylococcal nuclease, the enthalpy of unfolding, H, was reduced to 50% that of wild-type, but with no concomitant change in G for the unfolding process (Tanaka et al, 1993). (Remember that). Although possible, to explain this result in terms of changes in the folded state alone is difficult: requiring (i) that over half the stabilizing interactions present in the wild-type be absent in the mutant and (ii) that there be a huge increase in the entropy of the (folded) mutant over the native protein. A more convincing explanation of the result is that the enthalpy and entropy changes between mutant and wild-type are due to changes in the unfolded state, where introduction of a few interactions in the unfolded mutant could significantly decrease both enthalpy and entropy vis-à-vis the wild-type. There is further evidence of this type from unfolding studies with chaotropic denaturants which suggest that, for a small subset of mutants, the amount of change in accessible surface area upon unfolding is different to that in wild-type protein (Shortle & Meeker, 1986).
However, due to the brevity of this dissertation, I will simplify and assume that all of the intramolecular Van der Waals and hydrogen bonds that stabilize the native state are fully disrupted in the unfolded protein.
Stability Defined The Major Factors Affecting Protein StabilityBeginning