Heat capacity and thermodynamic characteristics of denaturation and glass transition of hydrated and anhydrous proteins

Thermodynamic and dynamic factors involved in the stability of native protein structure in amorphous solids in relation to levels of hydration

“…131 Interestingly, the respective values increased with an increase in the protein ratio, indicating that the T g value of the protein is greater than that of the sugars, and extrapolation of the data to zero sucrose concentration indicates a T g of about 1508C for the native protein state, which is a value similar to those recovered for the unfolded and aggregated proteins described above. 123,128,[132][133][134] That sugars and proteins indeed interact in the amorphous state to form molecular dispersions is also supported by FTIR measurements 135 and by the observation that the presence of protein with sucrose in lyophilized samples prevents crystallization of sucrose, 136 just as has been observed when PVP was mixed with sucrose 121 and other small molecules. 120 If we assume that a lowering of the overall T g in a molecular dispersion, relative to that of the protein alone, would ordinarily increase the molecular mobility of the protein at a given temperature, it would then appear that the sugar would not have the ability to stabilize the system as suggested in the ''vitrification'' model used to explain the stabilizing effects of sugars as lyoprotectants.…”

“…In Figure 3 we depict DSC scans of the type reported for samples of a folded protein, legumin in the amorphous state. 123 For the first scan, the ''dry'' or ''wet'' protein exhibits no discontinuity reflective of a DC p , and hence a T g , right up to the point where the characteristic thermal denaturation endotherm at T m is observed. However, when the denatured protein sample was cooled and then re-scanned, a clear indication of a DC p appears.…”

“…116 It would appear therefore that a-like motions, which are activated at T d , are not obvious by DSC measurements due to the continuous buildup of the lower energy, b-like motions through T d , where ultimately the two motions merge, giving rise to an apparent small change in DC p . 129,195 This behavior may help to explain the apparent fragility and ''T g ,'' determined by DSC measurements, for denatured aggregated proteins; which were generated by first unfolding the native protein with heat followed by cooling, such that the resulting protein structures are an assembly of non-native states 123,196 (Fig. 3).…”

Solid state chemistry of proteins IV. what is the meaning of thermal denaturation in freeze dried proteins?

“…for unfolding, as the geometric mean of the ratio k r =k m 1 is about 10 6 . Most important, it should be noted that this ratio is by no means constant for the systems studied; variation is from 10 5 to 10 8 .…”

“…Since the thermal unfolding in systems that are solid (glassy) at ambient temperature is completely irreversible and does not take place until the temperature far exceeds the glass transition temperature of the system, 1,[3][4][5][6][7][8] it is tempting to conclude that the unfolding event is under complete kinetic control. In fact, even studies with proteins in dilute solution 10 often display highly irreversible unfolding.…”