Unfolding isotherm

Folded proteins can be caused to spontaneously unfold upon being exposed to chaotropic agents, such as urea or guanidine hydrochloride (Gdn), or to elevated temperature (thermal denaturation). As solution conditions are changed by addition of denaturant, the mole fraction of denatured protein increases from a minimum of zero to a maximum of 1.0 in a characteristic unfolding isotherm (Fig. 7a). From a plot such as Figure 7a one can determine the concentration of denaturant, or the temperature in the case of thermal denaturation, required to achieve half maximal unfolding, ie, where... 

If an oligomeric protein has been shown to unfold reversibly in a two-state equilibrium, if the m-value does not depend on temperature and is reliably determined, and if all unfolding isotherms were measured at the same total protein concentration, then a plot of the C value yielded from all binding isotherms vs T can be fitted to a modified Gibbs-Helmholtz equation (cf. ref. 6)... 

Proteins often have the same high-affinity isotherms as do synthetic polymers and are also slow to equilibrate, due to many contacts with the surface. Proteins, however, have the additional complication that they can partially or completely unfold at the solid-liquid interface to expose their hydrophobic core units to a hydrophobic surface... 

Fig. 7. Unfolding (a) isotherm, where the half-maximal unfolding for this protein occurs at 2.6 M denaturant and (b) free energy where in the absence of denaturant, the protein has an extrapolated stability,, of 17.6 kj/mol (4.2 kcal/mol) as shown. To convert to cal, divide by 4.184.

Both Reynolds and Karim worked at neutral pH, with denatured proteins, and with reduced disulfide bonds. Under these conditions, proteins are in a random coil conformation (Mattice et al., 1976), so that their hydrodynamic radius is monotoni-cally related to their molar mass. Takagi et al. (1975) reported that the binding isotherm of SDS to proteins strongly depends upon the method of denaturing disulfide bonds. Presumably, protein-SDS complexes are not fully unfolded when disulfide bonds are left intact, which breaks the relationship between molar mass and hydrodynamic... 

Christensen, T., B. Svensson, and B.W. Sigurskjold. 1999. Thermodynamics of reversible and irreversible unfolding and domain interactions of glucoamylase from Aspergillus niger studied by differential scanning and isothermal titration calorimetry. Biochemistry 38 6300-6310. 

We now turn to the non-isothermal reaction system in a non-adiabatic CSTR, as studied in 7.2.4—6. We begin with the simplified model with exponential approximation to the Arrhenius law, and to systems for which the inflow and ambient temperatures are the same (y = 0 and gc = 0), This system has two unfolding parameters gad and rN. The stationary-state equation and its various derivatives are... 

Fig. 9.4. Specific volume of Snase as a function of pressure at 40°C [15], The protein is folded up to 50 MPa, and the slope up to that pressure is indicative of the isothermal compressibility of the folded state. The arrow at 100 MPa indicates the volume change of unfolding assuming constant compressibility of the folded state and nearly complete unfolding by 100 MPa. Unfortunately, the high-pressure densitometer was limited to 100 MPa, so the compressibility of the unfolded state could not be determined...

Figure 6. Equilibrium denaturation isotherms for Pal-HI (solid symbols) anrt HT t symbols) in pH 7.5 buffer containing 20% EtOH determined from the ellipticity at 224 nm°The fraction of protein unfolded is plotted as a function of guanidine hydrochloride concentration...

At fluid/fluid interfaces, it is well established that proteins lose their tertiary structure. Measurements of II — A isotherms give areas that can be ascribed to unfolded polypeptide chains e.g., limiting... 

It is seen that the calculated deviation for the reorientation model is two times lower than for the Frumkin model. The dependencies of the isotherm parameters of C TAB on n are similar to those obtained for other surfactants the Frumkin constant a increases with n (cf Table 3.21), the minimum area of the surface active ion C0 2 is almost independent of n, and the molar area in the unfolded state [Pg.246]

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