48 CHARLES TANFORD [3 



this effect is provided by the viscosity data of Arnold and Overbeek^ for 

 polymethacrylic acid, which are shown in Fig. 3. 



An increase in ionic strength has a much greater effect on electrostatic 

 interactions in configurations in which the molecular domain is permeated 

 by solvent than it does for the impenetrable configuration to which the cal- 

 culation of Table 2 applies. As a result, a moderate ionic strength suffices 

 to eliminate electrostatic forces almost entirely with a resultant return to 

 essentially the randomly coiled configuration of the uncharged molecule. 

 This effect is also illustrated by Fig. 3. 



0.2 0.4 0.6 



DEGREE OF DISSOCIATION 



Fig. 3. Reduced viscosities in dilute solutions of polymethacrylic acid (Arnold and 

 Overbeek^). Ionic strength is 0010 to 0-017 for the top curve, 0-10 for the middle curve, 

 and 10 for the lowest one. The concentration c was 000085 g./cc. in all experiments. 



Exactly parallel behavior on the part of globular proteins is, of course, 

 not to be expected. Isoelectric polymethacrylic acid is randomly coiled, i.e. 

 all possible configurations have approximately the same energy. Thus, even 

 small energy changes in the more compact configurations are immediately 

 (and reversibly) reflected in an increase in the proportion of molecules with 

 more extended configurations. In the globular proteins here under discus- 

 sion, on the other hand, the various possible configurations in the isoelectric 

 state do not have identical energy. Instead, the most compact configuration 

 is stabiHzed by what may be a large energy inherent in the various forms 

 of intramolecular attraction described earlier in this paper. The effect of 

 titration is to make the electrostatic contribution to the free energy a posi- 

 tive one (and, perhaps, to change the strength of specific bonds between 

 side chain groups). Only when the resulting change in free energy becomes 



