OPTICAL PROPERTIES OF NUCLEIC ACIDS 



523 



. Forward titration from pH 7 U hotometric 



* Backtitration I 



* Forward titration from pH 7 U|,c,rometric 

 » Backtitration ' 



8 9 10 11 12 



pH 



Fig. 23. Spectrophotometric and electrometric titration data for calf thymus 

 DNA (Shack and Thompsett'") . (a represents the proportion of the high-e(P) form 

 of the nucleate, or the proportion of nucleate anion.) 



photometricalh^^^•^* Fig. 23, taken from a paper by Shack and Thomp- 

 sett,^" compares the electrometric and spectrophotometric titration curves 

 of DNA when titrated with alkaU. It can be seen from these results that 

 over the pH range 9-12 the increase in e(P) corresponds with ionization, 

 presumably of the hydroxyl groups of guanosine (pKa 9.5) and thymidine 

 (pKa 9.8). It is evident that the pK^ values of these groups are significantly 

 higher in the DXA macromolecule. 



On the acid side of neutrality comparative electrometric and spectro- 

 photometric titration data at pH values lower than 3.5 are lacking and such 

 a correspondence does not emerge. 



Fig. 24 (excluding curve 1) shows spectrophotometric titration curves 

 obtained in different laboratories ^^ 74.75 qj^ samples of calf thymus DNA in 

 absence of salts. These show that on titration of DNA with acid at the 

 lowest possible salt concentration, the e(P) of the nucleic acid begins to 

 rise at a pH as high as 8.0 and has almost attained its maximum value at 

 pH 5.0. A typical electrometric titration ".72,73 qj^ i\^q other hand shows that 

 in this pH region no ionic groups are being titrated. On the acid side of 

 pH 8.0, therefore, it appears that an increase in €(P) occurs, in the absence 

 of salts, at a higher pH than ionization. The effect of the addition of salts 



''* G. Frick, Biochim. et Biophys. Acta 8, 625 (1952). 



^* E. R. Blout and A. Asadourian, Biochim. et Biophys. Acta 13, 161 (1954). 



