OPTICAL PROPERTIES OF NUCLEIC ACIDS 525 



The lowered absorptivity of the bases, resulting in the low €(P) of DNA 

 is most plausibly interpreted as due to a disturbance of the tautomeric 

 equilibrium of one or more of the bases. It is difficult to envisage any other 

 type of explanation: no exception to Lambert's law has been recorded so 

 that the lowering of €(P) cannot be accounted for on the basis of "optical" 

 interference between chromophoric groups (i.e., screening affects^^). 



Purine and pyrimidine bases exhibit keto-enol tautomerism, and it has 

 been shown^-^ that free pyrimidines occur in a predominantly keto form, 

 although ionization to the anion may be presumed to involve an increase 

 in enolic character. [Cf. Jordan, Chapter 13.] The observation that the 

 electrometric and spectrophotometric titrations proceed concurrently in 

 the alkaline pH region suggests that hydrogen ions are being released from 

 the ionizable hydroxyl groups (with modified ^Ka values) of bases in a 

 predominantly enolic form. Such a degree of enolic character for the bases 

 in intact DNA would correspond, at least qualititively, to certain structures 

 for DNA proposed by Stern^^ and by Watson and Crick." In the structure 

 proposed by the latter, specific pairs of adjacent bases are hydrogen-bonded 

 through the 1,1- and 6,6-positions. [Compare Fig. 8 in Chapter 13.] The 

 effect of ionization is always to raise the value of €(P), but the salt effects 

 at constant pH indicate that the factors operating in the native macro- 

 molecule which are responsible for the absorption anomaly can also be 

 counteracted (in part reversibly) in other ways. 



4. Effect of Salts on the Absorptivity of DNA 



It has been known for some time that exposure of nucleic acids to ex- 

 tremes of pH, temperature, dehydration, etc. causes marked irreversible 

 changes in physical properties, including €(P). More recently the effects of 

 salts in particular on the €(P) value of DNA have been studied in several 

 laboratories. 



Thomas^^-''^ has shown that exposure of DNA to low sodium chloride con- 

 centration (<10~' M) results in irreversible changes in its structure. The 

 evidence for this may be summarized as follows. A DNA preparation, which 

 has never been exposed to low salt concentration, extremes of pH, or other 

 denaturing influences, when dissolved in sodium chloride solution (con- 

 centration > 10~^ M) has an e(P) value in the region of 6000. Such a solution 

 is quite stable for several days as judged by the constancy of e(P) and by 

 the fact that increasing the concentration of sodium chloride does not lower 

 the e(P) value. However, a solution of DNA in which the concentration 

 of sodium chloride is less than 10~^ M has an €(P) value considerably higher 

 than 6000, reaching a value of 8000 at a concentration of ca. 10~^ M . When 



76 K. G. Stern, Yale J. Biol. Med. 19, 937 (1947). 



" J. D. Watson and F. H. C. Crick, Nature 171, 737, 964 (1953). 



" R. Thomas, Biochim. et Biophys. Acta 14, 231 (1954). 



