Mutagenic Alkylating Agents 305 



with a positively and a negatively charged synthetic polymer 

 (polyethylene imine and polymethacrylic acid respectively) 

 show that reaction is very largely influenced by electrostatic 

 repulsion or attraction between the reactants. 



Reaction vs^ith nucleoproteins 



In the cell, of course, DNA is not present as such but com- 

 bined with proteins which will reduce its electrical charge. 

 Consequently we decided to study the reaction of a nucleo- 

 protein with the radiomimetic alkylating agent. Since the 

 extraction of nuclear proteins from the cell gives ill defined 

 products which are difficult to handle, we used sperm heads 

 obtained by plasmolysing soft roe from herrings (Felix, Fischer 

 and Krekels, 1956). These are hard balls with a diameter of 

 the order of 2 [x, containing only DNA and the low-molecular 

 w^eight proteins, the protamines, in the ratio of 39 per cent to 

 61 per cent. They are surrounded by a tough membrane the 

 weight of which is insignificant. When placed in water they 

 swell only to a very limited extent and take up less than their 

 own weight of water. Suspensions of sperm heads can there- 

 fore be handled easily and they are not at all viscous. 



The reaction of the sperm heads with the alkylating agents 

 can be followed on the automatic titrator in exactly the same 

 way as with the solution of DNA. From the amount of acid 

 liberated the esterification of the phosphate groups can be 

 determined and is seen to occur extremely readily (Fig. 4). 

 For the aromatic mustards it could again be shown that the 

 only reaction which occurred was esterification of acid groups 

 and there was no evidence of alkylation of amino or imino 

 groups in either the protein or the nucleic acid. Since the 

 number of carboxyl groups in the protein is very much less 

 than that of the phosphate groups from DNA the esterifica- 

 tion of the latter would again appear to be the predominant 

 reaction. Other evidence (see p. 311) supports the view that 

 the reaction of the mustard is confined to DNA and that the 

 protamine is not involved. 



