92 MUTATIONS 



traorclinarily more efficient per amount incorporated than bromouracil? 



Freese: Let me start with bromouraciL I want to say that all we 

 tried to do was to make a correlation between, in this case, a chemical 

 theory and genetic observations. In order to prove our point com- 

 pletely we would have to make a sequence analysis of normal and 

 mutant DNA. But this is completely impossible at the present time. 



Figure 16 shows the normal pairing between adenine and bromo- 

 uracil. Occasionally, 5-bromouracil may either lose the hydrogen on 

 the 3-position, which means that it may be ionized or that it may 

 undergo a tautomeric shift, in which case it loses this hydrogen and 

 attaches one at the 4-position. In either of these two cases, it can 

 pair with guanine, when it is ionized by two hydrogen bonds, or when 

 it has undergone a tautomeric shift, with three hydrogen bonds, the 

 same hydrogen bonds which we observe in a guanine-cytosine pair. 

 This pairing should be perfectly normal with respect to the C-N dis- 

 tances and angles. 



Glass: Why can't ordinary uracil do that, then? 



Benzer: Ordinary uracil can't get in because it doesn't have a group 

 in the 5-position that the enzyme requires. 



Freese: It is very critical to have the right molecule at the 5-position. 

 Bromine has about the same Van der Weels radius as the methyl group 

 in thymine, and this is extremely important. If you take iodouracil or 

 chlorouracil, it does not get incorporated to the same extent, and 

 fiuorouracil does not get incorporated, to any measurable amount, into 

 DNA. 



Auerbach: Do the others produce mutations — the chlorouracil, for 

 instance? 



Freese: Iodouracil does, but I'm not sure about chlorouracil. The 

 question is, why is bromouracil mutagenic? We know that bromine is 

 more electronegative than the carbon in the methyl group, and there- 

 fore pulls the electrons out of the ring. In some way, not understood 

 in detail, the hydrogen at the 3-position thereby becomes more labile 

 than it is in thymine. I think I should really not say more about this 

 because it is all conjecture. 



One way to prove this view would be to employ compounds with 

 different groups at the 5-position which differ in their electronegativity 

 but do not differ very much in size (this is obviously quite difficult to 

 do) ; then one could see whether the frequency of mutations increases 

 with the electronegativity. 



Auerbach: Is this what you call mutagenesis by replication? 



Freese: Whenever the mistaken base pair bromouracil-guanine is 



