Synthesis of Nucleosides and Nucleotides 173 



are reported in Proc. Soc. exp. Biol., N.Y., 76, 464-5 (1951). We found 

 in all eases except one (in which we had an unexpectedly high value for 

 the purines) that 35-75 per cent of the activity in the purine is actually 

 contained in the 5-carbon atom, and in most cases the value from the 

 DNA is somewhat higher than the value from the PNA. This proves, 

 then, that to the extent of 35-75 per cent this glycine is being incor- 

 porated into the purine as glycine, and the work of Dr. Wilson and 

 Heinrich and also the early work of Carlson and Barker indicate that 

 presumably the remainder of the purine carbon is derived from formate. 

 This, I think, necessitates the postulation that there are two separate 

 pathways of DNA purine synthesis, one involving the incorporation of 

 adenine, which essentially does not exchange; the other involves these 

 smaller precursors, which apparently enter and leave the DNA, which 

 by chemical analysis by phosphorus incorporation is known not to be 

 turning over. Thus there is considerably more lability of the purine 

 of the DNA than there is of the phosphorus, and of course there is no 

 information yet on the ribose. At the present time we are doing a 

 rather extensive turnover study to try to find the time sequence of 

 these reactions. 



Hammarsten: I would like to ask Dr. Heidelberger if he extracted 

 all of the DNA from his tissue or only part of it. It might not be quite 

 homogeneous. 



Heidelberger: We feel that we have extracted in these procedures 

 about 75 per cent of the DNA, and in a separate experiment we found 

 identical specific activities in the purines of this extract and in com- 

 pletely extracted aliquots of the same sample. Of course, the work of 

 Chargaff and others has shown that DNA is probably not a homogeneous 

 compound, and we hope to try to study the differences in specific 

 activities in different parts of the DNA molecule. We do have con- 

 tamination of up to 10 per cent of RNA in our DNA sample, but in order 

 to invalidate these results we would have to have about 200 per cent 

 contamination. The contamination alone cannot be the explanation. 



Brown: I agree with Dr. Heidelberger. Contamination alone cannot 

 be the explanation. 



I would like to stand up for the Schmidt-Thannhauser method, a 

 modification of which we are using now, involving a salt extraction of 

 nucleic acid, precipitation with alcohol to get it away from protein, 

 then an alkaline digestion of the mixture of nucleic acids. You can 

 repeat the salt extraction until you are sure you have all the nucleic 

 acids out of the tissue. Dr. Roll is now using an alkahne Dowex column 

 which will separate five bases: adenine and guanine, as well as cytosine, 

 uracil and thymine. The nucleotide experiments which I described 

 were worked up by that method. There uracil and thymine are the 

 cross-contamination criteria and 1 per cent of either can be readily 

 picked up on the column, and each nucleic acid is free from the other 

 to within 0-4 per cent. We do not have that data on the nucleic acids 

 that were used for our PNA-DNA turnover studies, because at that time 

 we were depending on either a modified isotope dilution experiment or 

 on colorimetric methods, neither one of which were perfect. 



