On Determining the Chemical Structure of Proteins 287 



eluents, both of which can be removed by volatilization. Alternatively, 

 eluents containing sodium can be desalted by passage over a small 

 bed of NH 4 -Dowex 50, whereby Na is exchanged for the volatile 

 ammonium. 



Once the amino acid composition of the various peptides obtained 

 from peptic and chymotryptic digests of ribonuclease has been deter- 

 mined and correlated with the data obtained by the use of trypsin, 

 it should be possible to learn a great deal about the way the molecule 

 is constructed even before sequence studies on the individual peptides 

 have been performed.* Moreover, from the array of data thus pre- 

 sented, it should be possible with some assurance to choose for detailed 

 investigation those and only those peptides the amino acid sequence 

 of which will contribute the most to an understanding of the final 

 structure of the protein. It should also be possible to choose those 

 peptides the size and composition of which are most suitable for study 

 by existing techniques. In this manner the number of sequence deter- 

 minations required will be reduced to a minimum. There is not space 

 to go into the question of the actual determination of the sequence 

 of amino acids in peptides. If the peptides chosen for such an investi- 

 gation contain from four to ten amino acid residues, there is little 

 doubt that a sequence for each can be determined by one or a com- 

 bination of several existing methods. 



Structural analysis would be simplified greatly if there were avail- 

 able more proteolytic enzymes possessing a specificity as sharp as that 

 of trypsin, only directed towards other linkages, such as those involv- 

 ing proline, or histidine, or the aromatic amino acids. Possibly the 

 enzymologist will be able to come to the aid of the analyst and provide 

 the necessary tools. If, however, enzymes of the requisite specificity 

 cannot be found in nature, possibly they can be created artificially. 

 Microorganisms seem to be able to do almost anything, given sufficient 

 provocation. Perhaps with the aid of synthetic substrates they can 

 be induced to synthesize enzymes capable of hydrolyzing the particular 

 types of peptide linkages in which the protein chemist is interested. 



It should be emphasized that there is nothing extremely novel in 



* The separation and analysis of the peptides formed by the action of chymo- 

 trypsin has recently been completed by Dr. Hirs. On the basis of his data, it 

 has been possible to construct, as a working hypothesis, a tentative, partial 

 structural formula showing how the peptides present in the tryptic and chymo- 

 tryptic hydrolyzates might have originally been joined together in the molecule 

 of oxidized ribonuclease [Hirs, Stein, and Moore, /. Biol. Chem., in press]. The 

 expectations expressed in this paragraph have, therefore, been realized. 



