228 



C. B. ANFINSEN 

 Table 4 



[13 



DETERMINATION OF A^-TERMINAL AMINO ACIDS IN A TRYPSIN 

 DIGEST OF FULLY REDUCED, CARBOXYMETHYLATED LYSOZYME 



DIGESTION conditions: 1% LYSOZYME, 0-01% TRYPSIN, pH 8.0, 25°. 



PROTEOLYSIS COMPLETE (BY pHSTAT MEASUREMENT) AFTER 5 MINUTES. 



END-GROUP ANALYSIS BY METHOD OF LEVY^ 



In concluding these preliminary remarks about methods I should em- 

 phasize that a real measure of the value of the reductive cleavage of disulfide 

 bridges and of the trypsin digestion of carbobenzoxylated polypeptide chains 

 will become apparent only when these methods have been properly applied 

 to tryptophan-containing proteins larger than ribonuclease and lysozyme. 



To proceed with the main purpose of this discussion, as indicated in its 

 title, I should like now to consider some aspects of ribonuclease structure 

 in relation to biological and physical properties. As you have seen, the 

 sequential work on this protein is well along and the full sequence should 

 be forthcoming in a relatively short time. As all of us will agree, however, 

 a knowledge of amino acid sequence alone is of little use in the understand- 

 ing of three-dimensional structure and of the basic reasons for biological 

 activity. It has become increasingly evident that the activity of an enzyme, 

 for example, must ultimately be explained in terms of the secondary and 

 tertiary structure of the molecule, determined and rigidly fixed by covalent 

 cross linkages and by non-covalent bonds of differing strengths. 



The Rockefeller Institute group,^' and ourselves,^* have recently under- 

 taken independent investigations of the disulfide bridges of ribonuclease by 



* From sequence data of Thompson,^ W. Schroeder.^^ and R. Acher et al.^^ Amino 

 acids listed are those following either lysine or arginine in the sequences hsted in the 

 above papers. 



t Lysozyme contains 6 lysine and 1 1 arginine residues. Theoretical maximum number 

 of end-groups in a complete trypsin digest should, therefore, be 18. 



