On Determining the Chemical Structure of Proteins 277 



apparent contradiction may be resolved by the observation 20 -- 1 that 

 traces of copper inhibit the enzyme strongly. It seems entirely possi- 

 ble that HCN and sulfhydryl compounds could enhance enzymatic 

 activity by sequestering inhibitory traces of copper that might be 

 present in either the enzyme, the substrate, or some of the reagents 

 used in the assay procedure. According to this view, copper could not 

 inhibit enzymatic activity by reacting with sulfhydryl groups of the 

 enzyme, which is the usual assumption advanced to explain inhibition 

 by copper, but must be bound at some other site in the molecule. The 



LNH 2 ] 



Fig. 1. Generalized gross structure of ribonuclease (from Anfinsen, Redfield, 

 Choate, Page, and Carroll 18 ). 



imidazole ring of one or more of the four histidine residues in the 

 enzyme seems a likely binding site, particularly in view of the findings 

 of ^'eil and Seibles 2 - that photooxidation of only one of the histidine 

 residues in ribonuclease leads to a 74% decrease in enzymatic activity. 

 If, in fact, a histidine residue is associated with the active center of 

 the enzyme, it seems possible that the studies now in progress might 

 reveal something of the structure associated with this active center. 

 If disulfide bridges exist in a protein molecule, it is desirable that 

 they be ruptured before partial degradation is attempted. The peptide 

 chains, if there is more than one, should then be separable, as they 

 were in the case of insulin." If, as in ribonuclease, only one chain 

 exists, hydrolysis of peptide bonds will not necessarily lead to a reduc- 



* The peptide chains of insulin were separable on the basis of solubility. To 

 separate the chains obtained from other proteins, techniques that have been 

 found useful for the separation of large peptides, such as countercurrenl distribu- 

 tion, or chromatography on IRC-50, or low-eross-linked polystyrene sulfonic acid 

 resins, may prove advantageous. 



