572 G. SCHMIDT 



the earlier results in the light of the recent information on the reaction 

 mechanism of the enzyme. 



Influence of pH. The pH optimum of crystallized pancreas ribonuclease 

 was found by Kunitz^ to be in the region of pH 7.7 (Fig. 6). Appreciable 

 spontaneous degradation of ribonucleic acids occur already at this pH in 

 experiments of longer duration. Many studies on ribonuclease action are 

 therefore carried out at slightly acid pH values in the range between 5 and 

 6 in which the enzyme has still considerable activity. 



Influence of Temperature. At pH 5, the optimal temperature of ribonu- 

 clease action is 65°.^ In the range between pH 2 and pH 5 ribonuclease is 

 rather heat-stable. Only about 20% of the activity is lost when ribonuclease 

 is kept under these conditions at a temperature of 100° for thirty minutes. 



Activators, Inhibitors. Ribonuclease I requires for its optimal activity an 

 ionic strength of about 0.1. The activating effects of various univalent cat- 

 ions are quantitatively similar and larger than those observed with mag- 

 nesium ions.^^ '"^^ Zittle observed that contamination of ribonucleates with 

 copper ions impaired their susceptibility to ribonuclease action.^^' ■** Sodium 

 fluoride even at 0.1 M concentration is without appreciable effect on 

 ribonuclease. Zollner and Fellig^^ showed good evidence for a specific 

 competitive inhibition of ribonuclease activity by heparin. Some other 

 acidic polysaccharides were without effect. Massart et aU^ reported an 

 inhibitory influence of penicillin on ribonuclease on the basis of a histochem- 

 ical assay of enzyme activity. 



Ledoux"^ studied the influence of the sulfhydryl reagent p-chloromercuri- 

 benzoate. He concluded that the complex influence of this substance 

 resulted on the one hand from the inhibitory efi'ect of its combination with 

 the sulfur groups of the enzyme, on the other hand, from the activating 

 influence of its combination with polar groups of the substrate. (See also *^^) 



Ribonuclease as a Protein. The crystaflization of ribonuclease was 

 achieved by Kunitz^ in 1940 by fractionation with ammonium sulfate. 

 The enzyme prepared according to this procedure retains traces of con- 

 taminating proteolytic enzymes even after several recrystallizations.^^ 

 Since crystalline ribonuclease is frequently applied as a histochemical 

 reagent, an important advance was made when McDonald succeeded in 

 removing these contaminations by introducing a heating step into the pro- 



«=» S. R. Dickman, R. B. Knopf, and J. B. Aboskar, Abstr. 126th Meeting Am. Chem. 



Soc, New York p. 71C (1954). 

 ■*« The effects of copper ions can be abolished by versene."^ 

 " N. Zollner and J. Fellig, Am. J. Physiol. 173, 223 (1953). 

 46 L. Massart, G. Peters, and A. Vanhauke, Experientia 3, 494 (1947). 

 4' L. Ledoux, Biochim. et Biophys. Acta 11, 517 (1953). 

 « M. R. McDonald, J. Gen. Physiol. 32, 33 (1948). 



