ENZYMES ATTACKING NUCLEIC ACIDS 559 



nuclease of pancreas was reawakened by the observation of Dubos and 

 his associates,^ '^ who found that the Gram-positive staining properties of 

 killed pneumococci disappeared after incubation of the cells with boiled 

 pancreas extracts. They succeeded in a considerable purification of Jones' 

 enzyme for which they proposed the term "ribonuclease,"'and they found 

 evidence supporting the assumption that the ribonucleic acid fraction of 

 pneumococci was related to their Gram-positive properties. The work of 

 these authors stimulated P. A. Levene** to undertake a study of pancreas 

 ribonuclease in collaboration with Schmidt. These authors confirmed 

 Jones' concept that the action of ribonuclease was limited to the cleavage 

 of the interlinkages between the nucleotide groups of ribonucleic acid, but 

 they emphasized that only part of these interlinkages could be split by 

 the enzyme. 



A new phase in the investigation of ribonucleases began in 1940 when 

 Kunitz^ obtained the enzyme in crystalline form (Fig. 1) and found that 

 its action was accompanied by the liberation of acidic groups. It will be 

 seen in the following sections and in other chapters of this book that this 

 achievement not only opened the way for a clearer understanding of the 

 properties of Jones' enzyme, but that it also created a valuable tool for 

 the study of the structure of its substrates, the ribonucleic acids. 



Specificity. Ribonuclease I is a highly specific phosphodiesterase which 

 hydrolyzes all known ribonucleic acids, certain ribonucleotides, ^° and syn- 

 thetic ribonucleotide-P-esters" which will be defined in the following 

 sections (see also Chapter 12), and a natural polymer of ribosephosphoric 

 acid which was discovered by Zamenhof et al}- (Fig. 2). It also hydrolyzes 

 polynucleotides obtained by deamination of PNA uith nitrous acid.^*'^'' 

 On the other hand, ribonuclease is entirely inactive toward deoxyribo- 

 nucleic acid and its hydrolysis products,^ except apurinic acid ("thymic 

 acid"),^^ and toward the P-esters of a-glycerophosphoric acid such as 

 L-a-glycerylphosphorylcholine and L-a-glycerylphosphorylethanolamine.'^ 

 Diphenyl phosphate and dinitrophenyl phosphate, which are used as 



' R. H. S. Thompson and R. Dubos, J. Biol. Chem. 125, 65 (1938). 



« G. Schmidt and P. A. Levene, J. Biol. Chem. 126, 423 (1938). 



9 M. Kunitz, /. Gen. Physiol. 24, 15 (1940). 



10 D. M. Brown, D. I. McGrath, and A. R. Todd, J. Chem. Soc. 1952, 2708. 

 " D. M. Brown, E. A. Dekker, and A. R. Todd, J. Chem. Soc. 1952, 2715. 

 '2 S. Zamenhof, G. Leidy, P. L. FitzGerald, H. E. Alexander, and E. ChargafT, J. 



Biol. Chem. 203, 695 (1953). 

 '3 W. E. Fletcher, J. M. Gulland, D. O. Jordan, and H. E. Dibben, J. Chem. Soc. 



1944, 30. 

 '■* L. Vandendriessche, Compt. rend. trav. lab. Carlsberg, Ser. chim. 27, 341 (1951). 

 '6 M. C. Durand and R. Thomas, Biochim. et Biophys. Acta 12, 416 (1953). 



