582 G. SCHMIDT 



Optimal activation by magnesium ions was observed at a molar proportion 

 of 3 between the magnesium ions and the total phosphorus of the sub- 

 strate. Increases of the total salt concentration of the incubation mixture 

 beyond 0.02 M are progressively inhibitory. 



The observations just reported suggest that the salt activation of deoxy- 

 ribonuclease is produced by the combination of the ions with the substrate 

 and not by the formation of an enzyme-activator compound.'"'^ '^"^ 



General Stability and Influence of Heat. Solutions of deoxyribonuclease 

 of relatively high concentration (0.1 mg. of enzyme protein per ml. of 

 buffer solutions) retain their activities for several days at 5°. Highly dilute 

 solutions, however, require the presence of stabilizing colloids (gelatin, 

 peptones) for preservation. 



When solutions of deoxyribonuclease I are kept at 90° for 5 minutes at 

 pH 2.8, the activity disappears completely, but most of the original activity 

 is restored by standing at 20° for 30 hours. 



Inhibitors. (1) Substances removing activating bivalent cations. The re- 

 quirement of certain bivalent cations for the activity of deoxyribonuclease 

 I explains the inhibitory effect of certain anions such as citrate and fluoride 

 on this enzyme. 



(2) Other inhibitors. Copper, selenite, and arsenate ions are strongly 

 inhibitory at 0.001 M, borate at 0.02 M. Hydroxylamine, cysteine, and 

 iodoacetic acid have no appreciable effects. Mitotic poisons, such as col- 

 chicine and aminopterine, and SH-reactants, such as iodoacetamide and 

 chloroacetophenone, caused no inhibition of the action of deoxyribonuclease 

 I on deoxyribonucleohistone. Incubation of nucleohistone with nitrogen 

 mustards resulted in a diminished susceptibility of the nucleoprotein 

 toward deoxyribonuclease. 



(3) Specific inhibitors of deoxyribonuclease in tissues. Tissue inhibitors of 

 deoxyribonucleases are of great biological interest because of their possible 

 regulatory function in the DNA metabolism of growing cells. ^"^ Recently, 

 protein inhibitors of deoxyribonuclease I were found in several tissues. 

 Dabrowska, Cooper, and Laskowski purified such a protein from growing 

 pigeon crop glands."" On the basis of kinetic studies, Laskowski and his 

 associates concluded that this effect was caused by a reversible association 

 of a specific protein with deoxyribonuclease I. The inhibitor protein is easily 

 extractable by water from the tissue, and is precipitated by ammonium 

 sulfate between 0.3 and 0.6 saturation. It is destroyed by heat and by the 

 action of trypsin. Protein fractions with similar inhibitory effects on 



107 N. Weissman and J. Fisher, /. Biol. Chem. 178, 1007 (1949). 



108 V. S. Shapot, Biokhimiya 17, 290 (1952). 



109 S. Zamenhof and E. Chargaff, J. Biol. Chem. 180, 727 (1949). 



110 W. Dabrowska, E. J. Cooper, and M. Laskowski, /. Biol. Chem. VII, 991 (1949). 



