ENZYMES ATTACKING NUCI EIC ACIDS 581 



of polymerization of DXA; its liberation (as measured by the appearance 

 of methylene blue in the supernatant solution after the precipitation of 

 the dye-polynucleotide compounds with acid) begins only in the later 

 phases of deoxyribonuclease action. The characteristic initial drop of 

 viscosity is therefore not accompanied by the liberation of methylene 

 blue. This interpretation is in analogy with the observations of Jungner 

 et al.,^^'^'' who found that the dielectric constants of deoxyribonuclease 

 digests remained unchanged during phases of rapid decreases of viscosity. 



Most methods of assay yield satisfactory results with purified deoxyribo- 

 nuclease preparations but are not easily adaptable to enzyme assays in 

 crude tissue extracts or serum. Kurnick's procedure, however, is applicable 

 to serum. It should be mentioned that Alfert reported evidence suggesting 

 that changes of methyl green binding by DXA can be caused by factors 

 other than depolymerization.^* 



Kinetics. The kinetics of the degradation of DXA by deoxyribonuclease 

 I is obviously an overall picture of many different individual enzyme 

 processes. Xo simple substrates for deoxyribonuclease are available as yet. 



At low concentrations of DXA (160 )ug. per ml.) the hydrolysis curve 

 follows the course of a pseudounimolecular reaction, and the velocity 

 constants are proportional to the concentrations of deoxyribonuclease in 

 the digestion mixture.^^ At high concentrations of the substrate, the hydrol- 

 ysis curve is an asymmetric S-shaped curve owing to a short initial lag 

 period. ^^ 



Activators. Unlike ribonuclease I, deoxyribonuclease recjuires for its 

 action the presence of bivalent cations.^'' •^^•^^•^^•^°°-'''^ Magnesium, man- 

 ganese, ferrous, and cobaltous ions are the most effective activators and 

 resemble each other in regard to the quantitative intensity of activation. 

 The observations regarding the effects of some other cations, such as zinc 

 and calcium, are contradictory.^"^ The concentration rerjuired for optimal 

 effects of the activating cations is practically independent of the enzyme 

 concentration but increases with increasing substrate concentrations. 



9« G. Jungner, I. Jungner, and L. G. Allg^n, Kahire 163, 849 (1949). 

 "G. Jungner, Ada Physiol. Scand. 10, Suppl. 32, 1 (1945). 

 98 M. Alfert, Biol. Bull. 103, 145 (1952). 

 "9 J. Gr^goire, Compt. rend. 231, 384 (1950). 



100 F. G. Fischer, H. Lehmann-Echternacht, and I. Bottger, J. prakt. Chem. 159, 59 

 (1941). 



101 M. Laskowski and M. K. Seidel, Arch. Biochem. 7, 465 (1945). 



io» C. E. Carter and J. P. Greenstein, J. Natl. Cancer Inst. 7, 29 (1946). 



1" J. P. Greenstein, C. E. Carter and H. W. Chalkley, Cold Spring Harbor Symposia 



Quant. Biol. 12, 64 (1947). 

 10^ W. G. Overend and M. Webb, Research 2, 99 (1949). 



lo^ T. Miyaji and J. P. Greenstein, Arch. Biochem. and Biophys. 32, 414 (1951). 

 »<" L. M. Gilbert, W. G. Overend, and M. Webb, Exptl. Cell Research 2, 349 (1951). 



