184 



minutes, and the third was used unheated 

 as control. A partially purified prepara- 

 tion of transforming material which had 

 previously been dried in the lyophile ap- 

 paratus was dissolved in saline in a con- 

 centration of 3.7 mg. per cc. 1.0 cc. of this 

 solution was mixed with 0.5 cc. of the 

 various samples of heated and unheated 

 diluted sera, and the mixtures at pH 7.4 

 were incubated at 37°C. for 2 hours. 

 After the serum had been allowed to act 

 on the transforming material for this 

 period, all tubes were heated at 65 °C. for 

 30 minutes to stop enzymatic action. 

 Serial dilutions were then made in sahne 

 and tested in triplicate for transforming 

 activity according to the procedure de- 

 scribed under Method of titration. The 

 results given in Table III illustrate the 

 differential heat inactivation of the en- 

 zymes in dog and rabbit serum which de- 

 stroy the transforming principle. 



From the data presented in Table III 

 it is evident that both dog and rabbit 

 serum in the unheated state are capable 

 of completely destroying transforming 

 activity. On the other hand, when 

 samples of dog serum which have been 

 heated either at 60°C. or at 65 °C. for 

 30 minutes are used, there is no loss of 

 transforming activity. Thus, in this 

 species the serum enzyme responsible 

 for destruction of the transforming 

 principle is completely inactivated at 

 60°C. In contrast to these results, ex- 

 posure to 65 °C. for 30 minutes was 

 required for complete destruction of 

 the corresponding enzyme in rabbit 

 serum. 



The same samples of dog and rabbit 

 serum used in the preceding experi- 

 ment were also tested for their de- 

 polymerase activity on a preparation 

 of sodium desoxyribonucleate isolated 

 by Mirsky from shad sperm. 



A highly viscous solution of the nu- 

 cleate in distilled water in a concentra- 

 tion of 1 mg. per cc. was used. 1.0 cc. 

 amounts of heated and unheated sera 

 diluted in saline as shown in the preced- 



AVERY, MACLEOD, MCCARTY 



ing protocol were mixed in Ostwald vis- 

 cosimeters with 4.0 cc. of the aqueous 

 solution of the nucleate. Determinations 

 of viscosity were made immediately and 

 at intervals over a period of 24 hours dur- 

 ing incubation at 37°C. 



The results of this experiment are 

 graphically presented in Chart 1. In 

 the case of unheated serum of both 

 dog and rabbit, the viscosity fell to 

 that of water in 5 to 7 hours. Dog se- 

 rum heated at 60°C. for 30 minutes 

 brought about no significant reduction 

 in viscosity after 22 hours. On the 



Differential Heat Inactivation 



of Desoxyribonucleodepolymerase 



of Dog and Rabbit 5er>um 



Dog serum 



Heated 60° for 30' 

 Heated 65° for 30' 



Unheated 



Hre. 



iO 15 



Time 



20 



25 



Chart 1 



other hand, heating rabbit serum at 

 60°C. merely reduced the rate of de- 

 polymerase action, and after 24 hours 

 the viscosity was brought to the same 

 level as with the unheated serum. 

 Heating at 65 °C., however, com- 

 pletely destroyed the rabbit serum 

 depolymerase. 



Thus, in the case of dog and rabbit 

 sera there is a striking parallelism be- 

 tween the temperature of inactivation 

 of the depolymerase and that of the 

 enzyme which destroys the activity of 



