ENZYMES DORMANT IN THE INTACT SPORE 109 



Table V 



Effect of p-fluorophenylalanine on the enzymatic activity of 



heat shocked spores of B. terminalis 



Time of addition of 



p-fluorophenylalanine Endogenous Exogenous 



Before heat shock 8 



Immediately after heat shock 4 61 



60 mintues after heat shock 4 54 



Strength of spore suspension — 15 mg dry wt/ml 



Duration of heat shock — 5 hrs. at 65°C. Substrate — Glucose 0.5 mg/ml 

 Nutrients — 26 jug/ml L-alanine; 6 ;Ltg/ml adenosine 

 p-fluorophenylalanine— 1.0 x lO-M fi 1 oxygen uptake in 30 minutes 



heated 1 hour at 65° to inactivate the active glucose-oxidizing enzymes, and 

 equilibrated at 30° for 40-45 min., there was no significant decrease in the 

 activity. Thus, it appeared that the glucose-oxidizing enzymes were stable to 

 heat, but this was not the case when a more concentrated suspension (15 

 mg/ml) was used. During storage in the frozen state for almost a year, the 

 spores have undergone some changes that obviate the need for alanine in the 

 activation of the enzymes. When a dense suspension (15 mg/ml) of such 

 aged spores, activated by heat alone or by heat in the presence of L-alanine, 

 was heated again 1 hour at 65°, the activity was completely lost. However, 

 the activity could be regained by adding more alanine, before the second 

 heat shock, although in the case of the suspension activated by heat alone 

 the first time, a much longer second heat shock was required (Table VI). 



Table VI 



Effect of duration of heating, strength of spore suspension, and time 



of addition of nutrients on the enzymatic activity of heat activated 



spores of B. terminalis 



Duration Time of addition of nutrients and spore concentration 



of second No Before Before 2nd Before each 



heat shock Time 1st heat shock only heat shock only heat shock 



65 °C 15 mg/ml 6 mg/ml 15 mg/ml 15 mg/ml 15 mg/ml 



Duration of first heat shock — 5 hrs. at 65 °C 



Nutrients — 26 /xg/ml L-alanine: 6 /Ag/ml adenosine 



Substrate — Glucose 0.5 mg/ml fi 1 oxygen uptake in 30 minutes 



