57 



The inhibitory potential of the ADH-BSA conjugate 

 for calf thymus RNA polymerase II was quantitatively com- 

 pared to that of free a-amanitin or ADH by determining the 



apparent inhibition constant (K ) for each compound. The 



-9 

 value of 1.8 x 10 M obtained for a-amanitin agrees with 



published values (Chochet-Meilhac and Chambon , 1974) and 



does not differ significantly from that for ADH (K =3.0 x 



-9 .... ... 



10 M) . Both inhibitions were noncompetitive with respect 



to UTP. ADH-BSA inhibition deviated from the strictly non- 

 competitive inhibition seen for a-amanitin and ADH with an 



average of the x-intercepts giving an apparent inhibition 



-9 

 constant of 69 x 10 M. Conjugation to BSA resulted in a 



38-fold decrease in the affinity of bound a-amanitin for 

 calf thymus polymerase. 

 Interaction with Cells 



The toxicity of conjugated ADH-BSA was examined in 

 three cultured cell lines, CHO M7 , AV3 and EL4. Table 1 

 presents the effects of continuous 48 hour exposure to ADH- 

 BSA or free a-amanitin on cellular proliferation and H- 

 thymidine incorporation of CHO M-7 cells. Comparing equiva- 

 lent molar concentrations of free or conjugated a-amanitin, 

 conjugated a-amanitin was found to be slightly less effective 



an inhibitor of cell growth than free a-amanitin for CHO 



. . 3 

 cells. Greater inhibition is seen when examining H-TdR 



incorporation, with conjugated a-amanitin inducing 60% of 

 the inhibition obtained at 1 x 10 M concentration in com- 

 parison to free a-amanitin. 



