140 

 ADGG-Con A Conjugates 



Final verification of the procedures for EDC coupling 

 to Con A was obtained by reaction of a radioactively labeled 



derivative of a-amanitin, H-DM-ADGG, with Con A under the 



3 

 phosphate buffer, pH 5 condition. Since the H-DM-ADGG had 



8 3 



a specific activity of 7.8 x 10 dpm/ymole, the H-DM-ADGG- 



Con A conjugates contained sufficient activity to be used in 

 cell binding experiments. These conjugates with molar ratios 

 of amanitin to Con A of 1.1 were 78% stable to hydroxyla- 

 mine and compared favorably with the similar values for HA- 

 Con A conjugates of 65-88% stability to hydroxy lamine. They 

 absorbed to Sephadex G-75 and eluted with . 1M D-glucose 

 indicative of retention of saccharide binding affinity by 

 the conjugate. 



Reaction of H-DM-ADGG with other lectins possessing 

 varying ligand binding specificities was performed to evalu- 

 ate the effects of ADGG conjugation on hemagglutination 



activity. The fact that all of the lectins tested bound 



3 3 



significant amounts of H-DM-ADGG (molar ratios H-DM-ADGG: 



Con A greater the 1.0) and with two exceptions (SBA and PHA) 

 retained hemagglutinating capability equal to the native 

 lectin, clearly indicates the feasibility of binding a- 

 amanitin to proteins with retention of the original biolo- 

 gical activity of the protein. 



Inhibition of calf thymus RNA polymerase II by an 

 ADGG-Con A conjugate containing 3.6 moles of a-amanitin per 

 mole of Con A yielded a K value for the ADGG-Con A of 



