136 



Synthesis of conjugates of ADH and Con A resulted in 

 ADH-Con A conjugates that had a molar ratio of ADH to Con A 

 of 0.67 as determined from the absorption spectra (Figure 10). 

 The conjugates retained the specificity of interaction with 



calf thymus RNA polymerase II but at a greatly reduced 



-9 



affinity (K = 186 x 10 M) . However, conjugation of ADH 



via free protein carboxyl groups had a deleterious effect on 

 the ligand binding ability of the Con A. The failure of the 

 ADH-Con A conjugates to bind to Sephadex G-75 implied a loss 

 of Con A specific ligand binding. Synthesis of ADH-Con A 

 conjugates in the presence of the specific ligand D-glucose 

 (1M) failed to protect the Con A saccharide binding site. 

 Synthesis in the presence of 1M NaCl to provide the optimal 

 ionic strength for native Con A conformation also failed to 

 produce conjugates with ligand specificity. Since it 

 appeared likely that conjugation to carboxyl groups on Con 

 A resulted in a loss of ligand binding activity, this 

 approach was discontinued and derivatives of a-amanitin 

 that couple to free amino groups on proteins were examined. 



ADGG-BSA Conjugates 



The synthesis and chemical characterization of an a- 

 amanitin derivative possessing a free carboxyl group (ADGG) 

 provided an opportunity to produce conjugates whose proper- 

 ties could be directly compared to the ADH-BSA conjugates 

 already characterized. With ADGG-BSA conjugates containing 

 2.0 moles of ADGG per mole of BSA, there resulted a 21-fold 



