14 



quantitative and qualitative variations in cell membrane 

 receptors for the lectin. Con A would also provide a means 

 for assessing the effects of conjugation on the biological 

 activities of the native lectin. 



Rationale 



Alpha-amanitin presents several distinct advantages 

 over most inhibitors used for investigations of drug tar- 

 geting. The affinity of the interaction of a-amanitin with 

 eucaryotic RNA polymerase II make it an extremely potent 

 inhibitor. Moreover, a-amanitin is of low molecular weight 

 and after modification can be coupled via a single reactive 

 site to proteins. This will allow for reproducible produc- 

 tion of conjugates that have minimal structural interference 

 by the amanitin moiety. Work by other investigators has 

 clearly demonstrated that amanitin conjugates retain inhi- 

 bitory potential for RNA polymerase II. The fact that a- 



amanitin inhibits an enzyme present in limited quantities, 



4 

 approximately 10 RNA polymerase II molecules per cell 



(Cochet-Meilhac et al., 1974), that is critical to survival 

 of the cell will allow relatively low doses of inhibitor to 

 generate a measureable cytotoxic effect. Radioactively la- 

 beled derivatives of a-amanitin may be synthesized and can be 

 used to quantitate the coupling to protein as well as the 

 binding to specific cells. Therefore, the initial phase of 

 this investigation consists of developing procedures neces- 

 sary for the synthesis of a-amanitin conjugates of defined 



