Serine phosphoric acid from diisopropylphosphoryl derivative of eel cholinesterase.

Diisopropyl fluorophosphate (DFP) is a highly specific inactivator of e&erases, such as cholinesterase and chymotrypsin (2), and combines irreversibly with these enzymes (3,4). In a previous report from this laboratory (5) we have shown that, when the reaction product of DFP and chymotrypsin, namely, diisopropylphosphoryl chymotrypsin, is partially hydrolyzed, serine phosphoric acid may be obtained in a 30 per cent yield (based on phosphorus) from the hydrolysate. It was of interest to see whether or not the reaction between DFP and choline&erase yielded the same product after partial hydrolysis. Eel cholinesterase preparations, even though highly active enzymatically, ordinarily contain considerable inert protein. Since the reaction between DFP and such preparations can be prevented by the addition of an excess of acetylcholine (3), it is reasonable to assume that the DFP-binding property is associated with the cholinesterase present and not with concomitant, impurities. This reasoning justified the use of these preparations in an effort to determine the nature of the combination between cholinesterase and DFP. The name “diisopropylphosphoryl cholinesterase” (DPChE) has been given to the DFP derivative of cholinesterase. We have prepared DPChE containing P32, partially hydrolyzed the product, fractionated the hydrolysate on a cation exchange resin, and obtained serine phosphoric acid in a yield of approximately 40 per cent based on phosphorus.