CLEAVAGE OF CCK-33 BY RECOMBINANT PC2 IN-VITRO

The specificity and kinetic properties of CCK 33 cleavage by recombina nt prohormone convertase 2 (PC2) was investigated using a combination of Sephadex G-50 chromatography, HLPC and RIA methods. It is shown tha t CCK 33 can be cleaved effectively by PC2 to form CCK 8, the reaction of which has a K-m of 104.8 mu M. No CCK 22 or other products were de tected and the reaction is completely inhibited by the PC2 inhibitor, p-CMS (p-chloromercuriphenylsulfonic acid), suggesting that the cleava ge is PC2 specific, The time course of this reaction shows an initial lag phase followed by a rapid increase in velocity consistent with a p reviously reported spontaneous transformation from a 71 kDa relatively inactive form into a more active form of 62 kDa (1). PC2 did not clea ve recombinant pro CCK or CCK 1-21. These results demonstrate that PC2 , an enzyme usually associated with dibasic cleavages, can cleave easi ly at single basic residues. (C) 1997 Academic Press.