STRUCTURE-FUNCTION STUDIES ON THE BIOSYNTHESIS AND BIOACTIVITY OF THE PRECURSOR CONVERTASE PC2 AND THE FORMATION OF THE PC2/7B2 COMPLEX

Site directed mutagenesis of the prohormone convertase PC2 was used to define the effect of certain residues on the zymogen activation of proPC2 and on its binding to the neuroendocrine protein 7B2, These included the oxyanion hole Asp(309) (D309N), the N-terminal Glu(25) (E25Q and E25K) of proPC2 and the Asp(519) (D519E) of the RGD motif within the P-domain of PC2, Heterologous vaccinia virus expression of the wild type and mutant PC2's in endocrine pituitary cells such as AtT20 and GH3 cells demonstrated that the most dramatic effect was observed with the D309N mutant which no longer bound pro7B2 and which exhibited a significant reduction in its capacity to produce beta-endorphin from pro-opiomelanocortin (POMC).