AN SH3 DOMAIN AND PROLINE-RICH SEQUENCE MEDIATE AN INTERACTION BETWEEN 2 COMPONENTS OF THE PHAGOCYTE NADPH OXIDASE COMPLEX

Neutrophils possess a multicomponent NADPH oxidase system capable of p roducing large quantities of superoxide in a process known as the resp iratory burst (1). Upon stimulation of a phagocytic cell, two cytosoli c components of the oxidase, p67(phox) and p47(phax), associate with a membrane-bound flavocytochrome b and a small GTP-binding protein to f orm a functional enzyme complex. Each of the Phox proteins contains tw o src homology 3 (SH3) domains, which are of unknown function but are potential mediators of protein-protein interactions between components of the activated oxidase. We have isolated a 47-kDa protein from lysa tes of differentiated HL60 cells that specifically bound to the carbox yl-terminal SH3 domain of p67(phox) and not to any other SH3 domain te sted. This protein was identified as p47(phox), and the putative SH3 d omain binding site was located to a carboxyl terminal proline-rich reg ion. Proline-rich synthetic peptides based on this carboxyl-terminal r egion specifically inhibited the binding of p47(phox) to the carboxyl- terminal SH3 domain of p67(phox), and sequential truncation defined a unique minimal sequence, which, although similar, does not match the c onsensus sequence defined for other SH3-binding proteins.