A region C-terminal to the proline-rich core of p47phox regulates activation of the phagocyte NADPH oxidase by interacting with the C-terminal SH3 domain of p67phox

Activation of the phagocyte NADPH oxidase requires the regulatory proteins p47phox and p67phox, each harboring two SH3 domains. p67phox interacts with p47phox via simultaneous binding of the p67phox C-terminal SH3 domain to both the proline-rich region (PRR) of amino acid residues 360-369 and its C-terminally flanking region of p47phox; the role of the interaction in oxidase regulation has not been fully understood. Here we show that the p47phox-p67phox interaction is disrupted not only by deletion of the PRR but also by substitution for basic residues in the extra-PRR (K383E/K385E). The substitution impaired oxidase activation partially in vitro and much more profoundly in vivo, indicating the significance of the p47phox extra-PRR. Replacement of Ser-379 in the extra-PRR, a residue known to undergo phosphorylation in stimulated cells, by aspartate attenuates the interaction and thus results in a defective superoxide production, suggesting that phosphorylation of Ser-379 is involved in oxidase regulation.