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SH3 domain-mediated interactions involving the phox components of the NADPH oxidase
Objective And Design:
To derive a model describing the SH3 domain-mediated assembly of the activated NADPH oxidase.
Materials:
Recombinant SH3 domain and Pro-rich fusion proteins were used to investigate potential co-associations.
Methods:
Interactions were assessed using biotinylated overlay assays and the yeast two hybrid system. Association with p47phox from cell lysates was examined by immunoblot analysis.
Results:
The association between p47- and p22phox involves the SH3 domains of p47phox functioning in tandem. The Prorich motif in p47phox interacts with both p40phox and the COOH-terminal SH3 domain of p67phox.
Conclusions:
In the resting cell, the Pro-rich motif of p47phox interacts with the SH3 domain of p40phox, which in turn associates with p67phox. Upon activation, the p47-p40phox regulatory complex dissociates, permitting the association of p47phox with the COOH-terminal SH3 domain of p67phox. This complex translocates to the plasma membrane and associates with cytochrome b558, via interaction of the tandem SH3 domains of p47phox with the p22phox Pro-rich motif.