The Domain Organization of p67phox, a Protein Required for Activation of the Superoxide-Producing NADPH Oxidase in Phagocytes

The phagocyte NADPH oxidase, crucial for innate immunity, is dormant in resting cells, but becomes activated during phagocytosis to produce superoxide, a precursor of microbicidal oxidants. In activation of the oxidase, the multidomain protein p67 phox plays a central role: it translocates to the membrane as a ternary complex with p47 phox and p40 phox , and interacts with the small GTPase Rac to assemble with the membrane-integrated catalytic protein gp91 phox , leading to superoxide production. Here we show, using small-angle X-ray scattering (SAXS) analysis, that p67 phox adopts an elongated conformation when it exists not only as a monomer but also as the heterotrimer. Although p67 phox harbors an N-terminal TPR domain for binding to Rac and a p40 phox -interacting PB1 domain, followed by an SH3 domain that associates with p47 phox , the present model suggests that no or few apparent associations occur between the domains. The positions of the protein-interaction domains in p67 phox contribute to activation of the phagocyte NADPH oxidase: the first SH3 domain that is located between the TPR and PB1 domains positively regulates oxidase activation only when it is present at the correct position; the PB1 domain placed at this SH3 domain position inhibits the oxidase by interacting with p40 phox .