Mapping the domains of interaction of p40phox with both p47phox and p67phox of the neutrophil oxidase complex using the two-hybrid system.

Abstract The superoxide-generating NADPH oxidase complex in phagocytic cells is constituted of a heterodimeric flavocytochrome b and cytosolic factors, p67, p47 and p40 as well as a small G protein Rac (for review, see (1, 2, 3)). A truncated form of the p40 cDNA was isolated by a two hybrid screen of a B lymphocyte library using a full length clone of p47 as target. This truncated form of p40 consisting of the Src Homology 3 (SH3) domain to the 3′ stop codon was also shown to interact with p67 in the same system. A library of smaller fragments of the truncated p40 cDNA was constructed and screened against either p47 or p67. Results show that the SH3 domain of p40 is sufficient for interaction with p47, whereas the C terminus of p40 but not its SH3 domain is involved in the interaction with p67.