The mechanism of activation of NADPH oxidase in the cell-free system: the activation process is primarily catalytic and not through the formation of a stoichiometric complex

It is commonly assumed that activation of the superoxide-generating NADPH oxidase requires the formation of a stable complex between flavocytochrome b(-245) (the gp91(phox)/p22(phox) heterodimer) and the cytosolic cofactors p47(phox), p67(phox) and Rac2. This association is thought to convert flavocytochrome b(-245), which contains the NADPH-binding site, flavin and haem centres, from an inactive into an active state. Here we provide evidence that, in the cell-free system, this activation process does not necessarily require the formation of a stable stoichiometric complex between the phox proteins. To explain this data we propose the hypothesis that p67(phox) (and possibly Rac2), are capable of activating flavocytochrome b(-245) in a catalytic fashion, where a single molecule of p67(phox) (or Rac2) is capable of activating multiple flavocytochrome b(-245) molecules.