Reactive oxygen species (ROS) are conventionally regarded as inevitable deleterious by-products in aerobic metabolism with a few exceptions such as their significant role in host defense. The phagocyte NADPH oxidase, dormant in resting cells, becomes activated during phagocytosis to deliberately produce superoxide, a precursor of other microbicidal ROS, thereby playing a crucial role in killing pathogens. The catalytic center of this oxidase is the membrane-integrated protein gp91^phox, tightly complexed with p22^phox, and its activation requires the association with p47^phox, p67^phox, and the small GTPase Rac, which normally reside in the cytoplasm. Since recent discovery of non-phagocytic gp91^phox-related enzymes of the NAD(P)H oxidase (Nox) family—seven homologues identified in humans—deliberate ROS production has been increasingly recognized as important components of various cellular events. Here, we describe a current view on the molecular composition and post-translational regulation of Nox-family oxidases in animals.