Chronic granulomatous disease is an inherited disorder characterized by the failure of phagocytic cells to produce superoxide upon the ingestion of microorganisms due to a lesion in a membrane-associated NADPH-oxidase. The components of the oxidase have been incompletely characterized by standard biochemical approaches. A genetic strategy has recently led to the identification of the gene affected in the common X-linked form of CGD without reference to its protein product. The X-CGD gene, assigned to chromosome position Xp21.1, encodes a phagocyte-specific RNA transcript that is mutated in patients with X-CGD. Antisera directed toward the predicted protein product of the X-CGD gene recognize a 90 kD membrane glycoprotein, which corresponds to the larger subunit of the phagocyte b-cytochrome heterodimer. The recent genetic and biochemical findings provide an explanation for the consistent absence of the b-cytochrome spectrum in X-CGD, and establish this cytochrome as an essential component of the phagocyte oxidase. The primary amino acid sequence of both the 90 kD b-cytochrome subunit and the 22 kD subunit (cloned as the cDNA using a specific antisera) have no significant similarity to other proteins, including previously studied cytochromes. As both subunits of the b-cytochrome heterodimer are absent in X-CGD, despite a genetic deficiency of only the larger polypeptide, a close interaction between the two subunits may be important for b-cytochrome stability and function. Expression of the b-cytochrome large subunit mRNA is increased by interferon-gamma, an important macrophage activator. Partial or complete restoration of oxidase activity in some X-CGD patients treated with interferon-gamma suggests new therapeutic approaches in the management of this disorder. Molecular reagents prepared from the cloned X-CGD cDNA or gene may prove to be clinically useful in prenatal diagnosis and may provide a basis for somatic gene therapy in future.