Aspartate transcarbamoylase (ATCase) was purified from Streptomyces griseus. The enzyme is a dodecamer with a molecular mass of approximately 450 kDa. The holoenzyme is st complex of ATCase and active dihydroorotase (DHOase) subunits. The ATCase and DHOase activities co-purify after gel filtration and ion-exchange chromatography. Denaturing gel electrophoresis separates the holoenzyme into a 38-kDa ATCase polypeptide and a 47-kDa DHOase polypeptide. The holoenzyme retained ATCase and DHOase activity after being heated to 65 degrees C for 5 min, but after storage at 4 degrees C for 24 hours lost ATCase activity. Previously, the Pseudomonas putida Class A ATCase was defined by Schurr et al. (J Bacteriol 177, 1751-1759) as requiring an inactive DHOase to be functional. Here, we show that an active DHOase is part of the dodecameric ATCase/DHOase complex in Streptomyces. To distinguish those Class AATCases with active DHOases from those with degenerate DHOases, we suggest the subdivision, Class A(1), for the former and Class A(2) for the latter.