Early in sporulation, the mother cell compartment of Bacillus subtilis transcribes the mother cell metabolic gene ( mmg) operon. The gene mmgA was assigned by other workers using sequence homology as an acetyl- CoA acetyltransferase [ E. C. 2.3.1.9]. The gene was overexpressed in Escherichia coli, and the protein was purified by Ni2+- affinity chromatography. However, the expected MmgA-catalyzed biosynthesis of acetoacetyl- CoA from acetyl- CoA was undetectable by a standard UV assay, HPLC, and mass spectrometry. These methods indicated a preference for the reverse degradative thiolytic reaction, with a k(cat) of 80 s(-1), and a Km of 70 and 50 mu M for CoA and acetoacetyl-CoA, respectively.