Abstract The fluxes of radioactive pantothenate between the cellular components 4'-phosphopantetheine, coenzyme A, and acyl carrier protein (ACP) and the medium have been measured in a pantothenate auxotroph of Escherichia coli. Radioactive fluxes generated with two alternate computer models for pantothenate metabolism were compared with the experimental data. One model was consistent with all the data with the same set of rate constants; the other model was not. The successful model has CoA as the immediate precursor of ACP and features rapid turnover of the 4'-phosphopantetheine prosthetic groups of ACP and of CoA in exponentially growing cells; such turnover was directly observable under certain conditions. The rate of ACP turnover was four times the rate of growth of the ACP pool.