Binding of radioactive α-difluoromethylorithine to rat liver ornithine decarboxylase

Inactivation of rat liver ornithine decarboxylase by incubation with [5-¹⁴C]-α-difluoromethylornithine resulted in the covalent binding of radio-activity to the enzyme. The extent of binding correlated with the degree of inactivation and with the amount of enzyme present. The labeled protein eluted as a single peak which coincided exactly with the active enzyme when chromatographed on Sephadex G-200 and ran as a single band on polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate at a position corresponding to a M.W. of about 55,000. The stoichiometric binding of [5-¹⁴C]-α-difluoromethylornithine therefore provides a convenient method for quantitating ornithine decarboxylase protein and for determining the purity of preparations of the enzyme. Assuming that 1 molecule of the drug is needed to inactivate each sub-unit, it was calculated that after stimulation with thioacetamide ornithine decarboxylase represents about 0.00014% of the liver soluble protein.