Purification and properties of a folate-catabolizing enzyme

We have identified and purified to homogeneity an enzyme from rat liver tha t catalyzes the oxidative catabolism of 5-formyltetrahydrofolate to p-amino benzoylglutamate and a pterin derivative. Purification of the enzyme utiliz ed six column matrices, including a pterin-6-carboxylic acid affinity colum n. Treatment of crude rat liver extracts with EDTA or heat decreased the sp ecific activity of the enzyme by up to 85%. Peptides generated from-the pur ified protein were sequenced and found to be identical to primary sequences present within rat light chain or heavy chain ferritin. Commercial rat fer ritin did not display catabolic activity, but activity could be acquired wi th iron loading. The purified enzyme contained 2000 atoms of iron/ferritin 24-mer and displayed similar electrophoretic properties as commercial rat l iver ferritin. The ferritin-catalyzed reaction displayed burst kinetics, an d the enzyme catalyzed only a single turnover In vitro. Expression of rat h eavy chain ferritin cDNA resulted in increased rates of folate turnover in cultured Chinese hamster ovary cells and human mammary carcinoma cells and reduced intracellular folate concentrations in Chinese hamster ovary cells. These results indicate that ferritin catalyzes folate turnover in vitro: a nd in vivo and may be an important factor in regulating intracellular folat e concentrations.